- Email: [email protected]
A 4 kb Fragment of the Desmocollin 3 Promoter Directs Reporter Gene Expression to Parakeratotic Epidermis and Primary Hair Follicles
LETTER TO THE EDITOR
A 4 kb Fragment of the Desmocollin 3 Promoter Directs Reporter Gene Expression to Parakeratotic Epidermis and Primary Hair Follicles Journal of Investigative Dermatology (2007) 127, 245–247. doi:10.1038/sj.jid.5700483; published online 13 July 2006
TO THE EDITOR The mechanisms by which epidermal differentiation is regulated remain obscure. Intriguingly in some situations, two different types of differentiation exist side by side. Parakeratotic epidermis is characterized by a thick, densely packed stratum corneum and the absence of a granular cell layer. In the tail, legs, and feet of mice and other rodents, a regular, highly ordered pattern of parakeratotic scale epidermis alternates with normal orthokeratotic interscale epidermis. Parakeratotic epidermis is histologically and biochemically similar to the parakeratotic scales of psoriatic epidermis, and mouse tail has been used as an experimental model for testing antipsoriatic drugs for their ability to enhance orthokeratotic conversion. Using transgenic mice, we have identified a 4 kb fragment of desmocollin (Dsc) 3 50 -flanking DNA that specifically directs expression of a b-galactosidase reporter gene to parakeratotic epidermis and to the bulb region of primary hair follicles. The construct retains some of the features of the promoter from which it was derived, but also exhibits a remarkable pattern of expression that has not been documented previously. Dsc3 is one of a family of three Dscs. The Dscs are adhesion molecules of desmosomes, and play an essential role in the maintenance of epidermal integrity (Chidgey et al., 2001). They exhibit tissue-specific patterns of expression, with Dsc2 being ubiquitously expressed in epithelial tissues, and Dsc1 and Dsc3 largely restricted to stratified epithelia. All three Dscs are expressed in the epidermis, with Dsc3 Abbreviation: Dsc, desmocollin
& 2006 The Society for Investigative Dermatology
being most strongly expressed in basal cell layers (North et al., 1996). We have previously shown that transcription factors of the CCAAT/enhancer-binding protein family are important in the regulation of Dsc3 and Dsc1 transcription in cultured keratinocytes (Smith et al., 2004). As a first step in identifying those sequences responsible for directing the tissue and stratum specificity of Dsc3, a construct containing 4 kb of the Dsc3 promoter upstream of a b-galactosidase reporter gene (Figure 1a) was microinjected into fertilized mouse eggs. Transgenic founder mice were identified by Southern blotting (Figure 1b) and two independent transgenic lines were established. Both expressed similar levels of b-galactosidase and exhibited identical staining patterns. One line, which contained 3–6 copies of the transgene at a single point of insertion, was studied in detail. First, b-galactosidase activity was analyzed in the tail epidermis of young (4 weeks) mice. Surprisingly, staining was restricted to parakeratotic scale regions and was absent from orthokeratotic interscale regions (Figure 1c and d). This unusual pattern of expression is in contrast to the normal pattern of expression exhibited by Dsc3, which is expressed in both scale and interscale epidermis (Figure 1e). In scale epidermis, b-galactosidase staining was restricted to suprabasal cells, and was absent from the basal layer (Figure 1f). In contrast, Dsc3 was detected in all cell layers of tail epidermis (Figure 1g). A similar pattern was found in the leg epidermis, with strong b-galactosidase staining restricted to parakeratotic scale regions and absent from orthokeratotic
interscale regions (Figure 1h). However, in leg scale epidermis, staining was located in all cell layers. The scale epidermis of leg has fewer cell layers than that of tail, and it may be that b-galactosidase expression in the basal layer of leg, but not tail, scale epidermis reflects a difference in parakeratotic differentiation programs. Parakeratotic scale epidermis begins to develop from normal orthokeratotic epidemis after birth and is not fully formed until the second postnatal week (Schweizer and Marks, 1977). b-Galactosidase was first detected 2 days after birth when small groups of positive cells were observed in suprabasal layers of tail epidermis (not shown). Weak b-galactosidase staining was detected in the center of newly forming scales