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Genes and deafness
REVIEWS 5 Woychik, R.P. et al. (1990)/~c. NaaAcad. S¢i. USA 87, 2588-2592 6 Bultman, S.J., Russell, L.B., Gutierrez-Espeleta, G.A. and Woychik, R.P. (1991) Proc. Natl Acad. Sci. USA 88, 8062--8066 7 Bultman, S.J., Michaud, E.J. and Woychik, R.P. (1992) Cell 71, 1195-1204 8 MillerM.W. et al. (1993) Genes Dev. 7, 454--467 9 Bultman, S.J. et al. (1994) Genes Dev. 8, 481--490 10 Vrieling,H. et al. (1994) Proc. Natl Acad. Sci. USA 91, 5667-5671 11 Robbins, L.S. et al. (1993) Cell72, 827-834 12 Lu, D. etal. (1994) Nature371, 799-802 13 Copeland, N.G., Hutchison, K.W. and Jenkins, N.A. (1983) Cell 33, 379-387 14 Yen, T.T. et al. (1994) FASFdlJ. 8, 479.-488 15 Michaud, E.J., Bultman, S.J., Stubbs, L.J. and Woychik, R.P. (1993) Genes Dev. 7, 1203-1213 16 Duhl, D.M.J. et al. (1994) Nature Genet. 8, 59.-65 /7 Michaud, E.J. et al. (1994) Genes Dev. 8, 1463-1472 18 Siracusa, L.D. et aL J. Hered. (in press) /9 Duhl, D.M.J. et al. (1994) Development 120, 1695-1708 ~0 Johnson, P.R. and Hirsch, J. (1972)J. LipidRes. 13, 2-11 21 Yen, T.T., Greenberg, M.M.,Yu, P.L. and Pearson, D.V. (1976) Horm. Metab. Res. 8, 159-166 22 Iierberg, L. and Coleman, D. (1977) Metabolism 26, 59-99 23 Friedman,J.M. and Leibel, R.L.(1992) Cell69, 217-220 24 Wolff,G.L., Medina, D. and Umhoitz, R.L. (1979)J. Naa Cancer Inst. 63, 781-785 25 Heston, W.E. and Vlahakis, G. (1968)J. Nail Cancerlnst. 40, 1161-1168
26 Heston, W.E. and Vlahakis, G. (1961)J. Natl Cancerlnst. 27, 1189--1196 27 Heston, W.E. and Vlahakis, G. (1961)J. Natl Cancerlnst. 26, 969-983 28 Wolff, G.L. (1970) CancerRes. 30, 1731-1735 29 Wolff, G.L., Kodell, R.L.and Cameron, A.M. (1982)J. Tox. Env. Health 10, 131-142 30 Wolff, G.L. etal. (1987)Carcinogenesis& 1889-1897 31 Siracusa, L.D. (1991) Ann. N.Y. Acad. Sci. 642, 419-.-430 32 Michaud, EJ. et al. (1994) Proc. Natl Acad. Sci. USA 91, 2562-2566 33 Miller, M.W. et al. (1994) EMBOJ. 13, 1806-1816 34 Papaioannou, V.E. and Mardon, H. (1983) Dev. Genet. 4, 21-29 35 Rinchik, E.M. and Russell, L.B. (1990) in Genome Analysis (Vol. I) (Davies, K. and Tilghman, S., eds), pp. 121-158, Cold Spring Harbor Laboratory Press 36 Siracusa, L.D. et al. (1987) Genetics 117, 93-100 37 Seafle, A.G. (1968) Comparative Genetics of Coat Colour in Mammals, Academic Press 38 Sage, R.D. (1981) in The Mouse in Biomedical Research (Vol. I) (Foster, H.L., Small,J.D. and Fox, J.G., eds), pp. 39-90, Academic Press
L.D. SXRACUSA tS IN THE JWFERSOS CASC~R ISSTITt~, THOMAS JEFI:ERSOJV UNIVERSITY, JDEPARTMENTOF MICRO. BIOLOGY AND IMMUNOLOGY, 233 SOUTH IOTH STREET, PHULAOE~HL¢,PA 19107, USA.
Genes and deafness T h e mammalian inner ear rightly deserves its alternative name, the labyrinth. It is a remarkably complex structure (see Box 1). Endol?mph- and perilymph-filled channels course around the cochlea, the attditory sense organ, and around the sacculus, utriculus and the three semicircular canals, which together form the vestibular part of the inner ear that detects head position and movement and hence aids balance, The sensory epithelia within the inner ear are highly organized arrays of senso W hair cells and supporting cells. The organ of Corti within the cochlear duct is particularly complex and precise in its arrangement of a variety of types of sensory and supporting cells, Given this complexity at the cellular and gross structural levels, it is not surprising that many genes have been found to be involved in controlling the development and function of the inner ear. Indeed, mutations at many different loci in both mice and humans are known to cause hearing impairment1-'1, About one in every 1000 children is born with a significant hearing impairment, and about half of these suffer from the relatively common genetic disorder of hereditatT deafness. Genetic factors are highly likely to play a role in susceptibility to hearing loss later in life, and understanding how single gene mutations cause deafness should help us to unravel the interactions between genetic predisposition and environmental experience that cause late-onset hearing loss. In the British population, 16°,6 of all adults suffer from a significant hearing impairmentS; this proportion is probably similar elsewhere,
KAREN P. STEEL A N D STEPHEN D . M . BROWN
Many d~erent &enosappear to be involved in the development anti, notion of the mammalian innerear. Some of the genes involvedduring early innerear morphogenests have boonIdent~l using mutations or targetted transgenic interruption, while a hanOI of genes involved in pigmentation anomalies associated with hearing impairment have been clone~ Severalgenes involved in syndromtc late.o~et hearing loss have also been ldantO~e~However, the maJori~ of cases of hereditary hearing impalrmentJ~om chiMhoodprobably involve g e n e s expressed in the sensory neurospithelia of the inner ear, and n o n e of the genes or mutations causing this type of de~ess haveyet b m id~t~e~ Here, we review the progress that has boonmade tn~ndinggenes for de~ess and in using mouse mutants to elucidate the biological basts of the bearing deficit. and thus represents a large health and communication problem for both sufferers and their families. Mice are likely to be extremely useful in identifying some of the genes involved in causing deafness, as well as in understanding how those genes and their mutations interfere with the normal process of development and functton of the tuner ear5 • ' • ,9, m. The m any similarities between the mouse and human auditory systems suggest that similar cascades of gene action are likely to be involved in the development of the system in the
TIG DECEMBER1994 VOL. 10 NO. 12
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(a) The human outer, middle and inner ear. The outer ear canal leads to the tympanic membrane (ear drum), and ~und vibrations are transmitted from there to the inner ear via the three middlL• ear ossicles, the malleus, incus and stapes. The stapes inserts into the oval window at the base of the cochlea. Co) One rum of the cochlear duet. During sound stimulation, the organ of Corti vibrates and the sensory hair cells are stimulated by a shearing movement between the top surface of the organ of Cotti and the rectorial membrane. Most afferent neurons innervate inner hair cells (shown here on the left of the black pillar cells), and their cell bodies form the main par of the spiral ganglion, which in mm leads to the cochlear nerve. The three or more row~ .of outer hair cells (to the right of the pillar cells) receive primarily efferent innervation, and are believed to act as motor cells, enhancing the response of the cochlea (see, for example, Sef. 55). The stria vascularis on the lateral wall of the cochlear duct has a key role in generating the high resting endocochlear potential in the endolymph that fills the scala media, and it also controls the high K+ and low Na+ concentration in the endolymph. Scab vestibuli and scala tympani both contain perilymph, a fluid that is much more like a normal extracellular fluid, with low K+and high Na+levels and no high resting potential. The stippled area represents bone. (c) A typical sensory hair cell. The actin-filledstereocilia are arranged in rows of waded heights at thc top of the hair cell. Tip links connect the shorter stereocilia to the adjacent taller stereo:ilia, and it is stretching of the tip links that is
(C)
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Tip links - -
Stereocilium
Haircell
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thought to activate the transduction channels of the hair ceil, leading to depolarIzation and initiationof synaptic activity at the base of the cell (see, for example, Refs 51, 52), two species. Furthermore, there are many parallels in the types of pathology seen in the two species (see Box 2). This review outlines some of the key findings concerning genes that cause deafness and our understanding of how they affect the development and function of the inner ear. The major types of pathology as defined in Box 2 are considered in turn.