at day 9 (Figure 1i), coincident with the first detectable loss of the granular layer (Didierjean et al., 1983). No b-galactosidase activity was found at any time in normal orthokeratinizing back skin epidermis, nor was it found in other epithelial tissues known to express Dsc3, nor was it found at any stage during the development of embryonic epidermis (not shown). An unusual pattern of b-galactosidase expression was observed in hair follicles. Dsc3 was expressed throughout the length of the hair shaft, and in the bulb cells of primary hair follicles of the tail (Figure 2a). In contrast, b-galactosidase activity was restricted to bulb cells surrounding the dermal papillae (Figure 2b). At present, it is not clear why b-galactosidase shows such a restricted pattern of expression in tail hair follicles. No b-galacosidase staining was found in hair follicles of dorsal skin. The Tabby mouse (www.informa tics.jax.org) acts as an animal model for www.jidonline.org
245
A Merritt et al. A 4 kb Fragment of the Desmocollin 3 Promoter
a
SV40 polyA
SV40 SD/SA
c
-galactosidase
Dsc3 promoter
Hind III
b
Sacl
Hind III –1
EcoRV
tg
wt
tg
tg
wt
tg
Probe sr
d
isr
sr
e
h
sr
isr
f
g
i
isr
Figure 1. b-Galactosidase is expressed in parakeratotic, but not orthokeratotic, epidermis. (a) Dsc3 50 -flanking DNA (4 kb; from HindIII to base 1) was cloned into the EcoRI and XhoI sites of pNASSb (Clontech, Basingstoke, Hampshire, UK), a promoterless eucaryotic expression vector containing an SV40 splice donor/splice acceptor (SD/SA), b-galactosidase coding sequence, and SV40 polyadenylation site. The Dsc3-bgal cassette was excised from the plasmid by cutting with KasI (partial) and SalI. DNA was recovered using the QIAEXII kit (Qiagen, Crawley, West Sussex, UK) and diluted in microinjection buffer (5 mM Tris, 0.1 mM EDTA, pH 7.4). Transgenic mice were generated as described (Merritt et al., 2002), with ethical approval from the Universities of Birmingham and Manchester, and under licence from the UK Home Office. (b) Mice carrying integrated transgene DNA were identified by Southern blotting of BamHI cut genomic DNA using an 827 bp SacI–EcoRV probe from the b-galactosidase gene. tg, transgenic; wt, wild type. (c) Mouse tail showing b-galactosidase activity in scale epidermis but not in interscale regions. Tissues from transgenic mice were fixed for 30 minutes in 0.2% glutaraldehyde, 0.1 M sodium phosphate (pH 7.3), 5 mM EGTA, and 2 mM MgCl2, washed (3 30 minutes) with 0.1 M sodium phosphate (pH 7.3), 2 mM MgCl2, 0.01% sodium deoxycholate, and 0.02% IGEPAL CA-630, and stained overnight at 371C in wash buffer with 5 mM potassium ferrocyanide, 5 mM potassium ferricyanide, and 1 mg/ml 5-bromo-4chloro-3-indolyl-b-D-galactopyranoside (X-gal). Stained tissues were washed, post-fixed in formal saline, embedded in paraffin, sectioned, and stained with eosin. Arrow indicates anterior to posterior direction. (d) Transverse section of tail showing b-galactosidase activity in scale epidermis but not in interscale regions. b-Galactosidase activity is also associated with primary hair follicles in the tail (arrowheads). Note the characteristic presence of three hair follicles associated with each scale. sr, scale region; isr, interscale region. (e) Tail epidermis showing Dsc3 native protein expression (brown immunostaining) in the epidermis. The streptavidin–biotin indirect immunoperoxidase method was used with a primary rabbit anti-mouse Dsc3 antibody (Hardman et al., 2005) at a 100-fold dilution. Antigen retrieval was performed by microwaving (3 minutes in 0.01 M citrate buffer, pH 6) and the primary antibody was conjugated with biotinylated goat anti-mouse/rabbit IgG (Dako, Ely, Cambridgeshire, UK) and streptavidin–peroxidase. Sections were stained with 3,30 -diaminobenzidine (Sigma, Poole, Dorset, UK) and counterstained with hematoxylin. Dsc3 is expressed in both scale and interscale epidermis. (f) High magnification of scale epidermis showing absence of b-galactosidase activity in the basal cell layer (arrowhead). (g) High magnification of scale epidermis showing Dsc3 expression in all cell layers, including the basal cell layer (arrowhead). (h) Expression of b-galactosidase in all cell layers of scale epidermis of leg. Staining is absent from interscale epidermis. The basal layer is indicated by an arrowhead. (i) b-Galactosidase activity in developing scales and primary hair follicles (arrowheads) of an 8-day mouse. Bar ¼ 100 mm in (d), (e), and (i) and 25 mm in (f), (g), and (h).