Morphosenetic defects The gross structural malformations seen in cases of morphogenetic defects of the inner ear suggest that the development of the labyrinth is disrupted very early in embryogenesis. The malformations may affect the vestibular part of the inner ear or the cochlea or both. Art example in which the vestibular part of the inner ear is primarily affected, the fidget mouse inner ear, is shown in Fig. 1. One interesting feature of these morphogenetic defects is the association of neural tube and inner ear anomalies: minor or transient neural tube defects are frequently detectable at the time that the inner ear is forming, and labyrinthine malformations usually occur in cases of severe neural tube defects 1tJ2. Figure 2
illustrates one example in which neural tube defects in embryos homozygous for the splotcb mutation are associated with malformations of the developing inner ear. It appears that the neural tube has an inductive influence on the development of the labyrinth. The developing neural tube has been extensively studied, and the expression patterns of a number of genes thought to be involved in controlling its development have been described 13. One of the earliest genes to be proposed as a potential inducing agent for inner ear morphogenesis is Int2, which is expressed in the neural tube. Exposure to Int2 antisense oligonucleotides does indeed disrupt otocyst development in vitro t4. Transgenic disruptions of lnt2 and another gene, Hoxa-I (previously known as Hoxl.6), which is known to be expressed in the neural tube opposite the developing labyrinth, have been shown to lead to abnormal morphogenesis of the inner ear 1s-re. In contrast, transgenic disruptions of Krox20 do not affect labyrinthine morphogenesis, despite the fact that this gene is known to be expressed in rhombomere 5, directly adjacent to the otocyst. One consequence of the interaction between the neural tube and the inner ear is that the
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humans homozygous for mutations in the homologofis gene GLI3 has not been reported. In this case, humans heterozygous for GLI3 mutations have Grieg cephalopolysyndactyly syndrome 21 (GCPS), which does not usually involve deafness. Mice homozygous for the b/s mutation have maternal histidinaemia resulting from abnormal histidase activity, which causes transient neural tube anomalies and abnormal formation of the vestibular part of the inner ear in the offspringZZ; it is not clear whether there is any human correlate of these ear defects in maternal hisfidinaemia. Disruption of both Int2 and Hoxa-1 causes abnormalities of the neural tube and inner ear, but, again, it is not known whether mutations in these genes cause deafness in humans. The interaction between development of the neural tube and the inner ear needs further investigation, but at least some of the key players are now known, , ¢oehk, o-samflar defects Cochleo-saccular abnormalities are found in the type of deafness that is often associated with white spotting of the hair or skin in many different mammals. Darwin noted that white cats are often deaf, and the association was probably known long before. White spotting of the coat is believed to be caused by mutations that affect migratory melanocytes, which originate in the neural crest. It has long been known that there genes responsible for morphogenetic defects may not are melanocytes in the inner ear, and that melanocytenecessarily be expressed in the inner ear, but only in like cells form part of the stria vascularis of the cochlea; the neural tube. however, albino mammals hear perfectly well so it is obviously not the pigment-producing ability of the Ident~ed 8enes melanocytes that is important in this respect. It appears Some of the identified genes involved in morpho- that some other feature of melanocsrtes is essential for genetic defects are listed in Table 1. The splotch (St,) normal strial function. mutation in mice causes severe neural tube defects and The stria vascularis in normal mammals is responinner ear malformations when homozygous tl. How- sible for generating the unusually high extracellular restever, humans with mutations in the homologous gene ing potential, the endocochlear potential (EP), and the PAX3, who have Waardenburg syndrome type 1 (WS1) characteristic ion composition of high concentrations of (Ref. 19), are heterozygous for the mutation, and the K÷ and low concentrations of Na + in the endolymph in cause of the hearing impairment in WS1 is not known the lumen of the cochlear duct. In normal mice, the EP (reviewed in Ref. 20). The extra toes mouse mutation is around +100 mV, but in mutant mice with no (Xt) also causes neural tube defects and inner ear melanocytes in their stria vascularis this potential is malformations in homozygotes, but the condition of abolished a3. A causal relationship between the lack of melanocytes and the absence of a normal EP is suggested by the discovery that some mu(a) Crus (b) tations are 'leaky'; that is, some melanoblasts Presumptive do manage to find their way to the stria _ . . .~ commune Sunerior eostenor I ca,.,ol superior canal during developmenta4, 2~. In these ears, a positive EP can always be recorded, although it may not be as high as normal, while a significant EP has never been found in any mice Lateral./~'li"~'~"~-"~-"~'~ I 0 ! without strial melanocytes. Figure 3 illustrates an example of this type of mutant, the viable dominant spotting, mtmse mutant: melanoRO~lodlv//'XJ C )"Utriculus , . ~ Oval cytes have populated the stria of the left cochlea, which has an EP, but not the right cochlea, which has no EP. Asymmetry is com"-"- "Cochlea"- v mon in this type of hearing impairment. The particular property of the melanocyte Ihctm~ 1. An example of a malformed inner ear in a morphogenetic mutant important for striai function is unknown, of the mouse. (a) Outline of the normal mouse inner ear, (b) Inner ear of the but we do know that these cells are refidget mutant, showing severe deformityof the vestibularpart of the inner quired throughout adult life for normal strial ear, including absence of the lateral semicircularcanal. In other mutants, the function rather than being needed only at a cochlea is more severelymalformed. (Modifiedfrom Ref. 10.) critical stage of development. Therefore, the TIG DECEMBER1994 VOL. 10 No. 12 430
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F t ~ 2. Illustration of the relationship between inner ear malformations and neural tube defects. (a) Section through an 11.5 d.p.c. (days post coitum) normal mouse embryo, showing the normal neural tube (NT) and developing otic vesicle(OV). (b) An example of a severely malformed neural tube in a splotchhomozygote embryo, with associated abnormal morphogenesis of the labyrinth. (Modified from Ref. 11.) melanocytes could be involved in some aspect of the heterozygous form in people with piebald trait (a pigion-pumping processes involved in generating the EP. mentation defect causing white skin and hair patches). An EP is necessary for the normal functioning of the If these mutations have effects similar to the W musensory hair cells, and the result of an EP dose to zero tations in mice, it is the homozygotes who would be is a gready raised threshold to sound. Eventually, strial deaf. However, only one putative homozygote (who dysfunction leads to secondary hair-cell degeneration was deaf) has ever been reported and it is not clear whether he carried mutations in K/Tor in another gene and severe to profound hearing impairment. Various alleles of the dominant spotting ( W ) and affecting pigmentation aT. No mutations of the human steel(Sl) loci, which produce white spotting of the coat, homologue of SI (MGF) have been reported. Thus, it have been analysed extensively to investigate the role seems that the homologues of Wand S! in the human of melanocytes in cochlear function, and to establish population do not play a major role in deafness. The that the hearing impairment is mediated by strial light allele of the brown locus, Btt, causes progressive dysfunction resulting from a lack of melanocytes. melanocyte death and premature greying of the coat, Melanoblast migration away from the neural crest begins as normal in Wand SI mutant mice and these precursor cells start to differentiate normally by explesslng melanocyte-specific genes; however, in these mutants there are always fewer melanoblasts than normal and their numbers decrease between 11 and 12-13 days post cottum (Ref. 26; J. Cable et al., unpublished). This decrease in numbers, compared with the rapid prolfferation in numbers seen in control embryos at this stage, suggests that the c-kit growth factor and its iig'and, which are encoded by Wand Sl, respectively, may act as survival factors for mdanoblasts. Since melanoblasts migrate from the neural crest to the inner ear, genes causing cochleo-- Me,tam.3. An example of the (a) leR and (b) right cocl'dea from a single cochleo--saccular viable dominant spotting ( ga/ Wv) mouse mutant. The single coiled saccular defects may not necessarily line represents the turns of the cochlear duct, and the cross through the line shows the be expressed in the inner ear. position from which the endocochlear potential was recorded. Dots represent the distribution of melanocytes in the stria vascularisalong the entire Identified genes length of the cochlea. The left cochlea had a near-normal endocochlear potential Some of the genes involved in of 90 mV and melanocytespresent in the middle half of the length of the cochlear cochleo-saccular defects are listed in duct, while the right cochlea had an endocochlear potential near zero (-2 mY) Table 1 (part b). A number of K/T and no sign of any melanocytesthroughout the cochlea. OW, oval window: mutations have been identified in RW, round window. (Modifiedfrom Ref. 25.) TIG DECEMBER1994 VOL. 10 No. 12
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T.Amm1. Genes invohred in deafness
cae
Wpeofpmmu
Pax-3 PAX3
Paired-box transcription factor Paired-box transcription factor
speam
tecm/O~me
Mouse Human
S p ~ ' ~ Waatdenburg~
Mouse Human
Xt (homozygo¢~ ~ Gli3 Grieg cephalopolysyndactylysyndrome GU3 (GCPS)b (hetemzygotes affected)
Mouse
Ha/
His
Mouse
lnt2tg
F~
Mouse
Hoxl.6 t8
Hoxl.6, Hoxa-1
(b) ~ Mouse
type I
(ws]), (hm~zygo~:a~ed)
Zinc-fingertrmxscfiptionfactor Histidase (enzyme in histidine metabolism) Fibroblast growth factor family Homeobox transcription factor
abmrmallt~ c-k# 107" 1Cttfigand, MGF mt M17F rvpl
Human
W Piebald trait
Mouse
51
Mouse Human Mouse
mt ?
Mouse
Sp Waardenburgsyndrome type 1 (WSl)C(heterozygotes affected)
Human
Zinc-fingertranscriptionfactor
.-
B~
Protein tyrosine kinase receptor Protein tyrosine kinase receptor Growth factor, ligand of c-kit
Pax3 PaX3
bHLH-ZIPtranscription factor bHLH-ZIPtranscription factor Membrane-bound copper-containing enzyme Paired-box transcription factor Paired-box transcription factor
(O uue-omet I m m q Io~ Human
X-linked Alport Syndrome
60L4A5
Collagen component
Human
Alportsyndrome, autosomal recessive
Human
Sticklersyndrome
Collagencomponents Collagencomponent
Human Mouse Mouse
Osteogenesisimpeffecta Mov13tg Other tramgenic#
COL4A3, COL4A4 COL2A1 COLIAI/COL1A2 COLIA1 O')LIA1
Mouse Mouse
Osteo/m/ms~~e
C.$rC
op e
c~m
with progressive loss of strlal function, but again the role of similar mutations in causing deafness in the human population is not known. However, the human counterparts of the splotch (Sp) and mtcropbthaimta (rat) loci, PAX3 and M/TF, respectively, may account for significant numbers of hearing-impaired humans. Migratory melanocyte abnormalities occur in splotch and mtcropbthalmia mutants, and in Waardenburg syndrome, and a lack of strial melanocytes may explain the deafness in Waardenburg syndrome. The human homologue of the mi gene, M/TF, has recently been described; interestingly, it maps to the same region of chromosome 3 as one of the genes involved in Waardenburg syndrome type 2 (Refs 28, 29). Waardenburg syndrome type 2 may be much more common among those with hearing impairments than has previously been supposed, since the pigmentary disturbances are not always obvious in this disorder.
Neuroeplthellal defects It is likely that the majority of cases of genetic deafness in humans are due to mutations that affect the sensory neuroepithelium, the organ of Corti. Early structural or functional anomalies of the organ of Corti
Collagen components Collagen component Collagen component Tyrosine kinase growth factor receptor Macrophage colony-stimulatinghctor
lead to progressive secondary hair-cell degeneration. This type of pathology is usually autosomal recessive in inheritance and not part of an obvious syndrome; of the one in 2000 children born with a genetic hearing impairment, approximately two-thirds have non-syndromic, autosomal recessive deafness. However, none of the genes responsible have yet been identified, in humans, mice or other vertebrates. This contrasts with the progress made in identifying genes involved in retinal degeneration, where many different mutations in key elements of the transduction cascade and proteins involved in the structure of the receptor cells have been described (e.g. rhodopsin, rod cGMP phosphodiesterase [3 subunit, peripherin and ROM1; see, for example, Ref. 50). None of the proteins involved in normal auditory transduction have been identified, although there is a long list of genes and proteins reported to be expressecl in the inner ear, including structural proteins such as collagens, actins and myosins. Unlike the visual or chemosensory systems, in which secondary messengers are involved in tran~duction, auditory transduction requires very fast responses to analyse the rapidly changing components of sounds. Therefore, directly
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XsmJ! 1. (contd) species
tecta/disease
Gem
Type of pteteta
Human Mouse Human Human
Charcot-Mafie--Toothdisease type 1A Tr Charcot-Marie-Toothdisease type 1B X-linkedCharcot-Marie--Tooth disease
Growth-controUing pet-ipheral myelh-iprotein Growth-controlling peripheral myelin protein Membrane glycoproteinof myelin Connexin 32, gap junction protein
Human
Cockaynesyndrome
Human
Huntersyndrome
PMP22 Prop22 Po Cx32 ERCC IDS, SIDS
Human
Hurlersyndrome
IDUA
et-L-iduronidase, enzyme in
DNA excision repair protein Iduronate-2-sulphatase, enzyme in mucopolysaccharide metabolism mucopolysaccharide metabolism
Human
Neurofibromatosis2
NF2
Merlin, tumour suppressor protein
Human
Norriedisease
NDP
Homology with mucins and growth faaors with cystine knot motif
Human
Crouzonsyndrome
FGFR2
Fibroblast growth faaor receptor
Human
Generalizedresistance to thyroid hormone (GRTH)f
bTR#
Thyroid receptor protein
Human
Aminoglycoside-induced deafness
Ribosomal component
Human
Diabetes mellims with hearing loss
Human
Wolfram s~oadrome
Human
Diabetes mellitus and deafness
Mitochondrial 12S rRNA Mitochondrial tRNALeu(UUR) 7.6 kb mitochondrial deletion 10.4 kb mitochondrial deletion
tRNA Severalmitochondrial genes deleted Several mitocbo~drial genes deleted
Table includes only identified genes: many additional loci are known to be involved, but are not yet cloned (see, for example, Refs 2, 7). ~, transgenlc interference with a known gcrie, aNot known whether a morphogenetic or cochleo..saccular defect or both am involved (reviewed in Re£ 20). bear pinna malformations sometimes seen, but no report of hearing impairment. Hot known ff a morphog~netic or ¢ochlco-.saccular defect or both is involved (reviewed in Ref. 20). dThe hearing ability of these tran~ genlc mice is not known, but Movl3 transgenics show a progressive hearing impairment, eThe hearing ability of these transgenic mice and the osteopetrosis mutant is not known, but osteopctrosis in humans leads to a hearing impairment, rDeafness is a feature only In autosomal recessive GRTH, In which the whole gene is deleted, and may not occur in autosomal dominant ORTH, due to point mutations In the ilgand-binding domain. Deafness is severe or profound and is present from childhood. gated channels are likely to be involved3t-33. The mechanosensory channels identified in lower organisms, such as that encoded by mscL in E. co8, by mec-4, mec-6, mec-lO and deg-1 in nematodes, and by the relamd amiloride-sensitive Na + channel components a, 13 and ? rENaC in rats (see, for example, Refs 34-37), may provide useful clues to identify the genes encoding elements of the auditory transduction process. Such genes will be obvious candidates for mutations causing neuroepithelial defects, as will genes encoding structural proteins (such as myosins3S and tubulins39) that may link the mechanoreceptor channels to the architecture of the sensory cells, thus permitting transmission of mechanical forces to the channels. For neuroepithelial deafness, it would be surprising ff the genes responsible were not expressed in the neuroepithelium or auditory nerve. The current absence of candidate mammalian genes for neuroepithelial defects, combined with the importance of this type of pathology in humans, has led us and others to adopt positional cloning strategies in attempts to identify these genes (see, for example, Ref. 40). Many mouse genes involved in neuroepithelial deafness have been localized, but none identified (see,
for example, Ref. 6). Identifk:ation of such genes directly in humans is likely to be extremely difficult because of the heterogeneity involved, and it is only in the past few months that any recessive loci have even been localized, using large consanguinous families from isolated communities. Three of these loci occur on llq13, 13q12 and at the pericentromeric region of chromosome 17 (Refs 41, 42; T. Friedman and J. Asher, pers. commun.). Genes involved in autosomal dominant, non-syndromic deafness have been localized to lp32 and 5q31 (Refs 43, 44). lhese mappings in mice and humans will be of great help in eventually identifying the genes involved.