hypohidrotic ectodermal dysplasmia; mice lack primary hair follicles and normal tail scales as a result of a mutation in the ectodysplasin-A gene (Srivastava et al., 1997). As expected, no b-galactosidase staining was found in the tails of Dsc3-bgal Tabby crosses. Herein, we show that the sequences required for Dsc3 expression in orthoand parakeratatotic epidermis of tail can be uncoupled, as indeed can those responsible for expression in basal and suprabasal cell layers of scale epider246
mis. It appears that 4 kb of the Dsc3 promoter contains some elements responsible for tissue-specific expression of Dsc3, but not all. Similarly, 4.2 kb of 50 -flanking DNA from human DSG1 contains sufficient information to direct a lacZ reporter gene to epidermis and hair follicles, but lacks the regulatory elements required for correct expression in other tissues (Adams et al., 1998). Desmosomal cadherin genes are clustered on chromosome 18q12.1 (Hunt et al., 1999), and it is possible that additional regulatory elements,
Journal of Investigative Dermatology (2007), Volume 127
such as a locus control region, will be required in future vectors to fully recapitulate their correct expression patterns in transgenic mice. The expression pattern of Dsc3 in the epidermis shows some similarities with that of the keratins K5 and K14, which are also strongly expressed in basal cells. In the case of K5, only 90 bp of 50 -flanking DNA is required to direct expression to normal orthokeratotic epidermis, although a switch in differentiation specificity from basal to terminally differentiating cells is observed
A Merritt et al. A 4 kb Fragment of the Desmocollin 3 Promoter
a
b
Chidgey M, Brakebusch C, Gustafsson E, Cruchley A, Hail C, Kirk S et al. (2001) Mice lacking desmocollin 1 show epidermal fragility accompanied by barrier defects and abnormal differentiation. J Cell Biol 155:821–32 Didierjean L, Wrench R, Saurat JH (1983) Expression of cytoplasmic antigens linked to orthokeratosis during the development of parakeratosis in newborn mouse tail epidermis. Differentiation 23:250–5 Hardman MJ, Liu K, Avilion AA, Merritt A, Brennan K, Garrod DR et al. (2005) Desmosomal cadherin misexpression alters b-catenin stability and epidermal differentiation. Mol Cell Biol 25:969–78
Figure 2. b-Galactosidase is expressed in the bulb region of primary hair follicles. (a) Dsc3 and (b) b-galactosidase expression in the bulb of a primary hair follicle from tail. Arrowheads indicate specific Dsc3 staining at the cell membrane. Bar ¼ 10 mm.
with the truncated promoter. Interestingly, in tail reporter gene expression is present in suprabasal cells of orthokeratotic interscale regions but is entirely absent from parakeratotic scales (Byrne and Fuchs, 1993). It would appear that both the K5 and Dsc3 promoters contain elements that are responsible for pan-epidermal expression in tail; absence of these elements apparently can lead to discrimination between the two types of epidermis. Our findings present early evidence for a transgene that can be used as a marker for parakeratotic epidermis, and Dsc3-bgal mice may prove to be a useful experimental tool for studying the unique differentiation program that leads to its formation. Topical treatment of scale epidermis with retinoids leads to conversion of parakeratotic to normal orthokeratotic epidermis (Schweizer et al., 1987) and the mice may prove to be a suitable model for testing these and other potential antipsoriatic drugs.
CONFLICT OF INTEREST The authors state no conflict of interest.