The sbaker-I mouse mutant We have focused on the sbaker-I mouse mutant as a model for non-syndromic deafness it) humans. Over 1000 meioses in backcrossed mice segregating the shaker-1 mutation were analysed, and no recombination between sbaker-I and the olfactoo, marker protein (Omp) gene on mouse chromosome 7 was found (Ref. 45). Using Omp as a staring point, a YAC contig spanning 1.5 Mb has been identified, and we are currently examining genes identified within the
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(b)
(a)
Distal ---
RT6
13.1
-
-
=
13.2
Hbb
11p15.5
Rt6 Pth, Calc
11p15.2-15.1
sh-1 Ornp
11q13.5
11q13
13.3 I lcM
13.4
USH1B
DFNB2
I 13.s
I OMP
vertigo42 (recently renamed DFNB2). "Ihe shaker-1 gene might be the homologue of either of these human loci, or indeed of both; if USHIB and DF3/732 are found to ..be allelic, it would not be the first time that distinct clinical diseases have been found to be caused by mutations of the same gene.
[Homologue of sh-I]
late.onset hearing loss A number of genes responsible for syndromes including 14.2 - Tyr 11q14.3 hearing loss have now been identified, and these are shown TYR 14.3 in Table 1 (par c). Most of these syndromes express hearing loss as a minor or inconProximal sistent or late-onset feature. As such, they account for a relatively small proportion of the total hearing impairments in (c) the human population, but Profound Vestibular Retinitis they are no less interesting deafness amflexea pigmentosum because of that. Several of the genes were identified because + + + USHIB the clinical features suggested + +/DFNB2 an obvious class of candidate genes. The collagen gene + + No RP up to shaker.1 mutations in Alport synone year old drome, Stickler syndrome and osteogenesis imperfecta are examples. The accumulation Fmtam 4. Comparison of the map position of shaker.1 mouse mutant with that of Usher of mucopolysaccharides in syndrome type IB (UgH1B) in humans, (a) Part of human chromosome llq, showing Hunter and Hurler syndromes approximate locations of the USHIB,DFNB2, OMP,RT6and TERgenes,as well as the that lead ultimately to hearing location of the predicted human homologue of the shaker.l (sh.l) gene, (b) Mouse loss suggested an inborn error chromosome 7, giving locations of relevant genes around sh-1with the positions of their human homologue~ listed on the right, No recombinations between sbaker.l and Omp have of carbohydrate metabolism, been found in over 1000 backcrossed mice. (c) Comparison of the phenotypes of shaker.l, which was subsequently found DFNB2 and USHIB, All three show hearing Impairmentand at least some vestibular to be the case. Peripheral dysfunction, and USH1Bcauses retlnltis pigmentosum(RP); however, there Is as yet no myelin protein defects in evidence of RP In shaker.1 mutants up to one year old (K,P. Steel,J. Cable and Charcot-Marie--Tooth disease M, Mahony, unpublished). were suggested by the observed progressive peripheral nonrecombinant region as potential candidates for the neuropathy in this disease, and identification of the shaker-1 gene itself. One promising candidate is a mouse Pmp22 gene and its mutation in trembler eDNA clone identified from an exon-trapped fragment mutants pointed to human PMP22 as a candidate for that shows homology with the unconventional group of one of the genes involved in this disease. Other myosins. The likely importance of myosin-like proteins genes have been identified by a classical positional in the function of the hair cells (see, for example, Ref. cloning approach, such as the gene involved in 38) supports this theory. Norrie disease. We predict that the human homologue of shaker-1 A surprising feature of the list is the range of mitowill be located within the syntenic group of genes on chondrial mutations that lead to progressive deafness; human chromosome llq13, as illustrated in Fig. 4. This this may be a result of the dependence of cochlear region of 1lq holds a number of sensory system genes, homeostasis on oxidative phosphorylation and hence including several genes causing visual defects [such as on genes encoded by mitochondrial DNA. One interestUsher syndrome type 1B (USH1B), adFEVR and Best's ing feature of the mitochondrial mutations is the A--cG disease] and two loci responsible for recessive deafness: substitution at position 1555 in the gene encoding 12S Usher syndrome type 1B (USH1B), which results in rRNA47A8. This particular change has been implicated in deafness, vestibular dysfunction and progressive enhanced sensitivity to aminoglycoside-induced cochretinitis pigmentosum46; and nonsyndromic recessive lear damage, since it may favour of the steric interaction deafness (NSRD2), which is deafness with occasional of the aminoglycoside with the ribosome by increasing 14.1
-
Mod2
6p2,~-24
/ j • •
i i i
m
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access to the ribosome cleft47. This observation illustrates how an investigation into an inherited form of deafness can provide very useful hypotheses to explain the molecular basis of environmentally triggered cochlear damage. The range of types of gene involved in causing nearing loss given in Table 1 and the fact that hearing loss occurs as a part of so many diverse syndromes in humans probably refects two aspects of the auditory system: (1) the complexity of the inner ear and the wide range of structural components involved in maintaining optimum function in detecting very small and rapidly changing movements; and (2) the susceptibility of the sensory hair cells to damage and their inability to regenerate. Middle ear defects Two genes, M s x l and endotbelin-1, have been identified by transgenic disruptions as important in the formation of the middle ear in mice49,5°. However, in neither case did the homozygous deficient mice survive for long after birth. The future Identification of some of the genes involved in auditory transduction, either by positional cloning or by cloning mammalian homologues of mechanosensory genes of lower organisms, can surely be only a few months away. When these genes are known, it will be an exciting time of progress, not only for understanding the molecular basis of auditory transduction, but also for understanding the pathological processes involved in the non-syndromic autosomal recessive deafness that is so important in the human population.
Acknowledgements We thank A, Read, C. Petit, T. Friedman and J. Cable for communicating their findings before publication, D. Hughes for comments on the manuscript, and A. Pearce and P, Moorjani for bibliographic assistance.
References I Morton, N.E, (1991) Ann. N.Y. Acad. Set. 630, 16--31 2 Duyk, G., Gastier,J,M. and Mueller, R.F. (1992) Nature Genet. 2, 5-8
3 Konigsmark, B.W. and Godin, R,J. (1976) Genetic and Metabolic Deafness, W,B. Saunders Co. 4 Fraser, G.R. (1976) The Causes of Profound Deafness in Childhood, Johns Hopkins UniversityPress 5 Deol, M.S. (1968).]. Med. Genet. 5, 137-158 6 Lyon, M.F. and Seade, A.G. (1989) Genetic Variants and Strains of the Laboratory Mouse (2nd edn), Oxford University Press 7 Nadeau, J.H., Kosowsky, M. and Steel, K.P. (1991) Ann. N. E Acad. Sct. 630, 49.-67 8 Davis, A.C. (1989) Int.J. Eptdemtol. 18, 911-917 9 Searle, A.G., Edwards, J.H. and Hall,J.G. (1994)J. Med. Genet. 31, 1-19 10 Steel, K.P. (1991) Ann. N.Y. Acad. Sci. 630, 68-79 11 Deol, M.S. (1966) Nature 209, 219--220 12 Steel, K.P. and Bock, G.R. (1983) Arch. Otolaryngoi. 109, 22-29 13 Keynes, R. and Krumlauf,R. (1994) Annu. Rev. Neurosci. 17, 109-132 14 Repressa, J., Leon, Y., Miner, C. and Giraldez, F. (1991) Nature 353, 561-563
15 Mansour, S.L., Goddard, J.M. and Capecchi, M.R. (1993) Development 117, 13-28 16 Lufkin,T. etal. (1991) Cell66, 1105--1119 17 Chisaka, O., Musci, T.S. and Capecchi, M.R. (1992) Nature
355, 516-520 18 Corey, D.P. and Breakefield, X.O. (1994) Proc. NatlAcad. Sci. USA 91, 433--436 19 Tassabehji,M. et al. (1992) Nature355, 635--636 20 Steel, K.P. and Smith, R.J.H. (1992) Nature Genet. 2,
75-79 21 Vortkamp, A., Gessler, M. and Grzeschik, K-H. (1991) Nature 352, 539--540 22 Kacser, H., Mya Mya, K. and Bulfield,G. (1979) in Models for the Study of Inborn Errors of metabolism (Hommes,
F.A., ed.), pp. 43-53, ElsevierScience Publishers 23 Steel, K.P., Barkway, C. and Bock, G.R. (1987) Hear. Res.
27, 11-26 24 Cable,J., Huszar, D., Jaenisch, R. and Steel, K.P. (1994) Pigment Cell Res. 7, 17-32 25 Cable,J., Barkway, C. and Steel, K.P. (1992) tlear. Res. 64,
6-20 26 Steel, K.P., Davidson, D.R. and Jackson, I.J. (1992) Development 115, 1111-1119 27 Hulten, M.A., Honeyman, M.M., Mayne, A.J. and Tarlow, M.J. (1987)J. Med. Genet. 24, 568-571 28 Tachibana, M. et al. (1994) Hum. Mol. Genet. 3,
553-557 2.9 Hughes, A.E., Newton, V.E., Liu,X.Z. and Read, A.P. (1994) Nature Genet. 7, 509-512 .fro Wright,A.F. (1992) Trends Genet. 8, 85-91 31 Hudspeth, A.J. (1989) Nature341, 397-404 32 Pickles,J.O. and Corey, D.P. (1992) Trends Neurosci. 15, 254-259 33 Ashmore,J.F. (1994) Experimental Physiol. 79, 113-134 34 Canessa, C.M. et al. (1994) Nature367, 463.-467 35 Huang, M. and Chalfie, M. (1994) Nature 367, 467-47O 36 Hong, K. and Driscoil, M. (1994) Nature 367, 47O-473 37 Sukharev, S.I. et al. (1994) Nature 368, 265-268 38 Gillesple, P.G., Wagner, M.C. and Hudspeth, A.J. (1993) Neuron 11,581-594 39 Savage, C. et al. (1989) Genes Dev. 3, 870--881 40 Brown, S.D.M. and Steel, K.P. (1994) Hum. Mol. Genet. 3, 1453-1456 41 Guilford, P. et ai. (1994) Nature Genet. 6, 24-28 42 Guilford, P. et ai. (1994) Hum. Mol, Genet. 3, 989-993 43 Leon, P.E. et al. (1992) Proc. Natl Acad. Set. USA 89, 5181-5184 44 Coucke, P. et al. (1994) N. Engi.J. Med. 331,425--431 45 Brown, K.A., Sutcliffe,M.J., Steel, K.P. and Brown, S.D.M. (1992) Genomics 13, 189-193 46 Kimbeding,w.J. et al. (1992) Genomics 14, 988--994 47 Hutchin, T. et al. (1993) Nucleic Acids Res. 21, 4174--4179 48 Prezant, T.R. et al. (1993) Nature Genet. 4, 289--294 49 Kurihara,Y. et al. (1994) Nature 368, 703-710 50 Satokata, I. and Maas, R. (1994) Nature Genet. 6, 348-356 51 Truslove, G.M. (1956) J. Genet. 54, 64--86
K.P. STEEL iS IN THE MRC INSTHtr~ OF HEARING RESr~tR~ UNIV~SHY PAR& NOITINGHA~ L/K NG7 2RD; £1~M. BROWNns US rue DF.PARrmmNrOFBmcHF.msmv XND MOLE~L&¢ GENL~C~ Sr Macy's HOSPITAL MF.DICAL Scao01~ NORFOLgPIAC~ LONOON, UK W2 IPGo
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26 Heston, W.E. and Vlahakis, G. (1961)J. Natl Cancerlnst. 27, 1189--1196 27 Heston, W.E. and Vlahakis, G. (1961)J. Natl Cancerlnst. 26, 969-983 28 Wolff, G.L. (1970) CancerRes. 30, 1731-1735 29 Wolff, G.L., Kodell, R.L.and Cameron, A.M. (1982)J. Tox. Env. Health 10, 131-142 30 Wolff, G.L. etal. (1987)Carcinogenesis& 1889-1897 31 Siracusa, L.D. (1991) Ann. N.Y. Acad. Sci. 642, 419-.-430 32 Michaud, EJ. et al. (1994) Proc. Natl Acad. Sci. USA 91, 2562-2566 33 Miller, M.W. et al. (1994) EMBOJ. 13, 1806-1816 34 Papaioannou, V.E. and Mardon, H. (1983) Dev. Genet. 4, 21-29 35 Rinchik, E.M. and Russell, L.B. (1990) in Genome Analysis (Vol. I) (Davies, K. and Tilghman, S., eds), pp. 121-158, Cold Spring Harbor Laboratory Press 36 Siracusa, L.D. et al. (1987) Genetics 117, 93-100 37 Seafle, A.G. (1968) Comparative Genetics of Coat Colour in Mammals, Academic Press 38 Sage, R.D. (1981) in The Mouse in Biomedical Research (Vol. I) (Foster, H.L., Small,J.D. and Fox, J.G., eds), pp. 39-90, Academic Press
L.D. SXRACUSA tS IN THE JWFERSOS CASC~R ISSTITt~, THOMAS JEFI:ERSOJV UNIVERSITY, JDEPARTMENTOF MICRO. BIOLOGY AND IMMUNOLOGY, 233 SOUTH IOTH STREET, PHULAOE~HL¢,PA 19107, USA.
Genes and deafness T h e mammalian inner ear rightly deserves its alternative name, the labyrinth. It is a remarkably complex structure (see Box 1). Endol?mph- and perilymph-filled channels course around the cochlea, the attditory sense organ, and around the sacculus, utriculus and the three semicircular canals, which together form the vestibular part of the inner ear that detects head position and movement and hence aids balance, The sensory epithelia within the inner ear are highly organized arrays of senso W hair cells and supporting cells. The organ of Corti within the cochlear duct is particularly complex and precise in its arrangement of a variety of types of sensory and supporting cells, Given this complexity at the cellular and gross structural levels, it is not surprising that many genes have been found to be involved in controlling the development and function of the inner ear. Indeed, mutations at many different loci in both mice and humans are known to cause hearing impairment1-'1, About one in every 1000 children is born with a significant hearing impairment, and about half of these suffer from the relatively common genetic disorder of hereditatT deafness. Genetic factors are highly likely to play a role in susceptibility to hearing loss later in life, and understanding how single gene mutations cause deafness should help us to unravel the interactions between genetic predisposition and environmental experience that cause late-onset hearing loss. In the British population, 16°,6 of all adults suffer from a significant hearing impairmentS; this proportion is probably similar elsewhere,
KAREN P. STEEL A N D STEPHEN D . M . BROWN
Many d~erent &enosappear to be involved in the development anti, notion of the mammalian innerear. Some of the genes involvedduring early innerear morphogenests have boonIdent~l using mutations or targetted transgenic interruption, while a hanOI of genes involved in pigmentation anomalies associated with hearing impairment have been clone~ Severalgenes involved in syndromtc late.o~et hearing loss have also been ldantO~e~However, the maJori~ of cases of hereditary hearing impalrmentJ~om chiMhoodprobably involve g e n e s expressed in the sensory neurospithelia of the inner ear, and n o n e of the genes or mutations causing this type of de~ess haveyet b m id~t~e~ Here, we review the progress that has boonmade tn~ndinggenes for de~ess and in using mouse mutants to elucidate the biological basts of the bearing deficit. and thus represents a large health and communication problem for both sufferers and their families. Mice are likely to be extremely useful in identifying some of the genes involved in causing deafness, as well as in understanding how those genes and their mutations interfere with the normal process of development and functton of the tuner ear5 • ' • ,9, m. The m any similarities between the mouse and human auditory systems suggest that similar cascades of gene action are likely to be involved in the development of the system in the
TIG DECEMBER1994 VOL. 10 NO. 12
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i
:
~D)
~
ossi
II
\ ~-Pinna
bca,a vesuou,,
)'Xt~M.~"--']li~ 1 (r'x'~~ \ d o
~
~ar canal
~./Stria vascularis
Spiral
Eustachian tube
o£-~-.-~.at~.. )
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~
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(a) The human outer, middle and inner ear. The outer ear canal leads to the tympanic membrane (ear drum), and ~und vibrations are transmitted from there to the inner ear via the three middlL• ear ossicles, the malleus, incus and stapes. The stapes inserts into the oval window at the base of the cochlea. Co) One rum of the cochlear duet. During sound stimulation, the organ of Corti vibrates and the sensory hair cells are stimulated by a shearing movement between the top surface of the organ of Cotti and the rectorial membrane. Most afferent neurons innervate inner hair cells (shown here on the left of the black pillar cells), and their cell bodies form the main par of the spiral ganglion, which in mm leads to the cochlear nerve. The three or more row~ .of outer hair cells (to the right of the pillar cells) receive primarily efferent innervation, and are believed to act as motor cells, enhancing the response of the cochlea (see, for example, Sef. 55). The stria vascularis on the lateral wall of the cochlear duct has a key role in generating the high resting endocochlear potential in the endolymph that fills the scala media, and it also controls the high K+ and low Na+ concentration in the endolymph. Scab vestibuli and scala tympani both contain perilymph, a fluid that is much more like a normal extracellular fluid, with low K+and high Na+levels and no high resting potential. The stippled area represents bone. (c) A typical sensory hair cell. The actin-filledstereocilia are arranged in rows of waded heights at thc top of the hair cell. Tip links connect the shorter stereocilia to the adjacent taller stereo:ilia, and it is stretching of the tip links that is
(C)
Corti
Tip links - -
Stereocilium
Haircell
,)
thought to activate the transduction channels of the hair ceil, leading to depolarIzation and initiationof synaptic activity at the base of the cell (see, for example, Refs 51, 52), two species. Furthermore, there are many parallels in the types of pathology seen in the two species (see Box 2). This review outlines some of the key findings concerning genes that cause deafness and our understanding of how they affect the development and function of the inner ear. The major types of pathology as defined in Box 2 are considered in turn.