ACKNOWLEDGMENTS This work was supported by the BBSRC and the Medical Research Council, UK.
Anita J. Merritt1, Kuichun Zhu2, David R. Garrod1 and Martyn A.J. Chidgey2 1 Faculty of Life Sciences, University of Manchester, Manchester, UK and 2Division of Medical Sciences, University of Birmingham, Clinical Research Block, Queen Elizabeth Hospital, Birmingham, UK. E-mail: [email protected]
REFERENCES Adams MJ, Reichel MB, King IA, Marsden MD, Greenwood MD, Thirlwell H et al. (1998) Characterization of the regulatory regions in the human desmoglein genes encoding the pemphigus foliaceous and pemphigus vulgaris antigens. Biochem J 329:165–74 Byrne C, Fuchs E (1993) Probing keratinocyte and differentiation specificity of the human K5 promoter in vitro and in transgenic mice. Mol Cell Biol 13:3176–90
Hunt DM, Sahota VK, Taylor K, Simrak D, Hornigold N, Arnemann J et al. (1999) Clustered cadherin genes: a sequence-ready contig for the desmosomal cadherin locus on human chromosome 18. Genomics 62:445–55 Merritt AJ, Berika MY, Zhai W, Kirk SE, Ji B, Hardman MJ et al. (2002) Suprabasal desmoglein 3 expression in the epidermis of transgenic mice results in hyperproliferation and abnormal differentiation. Mol Cell Biol 22:5846–58 North AJ, Chidgey MAJ, Clarke JP, Bardsley WG, Garrod DR (1996) Distinct desmocollin isoforms occur in the same desmosomes and show reciprocally graded distributions in bovine nasal epidermis. Proc Natl Acad Sci USA 93:7701–5 Schweizer J, Furstenberger G, Winter H (1987) Selective suppression of two postnatally acquired 70 kD and 65 kD keratin proteins during continuous treatment of adult mouse tail epidermis with vitamin A. J Invest Dermatol 89:125–31 Schweizer J, Marks F (1977) A developmental study of the distribution and frequency of Langerhans cells in relation to formation of patterning in mouse tail epidermis. J Invest Dermatol 69:198–204 Smith C, Zhu K, Merritt A, Picton R, Youngs D, Garrod D et al. (2004) Regulation of desmocollin gene expression in the epidermis: CCAAT/enhancer-binding proteins modulate early and late events in keratinocyte differentiation. Biochem J 380:757–65 Srivastava AK, Pispa J, Hartung AJ, Du Y, Ezer S, Jenks T et al. (1997) The Tabby phenotype is caused by mutation in a mouse homologue of the EDA gene that reveals novel mouse and human exons and encodes a protein (ectodysplasin-A) with collagenous domains. Proc Natl Acad Sci USA 94:13069–74
www.jidonline.org
247
A 4 kb Fragment of the Desmocollin 3 Promoter Directs Reporter Gene Expression to Parakeratotic Epidermis and Primary Hair Follicles Journal of Investigative Dermatology (2007) 127, 245–247. doi:10.1038/sj.jid.5700483; published online 13 July 2006
TO THE EDITOR The mechanisms by which epidermal differentiation is regulated remain obscure. Intriguingly in some situations, two different types of differentiation exist side by side. Parakeratotic epidermis is characterized by a thick, densely packed stratum corneum and the absence of a granular cell layer. In the tail, legs, and feet of mice and other rodents, a regular, highly ordered pattern of parakeratotic scale epidermis alternates with normal orthokeratotic interscale epidermis. Parakeratotic epidermis is histologically and biochemically similar to the parakeratotic scales of psoriatic epidermis, and mouse tail has been used as an experimental model for testing antipsoriatic drugs for their ability to enhance orthokeratotic conversion. Using transgenic mice, we have identified a 4 kb fragment of desmocollin (Dsc) 3 50 -flanking DNA that specifically directs expression of a b-galactosidase reporter gene to parakeratotic epidermis and to the bulb region of primary hair follicles. The construct retains some of the features of the promoter from which it was derived, but also exhibits a remarkable pattern of expression that has not been documented previously. Dsc3 is one of a family of three Dscs. The Dscs are adhesion molecules of desmosomes, and play an essential role in the maintenance of epidermal integrity (Chidgey et al., 2001). They exhibit tissue-specific patterns of expression, with Dsc2 being ubiquitously expressed in epithelial tissues, and Dsc1 and Dsc3 largely restricted to stratified epithelia. All three Dscs are expressed in the epidermis, with Dsc3 Abbreviation: Dsc, desmocollin