Morphosenetic defects The gross structural malformations seen in cases of morphogenetic defects of the inner ear suggest that the development of the labyrinth is disrupted very early in embryogenesis. The malformations may affect the vestibular part of the inner ear or the cochlea or both. Art example in which the vestibular part of the inner ear is primarily affected, the fidget mouse inner ear, is shown in Fig. 1. One interesting feature of these morphogenetic defects is the association of neural tube and inner ear anomalies: minor or transient neural tube defects are frequently detectable at the time that the inner ear is forming, and labyrinthine malformations usually occur in cases of severe neural tube defects 1tJ2. Figure 2
illustrates one example in which neural tube defects in embryos homozygous for the splotcb mutation are associated with malformations of the developing inner ear. It appears that the neural tube has an inductive influence on the development of the labyrinth. The developing neural tube has been extensively studied, and the expression patterns of a number of genes thought to be involved in controlling its development have been described 13. One of the earliest genes to be proposed as a potential inducing agent for inner ear morphogenesis is Int2, which is expressed in the neural tube. Exposure to Int2 antisense oligonucleotides does indeed disrupt otocyst development in vitro t4. Transgenic disruptions of lnt2 and another gene, Hoxa-I (previously known as Hoxl.6), which is known to be expressed in the neural tube opposite the developing labyrinth, have been shown to lead to abnormal morphogenesis of the inner ear 1s-re. In contrast, transgenic disruptions of Krox20 do not affect labyrinthine morphogenesis, despite the fact that this gene is known to be expressed in rhombomere 5, directly adjacent to the otocyst. One consequence of the interaction between the neural tube and the inner ear is that the
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humans homozygous for mutations in the homologofis gene GLI3 has not been reported. In this case, humans heterozygous for GLI3 mutations have Grieg cephalopolysyndactyly syndrome 21 (GCPS), which does not usually involve deafness. Mice homozygous for the b/s mutation have maternal histidinaemia resulting from abnormal histidase activity, which causes transient neural tube anomalies and abnormal formation of the vestibular part of the inner ear in the offspringZZ; it is not clear whether there is any human correlate of these ear defects in maternal hisfidinaemia. Disruption of both Int2 and Hoxa-1 causes abnormalities of the neural tube and inner ear, but, again, it is not known whether mutations in these genes cause deafness in humans. The interaction between development of the neural tube and the inner ear needs further investigation, but at least some of the key players are now known, , ¢oehk, o-samflar defects Cochleo-saccular abnormalities are found in the type of deafness that is often associated with white spotting of the hair or skin in many different mammals. Darwin noted that white cats are often deaf, and the association was probably known long before. White spotting of the coat is believed to be caused by mutations that affect migratory melanocytes, which originate in the neural crest. It has long been known that there genes responsible for morphogenetic defects may not are melanocytes in the inner ear, and that melanocytenecessarily be expressed in the inner ear, but only in like cells form part of the stria vascularis of the cochlea; the neural tube. however, albino mammals hear perfectly well so it is obviously not the pigment-producing ability of the Ident~ed 8enes melanocytes that is important in this respect. It appears Some of the identified genes involved in morpho- that some other feature of melanocsrtes is essential for genetic defects are listed in Table 1. The splotch (St,) normal strial function. mutation in mice causes severe neural tube defects and The stria vascularis in normal mammals is responinner ear malformations when homozygous tl. How- sible for generating the unusually high extracellular restever, humans with mutations in the homologous gene ing potential, the endocochlear potential (EP), and the PAX3, who have Waardenburg syndrome type 1 (WS1) characteristic ion composition of high concentrations of (Ref. 19), are heterozygous for the mutation, and the K÷ and low concentrations of Na + in the endolymph in cause of the hearing impairment in WS1 is not known the lumen of the cochlear duct. In normal mice, the EP (reviewed in Ref. 20). The extra toes mouse mutation is around +100 mV, but in mutant mice with no (Xt) also causes neural tube defects and inner ear melanocytes in their stria vascularis this potential is malformations in homozygotes, but the condition of abolished a3. A causal relationship between the lack of melanocytes and the absence of a normal EP is suggested by the discovery that some mu(a) Crus (b) tations are 'leaky'; that is, some melanoblasts Presumptive do manage to find their way to the stria _ . . .~ commune Sunerior eostenor I ca,.,ol superior canal during developmenta4, 2~. In these ears, a positive EP can always be recorded, although it may not be as high as normal, while a significant EP has never been found in any mice Lateral./~'li"~'~"~-"~-"~'~ I 0 ! without strial melanocytes. Figure 3 illustrates an example of this type of mutant, the viable dominant spotting, mtmse mutant: melanoRO~lodlv//'XJ C )"Utriculus , . ~ Oval cytes have populated the stria of the left cochlea, which has an EP, but not the right cochlea, which has no EP. Asymmetry is com"-"- "Cochlea"- v mon in this type of hearing impairment. The particular property of the melanocyte Ihctm~ 1. An example of a malformed inner ear in a morphogenetic mutant important for striai function is unknown, of the mouse. (a) Outline of the normal mouse inner ear, (b) Inner ear of the but we do know that these cells are refidget mutant, showing severe deformityof the vestibularpart of the inner quired throughout adult life for normal strial ear, including absence of the lateral semicircularcanal. In other mutants, the function rather than being needed only at a cochlea is more severelymalformed. (Modifiedfrom Ref. 10.) critical stage of development. Therefore, the TIG DECEMBER1994 VOL. 10 No. 12 430
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F t ~ 2. Illustration of the relationship between inner ear malformations and neural tube defects. (a) Section through an 11.5 d.p.c. (days post coitum) normal mouse embryo, showing the normal neural tube (NT) and developing otic vesicle(OV). (b) An example of a severely malformed neural tube in a splotchhomozygote embryo, with associated abnormal morphogenesis of the labyrinth. (Modified from Ref. 11.) melanocytes could be involved in some aspect of the heterozygous form in people with piebald trait (a pigion-pumping processes involved in generating the EP. mentation defect causing white skin and hair patches). An EP is necessary for the normal functioning of the If these mutations have effects similar to the W musensory hair cells, and the result of an EP dose to zero tations in mice, it is the homozygotes who would be is a gready raised threshold to sound. Eventually, strial deaf. However, only one putative homozygote (who dysfunction leads to secondary hair-cell degeneration was deaf) has ever been reported and it is not clear whether he carried mutations in K/Tor in another gene and severe to profound hearing impairment. Various alleles of the dominant spotting ( W ) and affecting pigmentation aT. No mutations of the human steel(Sl) loci, which produce white spotting of the coat, homologue of SI (MGF) have been reported. Thus, it have been analysed extensively to investigate the role seems that the homologues of Wand S! in the human of melanocytes in cochlear function, and to establish population do not play a major role in deafness. The that the hearing impairment is mediated by strial light allele of the brown locus, Btt, causes progressive dysfunction resulting from a lack of melanocytes. melanocyte death and premature greying of the coat, Melanoblast migration away from the neural crest begins as normal in Wand SI mutant mice and these precursor cells start to differentiate normally by explesslng melanocyte-specific genes; however, in these mutants there are always fewer melanoblasts than normal and their numbers decrease between 11 and 12-13 days post cottum (Ref. 26; J. Cable et al., unpublished). This decrease in numbers, compared with the rapid prolfferation in numbers seen in control embryos at this stage, suggests that the c-kit growth factor and its iig'and, which are encoded by Wand Sl, respectively, may act as survival factors for mdanoblasts. Since melanoblasts migrate from the neural crest to the inner ear, genes causing cochleo-- Me,tam.3. An example of the (a) leR and (b) right cocl'dea from a single cochleo--saccular viable dominant spotting ( ga/ Wv) mouse mutant. The single coiled saccular defects may not necessarily line represents the turns of the cochlear duct, and the cross through the line shows the be expressed in the inner ear. position from which the endocochlear potential was recorded. Dots represent the distribution of melanocytes in the stria vascularisalong the entire Identified genes length of the cochlea. The left cochlea had a near-normal endocochlear potential Some of the genes involved in of 90 mV and melanocytespresent in the middle half of the length of the cochlear cochleo-saccular defects are listed in duct, while the right cochlea had an endocochlear potential near zero (-2 mY) Table 1 (part b). A number of K/T and no sign of any melanocytesthroughout the cochlea. OW, oval window: mutations have been identified in RW, round window. (Modifiedfrom Ref. 25.) TIG DECEMBER1994 VOL. 10 No. 12
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T.Amm1. Genes invohred in deafness
cae
Wpeofpmmu
Pax-3 PAX3
Paired-box transcription factor Paired-box transcription factor
speam
tecm/O~me
Mouse Human
S p ~ ' ~ Waatdenburg~
Mouse Human
Xt (homozygo¢~ ~ Gli3 Grieg cephalopolysyndactylysyndrome GU3 (GCPS)b (hetemzygotes affected)
Mouse
Ha/
His
Mouse
lnt2tg
F~
Mouse
Hoxl.6 t8
Hoxl.6, Hoxa-1
(b) ~ Mouse
type I
(ws]), (hm~zygo~:a~ed)
Zinc-fingertrmxscfiptionfactor Histidase (enzyme in histidine metabolism) Fibroblast growth factor family Homeobox transcription factor
abmrmallt~ c-k# 107" 1Cttfigand, MGF mt M17F rvpl
Human
W Piebald trait
Mouse
51
Mouse Human Mouse
mt ?