& 2006 The Society for Investigative Dermatology
being most strongly expressed in basal cell layers (North et al., 1996). We have previously shown that transcription factors of the CCAAT/enhancer-binding protein family are important in the regulation of Dsc3 and Dsc1 transcription in cultured keratinocytes (Smith et al., 2004). As a first step in identifying those sequences responsible for directing the tissue and stratum specificity of Dsc3, a construct containing 4 kb of the Dsc3 promoter upstream of a b-galactosidase reporter gene (Figure 1a) was microinjected into fertilized mouse eggs. Transgenic founder mice were identified by Southern blotting (Figure 1b) and two independent transgenic lines were established. Both expressed similar levels of b-galactosidase and exhibited identical staining patterns. One line, which contained 3–6 copies of the transgene at a single point of insertion, was studied in detail. First, b-galactosidase activity was analyzed in the tail epidermis of young (4 weeks) mice. Surprisingly, staining was restricted to parakeratotic scale regions and was absent from orthokeratotic interscale regions (Figure 1c and d). This unusual pattern of expression is in contrast to the normal pattern of expression exhibited by Dsc3, which is expressed in both scale and interscale epidermis (Figure 1e). In scale epidermis, b-galactosidase staining was restricted to suprabasal cells, and was absent from the basal layer (Figure 1f). In contrast, Dsc3 was detected in all cell layers of tail epidermis (Figure 1g). A similar pattern was found in the leg epidermis, with strong b-galactosidase staining restricted to parakeratotic scale regions and absent from orthokeratotic
interscale regions (Figure 1h). However, in leg scale epidermis, staining was located in all cell layers. The scale epidermis of leg has fewer cell layers than that of tail, and it may be that b-galactosidase expression in the basal layer of leg, but not tail, scale epidermis reflects a difference in parakeratotic differentiation programs. Parakeratotic scale epidermis begins to develop from normal orthokeratotic epidemis after birth and is not fully formed until the second postnatal week (Schweizer and Marks, 1977). b-Galactosidase was first detected 2 days after birth when small groups of positive cells were observed in suprabasal layers of tail epidermis (not shown). Weak b-galactosidase staining was detected in the center of newly forming scales at day 9 (Figure 1i), coincident with the first detectable loss of the granular layer (Didierjean et al., 1983). No b-galactosidase activity was found at any time in normal orthokeratinizing back skin epidermis, nor was it found in other epithelial tissues known to express Dsc3, nor was it found at any stage during the development of embryonic epidermis (not shown). An unusual pattern of b-galactosidase expression was observed in hair follicles. Dsc3 was expressed throughout the length of the hair shaft, and in the bulb cells of primary hair follicles of the tail (Figure 2a). In contrast, b-galactosidase activity was restricted to bulb cells surrounding the dermal papillae (Figure 2b). At present, it is not clear why b-galactosidase shows such a restricted pattern of expression in tail hair follicles. No b-galacosidase staining was found in hair follicles of dorsal skin. The Tabby mouse (www.informa tics.jax.org) acts as an animal model for www.jidonline.org