Mouse
Sp Waardenburgsyndrome type 1 (WSl)C(heterozygotes affected)
Human
Zinc-fingertranscriptionfactor
.-
B~
Protein tyrosine kinase receptor Protein tyrosine kinase receptor Growth factor, ligand of c-kit
Pax3 PaX3
bHLH-ZIPtranscription factor bHLH-ZIPtranscription factor Membrane-bound copper-containing enzyme Paired-box transcription factor Paired-box transcription factor
(O uue-omet I m m q Io~ Human
X-linked Alport Syndrome
60L4A5
Collagen component
Human
Alportsyndrome, autosomal recessive
Human
Sticklersyndrome
Collagencomponents Collagencomponent
Human Mouse Mouse
Osteogenesisimpeffecta Mov13tg Other tramgenic#
COL4A3, COL4A4 COL2A1 COLIAI/COL1A2 COLIA1 O')LIA1
Mouse Mouse
Osteo/m/ms~~e
C.$rC
op e
c~m
with progressive loss of strlal function, but again the role of similar mutations in causing deafness in the human population is not known. However, the human counterparts of the splotch (Sp) and mtcropbthaimta (rat) loci, PAX3 and M/TF, respectively, may account for significant numbers of hearing-impaired humans. Migratory melanocyte abnormalities occur in splotch and mtcropbthalmia mutants, and in Waardenburg syndrome, and a lack of strial melanocytes may explain the deafness in Waardenburg syndrome. The human homologue of the mi gene, M/TF, has recently been described; interestingly, it maps to the same region of chromosome 3 as one of the genes involved in Waardenburg syndrome type 2 (Refs 28, 29). Waardenburg syndrome type 2 may be much more common among those with hearing impairments than has previously been supposed, since the pigmentary disturbances are not always obvious in this disorder.
Neuroeplthellal defects It is likely that the majority of cases of genetic deafness in humans are due to mutations that affect the sensory neuroepithelium, the organ of Corti. Early structural or functional anomalies of the organ of Corti
Collagen components Collagen component Collagen component Tyrosine kinase growth factor receptor Macrophage colony-stimulatinghctor
lead to progressive secondary hair-cell degeneration. This type of pathology is usually autosomal recessive in inheritance and not part of an obvious syndrome; of the one in 2000 children born with a genetic hearing impairment, approximately two-thirds have non-syndromic, autosomal recessive deafness. However, none of the genes responsible have yet been identified, in humans, mice or other vertebrates. This contrasts with the progress made in identifying genes involved in retinal degeneration, where many different mutations in key elements of the transduction cascade and proteins involved in the structure of the receptor cells have been described (e.g. rhodopsin, rod cGMP phosphodiesterase [3 subunit, peripherin and ROM1; see, for example, Ref. 50). None of the proteins involved in normal auditory transduction have been identified, although there is a long list of genes and proteins reported to be expressecl in the inner ear, including structural proteins such as collagens, actins and myosins. Unlike the visual or chemosensory systems, in which secondary messengers are involved in tran~duction, auditory transduction requires very fast responses to analyse the rapidly changing components of sounds. Therefore, directly
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XsmJ! 1. (contd) species
tecta/disease
Gem
Type of pteteta
Human Mouse Human Human
Charcot-Mafie--Toothdisease type 1A Tr Charcot-Marie-Toothdisease type 1B X-linkedCharcot-Marie--Tooth disease
Growth-controUing pet-ipheral myelh-iprotein Growth-controlling peripheral myelin protein Membrane glycoproteinof myelin Connexin 32, gap junction protein
Human
Cockaynesyndrome
Human
Huntersyndrome
PMP22 Prop22 Po Cx32 ERCC IDS, SIDS
Human
Hurlersyndrome
IDUA
et-L-iduronidase, enzyme in
DNA excision repair protein Iduronate-2-sulphatase, enzyme in mucopolysaccharide metabolism mucopolysaccharide metabolism
Human
Neurofibromatosis2
NF2
Merlin, tumour suppressor protein
Human
Norriedisease
NDP
Homology with mucins and growth faaors with cystine knot motif
Human
Crouzonsyndrome
FGFR2
Fibroblast growth faaor receptor
Human
Generalizedresistance to thyroid hormone (GRTH)f
bTR#
Thyroid receptor protein
Human
Aminoglycoside-induced deafness
Ribosomal component
Human
Diabetes mellims with hearing loss
Human
Wolfram s~oadrome
Human
Diabetes mellitus and deafness
Mitochondrial 12S rRNA Mitochondrial tRNALeu(UUR) 7.6 kb mitochondrial deletion 10.4 kb mitochondrial deletion
tRNA Severalmitochondrial genes deleted Several mitocbo~drial genes deleted
Table includes only identified genes: many additional loci are known to be involved, but are not yet cloned (see, for example, Refs 2, 7). ~, transgenlc interference with a known gcrie, aNot known whether a morphogenetic or cochleo..saccular defect or both am involved (reviewed in Re£ 20). bear pinna malformations sometimes seen, but no report of hearing impairment. Hot known ff a morphog~netic or ¢ochlco-.saccular defect or both is involved (reviewed in Ref. 20). dThe hearing ability of these tran~ genlc mice is not known, but Movl3 transgenics show a progressive hearing impairment, eThe hearing ability of these transgenic mice and the osteopetrosis mutant is not known, but osteopctrosis in humans leads to a hearing impairment, rDeafness is a feature only In autosomal recessive GRTH, In which the whole gene is deleted, and may not occur in autosomal dominant ORTH, due to point mutations In the ilgand-binding domain. Deafness is severe or profound and is present from childhood. gated channels are likely to be involved3t-33. The mechanosensory channels identified in lower organisms, such as that encoded by mscL in E. co8, by mec-4, mec-6, mec-lO and deg-1 in nematodes, and by the relamd amiloride-sensitive Na + channel components a, 13 and ? rENaC in rats (see, for example, Refs 34-37), may provide useful clues to identify the genes encoding elements of the auditory transduction process. Such genes will be obvious candidates for mutations causing neuroepithelial defects, as will genes encoding structural proteins (such as myosins3S and tubulins39) that may link the mechanoreceptor channels to the architecture of the sensory cells, thus permitting transmission of mechanical forces to the channels. For neuroepithelial deafness, it would be surprising ff the genes responsible were not expressed in the neuroepithelium or auditory nerve. The current absence of candidate mammalian genes for neuroepithelial defects, combined with the importance of this type of pathology in humans, has led us and others to adopt positional cloning strategies in attempts to identify these genes (see, for example, Ref. 40). Many mouse genes involved in neuroepithelial deafness have been localized, but none identified (see,
for example, Ref. 6). Identifk:ation of such genes directly in humans is likely to be extremely difficult because of the heterogeneity involved, and it is only in the past few months that any recessive loci have even been localized, using large consanguinous families from isolated communities. Three of these loci occur on llq13, 13q12 and at the pericentromeric region of chromosome 17 (Refs 41, 42; T. Friedman and J. Asher, pers. commun.). Genes involved in autosomal dominant, non-syndromic deafness have been localized to lp32 and 5q31 (Refs 43, 44). lhese mappings in mice and humans will be of great help in eventually identifying the genes involved.