245
A Merritt et al. A 4 kb Fragment of the Desmocollin 3 Promoter
a
SV40 polyA
SV40 SD/SA
c
-galactosidase
Dsc3 promoter
Hind III
b
Sacl
Hind III –1
EcoRV
tg
wt
tg
tg
wt
tg
Probe sr
d
isr
sr
e
h
sr
isr
f
g
i
isr
Figure 1. b-Galactosidase is expressed in parakeratotic, but not orthokeratotic, epidermis. (a) Dsc3 50 -flanking DNA (4 kb; from HindIII to base 1) was cloned into the EcoRI and XhoI sites of pNASSb (Clontech, Basingstoke, Hampshire, UK), a promoterless eucaryotic expression vector containing an SV40 splice donor/splice acceptor (SD/SA), b-galactosidase coding sequence, and SV40 polyadenylation site. The Dsc3-bgal cassette was excised from the plasmid by cutting with KasI (partial) and SalI. DNA was recovered using the QIAEXII kit (Qiagen, Crawley, West Sussex, UK) and diluted in microinjection buffer (5 mM Tris, 0.1 mM EDTA, pH 7.4). Transgenic mice were generated as described (Merritt et al., 2002), with ethical approval from the Universities of Birmingham and Manchester, and under licence from the UK Home Office. (b) Mice carrying integrated transgene DNA were identified by Southern blotting of BamHI cut genomic DNA using an 827 bp SacI–EcoRV probe from the b-galactosidase gene. tg, transgenic; wt, wild type. (c) Mouse tail showing b-galactosidase activity in scale epidermis but not in interscale regions. Tissues from transgenic mice were fixed for 30 minutes in 0.2% glutaraldehyde, 0.1 M sodium phosphate (pH 7.3), 5 mM EGTA, and 2 mM MgCl2, washed (3 30 minutes) with 0.1 M sodium phosphate (pH 7.3), 2 mM MgCl2, 0.01% sodium deoxycholate, and 0.02% IGEPAL CA-630, and stained overnight at 371C in wash buffer with 5 mM potassium ferrocyanide, 5 mM potassium ferricyanide, and 1 mg/ml 5-bromo-4chloro-3-indolyl-b-D-galactopyranoside (X-gal). Stained tissues were washed, post-fixed in formal saline, embedded in paraffin, sectioned, and stained with eosin. Arrow indicates anterior to posterior direction. (d) Transverse section of tail showing b-galactosidase activity in scale epidermis but not in interscale regions. b-Galactosidase activity is also associated with primary hair follicles in the tail (arrowheads). Note the characteristic presence of three hair follicles associated with each scale. sr, scale region; isr, interscale region. (e) Tail epidermis showing Dsc3 native protein expression (brown immunostaining) in the epidermis. The streptavidin–biotin indirect immunoperoxidase method was used with a primary rabbit anti-mouse Dsc3 antibody (Hardman et al., 2005) at a 100-fold dilution. Antigen retrieval was performed by microwaving (3 minutes in 0.01 M citrate buffer, pH 6) and the primary antibody was conjugated with biotinylated goat anti-mouse/rabbit IgG (Dako, Ely, Cambridgeshire, UK) and streptavidin–peroxidase. Sections were stained with 3,30 -diaminobenzidine (Sigma, Poole, Dorset, UK) and counterstained with hematoxylin. Dsc3 is expressed in both scale and interscale epidermis. (f) High magnification of scale epidermis showing absence of b-galactosidase activity in the basal cell layer (arrowhead). (g) High magnification of scale epidermis showing Dsc3 expression in all cell layers, including the basal cell layer (arrowhead). (h) Expression of b-galactosidase in all cell layers of scale epidermis of leg. Staining is absent from interscale epidermis. The basal layer is indicated by an arrowhead. (i) b-Galactosidase activity in developing scales and primary hair follicles (arrowheads) of an 8-day mouse. Bar ¼ 100 mm in (d), (e), and (i) and 25 mm in (f), (g), and (h).