The sbaker-I mouse mutant We have focused on the sbaker-I mouse mutant as a model for non-syndromic deafness it) humans. Over 1000 meioses in backcrossed mice segregating the shaker-1 mutation were analysed, and no recombination between sbaker-I and the olfactoo, marker protein (Omp) gene on mouse chromosome 7 was found (Ref. 45). Using Omp as a staring point, a YAC contig spanning 1.5 Mb has been identified, and we are currently examining genes identified within the
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(b)
(a)
Distal ---
RT6
13.1
-
-
=
13.2
Hbb
11p15.5
Rt6 Pth, Calc
11p15.2-15.1
sh-1 Ornp
11q13.5
11q13
13.3 I lcM
13.4
USH1B
DFNB2
I 13.s
I OMP
vertigo42 (recently renamed DFNB2). "Ihe shaker-1 gene might be the homologue of either of these human loci, or indeed of both; if USHIB and DF3/732 are found to ..be allelic, it would not be the first time that distinct clinical diseases have been found to be caused by mutations of the same gene.
[Homologue of sh-I]
late.onset hearing loss A number of genes responsible for syndromes including 14.2 - Tyr 11q14.3 hearing loss have now been identified, and these are shown TYR 14.3 in Table 1 (par c). Most of these syndromes express hearing loss as a minor or inconProximal sistent or late-onset feature. As such, they account for a relatively small proportion of the total hearing impairments in (c) the human population, but Profound Vestibular Retinitis they are no less interesting deafness amflexea pigmentosum because of that. Several of the genes were identified because + + + USHIB the clinical features suggested + +/DFNB2 an obvious class of candidate genes. The collagen gene + + No RP up to shaker.1 mutations in Alport synone year old drome, Stickler syndrome and osteogenesis imperfecta are examples. The accumulation Fmtam 4. Comparison of the map position of shaker.1 mouse mutant with that of Usher of mucopolysaccharides in syndrome type IB (UgH1B) in humans, (a) Part of human chromosome llq, showing Hunter and Hurler syndromes approximate locations of the USHIB,DFNB2, OMP,RT6and TERgenes,as well as the that lead ultimately to hearing location of the predicted human homologue of the shaker.l (sh.l) gene, (b) Mouse loss suggested an inborn error chromosome 7, giving locations of relevant genes around sh-1with the positions of their human homologue~ listed on the right, No recombinations between sbaker.l and Omp have of carbohydrate metabolism, been found in over 1000 backcrossed mice. (c) Comparison of the phenotypes of shaker.l, which was subsequently found DFNB2 and USHIB, All three show hearing Impairmentand at least some vestibular to be the case. Peripheral dysfunction, and USH1Bcauses retlnltis pigmentosum(RP); however, there Is as yet no myelin protein defects in evidence of RP In shaker.1 mutants up to one year old (K,P. Steel,J. Cable and Charcot-Marie--Tooth disease M, Mahony, unpublished). were suggested by the observed progressive peripheral nonrecombinant region as potential candidates for the neuropathy in this disease, and identification of the shaker-1 gene itself. One promising candidate is a mouse Pmp22 gene and its mutation in trembler eDNA clone identified from an exon-trapped fragment mutants pointed to human PMP22 as a candidate for that shows homology with the unconventional group of one of the genes involved in this disease. Other myosins. The likely importance of myosin-like proteins genes have been identified by a classical positional in the function of the hair cells (see, for example, Ref. cloning approach, such as the gene involved in 38) supports this theory. Norrie disease. We predict that the human homologue of shaker-1 A surprising feature of the list is the range of mitowill be located within the syntenic group of genes on chondrial mutations that lead to progressive deafness; human chromosome llq13, as illustrated in Fig. 4. This this may be a result of the dependence of cochlear region of 1lq holds a number of sensory system genes, homeostasis on oxidative phosphorylation and hence including several genes causing visual defects [such as on genes encoded by mitochondrial DNA. One interestUsher syndrome type 1B (USH1B), adFEVR and Best's ing feature of the mitochondrial mutations is the A--cG disease] and two loci responsible for recessive deafness: substitution at position 1555 in the gene encoding 12S Usher syndrome type 1B (USH1B), which results in rRNA47A8. This particular change has been implicated in deafness, vestibular dysfunction and progressive enhanced sensitivity to aminoglycoside-induced cochretinitis pigmentosum46; and nonsyndromic recessive lear damage, since it may favour of the steric interaction deafness (NSRD2), which is deafness with occasional of the aminoglycoside with the ribosome by increasing 14.1
-
Mod2
6p2,~-24
/ j • •
i i i
m
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access to the ribosome cleft47. This observation illustrates how an investigation into an inherited form of deafness can provide very useful hypotheses to explain the molecular basis of environmentally triggered cochlear damage. The range of types of gene involved in causing nearing loss given in Table 1 and the fact that hearing loss occurs as a part of so many diverse syndromes in humans probably refects two aspects of the auditory system: (1) the complexity of the inner ear and the wide range of structural components involved in maintaining optimum function in detecting very small and rapidly changing movements; and (2) the susceptibility of the sensory hair cells to damage and their inability to regenerate. Middle ear defects Two genes, M s x l and endotbelin-1, have been identified by transgenic disruptions as important in the formation of the middle ear in mice49,5°. However, in neither case did the homozygous deficient mice survive for long after birth. The future Identification of some of the genes involved in auditory transduction, either by positional cloning or by cloning mammalian homologues of mechanosensory genes of lower organisms, can surely be only a few months away. When these genes are known, it will be an exciting time of progress, not only for understanding the molecular basis of auditory transduction, but also for understanding the pathological processes involved in the non-syndromic autosomal recessive deafness that is so important in the human population.
Acknowledgements We thank A, Read, C. Petit, T. Friedman and J. Cable for communicating their findings before publication, D. Hughes for comments on the manuscript, and A. Pearce and P, Moorjani for bibliographic assistance.
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3 Konigsmark, B.W. and Godin, R,J. (1976) Genetic and Metabolic Deafness, W,B. Saunders Co. 4 Fraser, G.R. (1976) The Causes of Profound Deafness in Childhood, Johns Hopkins UniversityPress 5 Deol, M.S. (1968).]. Med. Genet. 5, 137-158 6 Lyon, M.F. and Seade, A.G. (1989) Genetic Variants and Strains of the Laboratory Mouse (2nd edn), Oxford University Press 7 Nadeau, J.H., Kosowsky, M. and Steel, K.P. (1991) Ann. N. E Acad. Sct. 630, 49.-67 8 Davis, A.C. (1989) Int.J. Eptdemtol. 18, 911-917 9 Searle, A.G., Edwards, J.H. and Hall,J.G. (1994)J. Med. Genet. 31, 1-19 10 Steel, K.P. (1991) Ann. N.Y. Acad. Sci. 630, 68-79 11 Deol, M.S. (1966) Nature 209, 219--220 12 Steel, K.P. and Bock, G.R. (1983) Arch. Otolaryngoi. 109, 22-29 13 Keynes, R. and Krumlauf,R. (1994) Annu. Rev. Neurosci. 17, 109-132 14 Repressa, J., Leon, Y., Miner, C. and Giraldez, F. (1991) Nature 353, 561-563
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K.P. STEEL iS IN THE MRC INSTHtr~ OF HEARING RESr~tR~ UNIV~SHY PAR& NOITINGHA~ L/K NG7 2RD; £1~M. BROWNns US rue DF.PARrmmNrOFBmcHF.msmv XND MOLE~L&¢ GENL~C~ Sr Macy's HOSPITAL MF.DICAL Scao01~ NORFOLgPIAC~ LONOON, UK W2 IPGo
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