hypohidrotic ectodermal dysplasmia; mice lack primary hair follicles and normal tail scales as a result of a mutation in the ectodysplasin-A gene (Srivastava et al., 1997). As expected, no b-galactosidase staining was found in the tails of Dsc3-bgal Tabby crosses. Herein, we show that the sequences required for Dsc3 expression in orthoand parakeratatotic epidermis of tail can be uncoupled, as indeed can those responsible for expression in basal and suprabasal cell layers of scale epider246
mis. It appears that 4 kb of the Dsc3 promoter contains some elements responsible for tissue-specific expression of Dsc3, but not all. Similarly, 4.2 kb of 50 -flanking DNA from human DSG1 contains sufficient information to direct a lacZ reporter gene to epidermis and hair follicles, but lacks the regulatory elements required for correct expression in other tissues (Adams et al., 1998). Desmosomal cadherin genes are clustered on chromosome 18q12.1 (Hunt et al., 1999), and it is possible that additional regulatory elements,
Journal of Investigative Dermatology (2007), Volume 127
such as a locus control region, will be required in future vectors to fully recapitulate their correct expression patterns in transgenic mice. The expression pattern of Dsc3 in the epidermis shows some similarities with that of the keratins K5 and K14, which are also strongly expressed in basal cells. In the case of K5, only 90 bp of 50 -flanking DNA is required to direct expression to normal orthokeratotic epidermis, although a switch in differentiation specificity from basal to terminally differentiating cells is observed
A Merritt et al. A 4 kb Fragment of the Desmocollin 3 Promoter
a
b
Chidgey M, Brakebusch C, Gustafsson E, Cruchley A, Hail C, Kirk S et al. (2001) Mice lacking desmocollin 1 show epidermal fragility accompanied by barrier defects and abnormal differentiation. J Cell Biol 155:821–32 Didierjean L, Wrench R, Saurat JH (1983) Expression of cytoplasmic antigens linked to orthokeratosis during the development of parakeratosis in newborn mouse tail epidermis. Differentiation 23:250–5 Hardman MJ, Liu K, Avilion AA, Merritt A, Brennan K, Garrod DR et al. (2005) Desmosomal cadherin misexpression alters b-catenin stability and epidermal differentiation. Mol Cell Biol 25:969–78
Figure 2. b-Galactosidase is expressed in the bulb region of primary hair follicles. (a) Dsc3 and (b) b-galactosidase expression in the bulb of a primary hair follicle from tail. Arrowheads indicate specific Dsc3 staining at the cell membrane. Bar ¼ 10 mm.
with the truncated promoter. Interestingly, in tail reporter gene expression is present in suprabasal cells of orthokeratotic interscale regions but is entirely absent from parakeratotic scales (Byrne and Fuchs, 1993). It would appear that both the K5 and Dsc3 promoters contain elements that are responsible for pan-epidermal expression in tail; absence of these elements apparently can lead to discrimination between the two types of epidermis. Our findings present early evidence for a transgene that can be used as a marker for parakeratotic epidermis, and Dsc3-bgal mice may prove to be a useful experimental tool for studying the unique differentiation program that leads to its formation. Topical treatment of scale epidermis with retinoids leads to conversion of parakeratotic to normal orthokeratotic epidermis (Schweizer et al., 1987) and the mice may prove to be a suitable model for testing these and other potential antipsoriatic drugs.
CONFLICT OF INTEREST The authors state no conflict of interest.
ACKNOWLEDGMENTS This work was supported by the BBSRC and the Medical Research Council, UK.
Anita J. Merritt1, Kuichun Zhu2, David R. Garrod1 and Martyn A.J. Chidgey2 1 Faculty of Life Sciences, University of Manchester, Manchester, UK and 2Division of Medical Sciences, University of Birmingham, Clinical Research Block, Queen Elizabeth Hospital, Birmingham, UK. E-mail: [email protected]
REFERENCES Adams MJ, Reichel MB, King IA, Marsden MD, Greenwood MD, Thirlwell H et al. (1998) Characterization of the regulatory regions in the human desmoglein genes encoding the pemphigus foliaceous and pemphigus vulgaris antigens. Biochem J 329:165–74 Byrne C, Fuchs E (1993) Probing keratinocyte and differentiation specificity of the human K5 promoter in vitro and in transgenic mice. Mol Cell Biol 13:3176–90
Hunt DM, Sahota VK, Taylor K, Simrak D, Hornigold N, Arnemann J et al. (1999) Clustered cadherin genes: a sequence-ready contig for the desmosomal cadherin locus on human chromosome 18. Genomics 62:445–55 Merritt AJ, Berika MY, Zhai W, Kirk SE, Ji B, Hardman MJ et al. (2002) Suprabasal desmoglein 3 expression in the epidermis of transgenic mice results in hyperproliferation and abnormal differentiation. Mol Cell Biol 22:5846–58 North AJ, Chidgey MAJ, Clarke JP, Bardsley WG, Garrod DR (1996) Distinct desmocollin isoforms occur in the same desmosomes and show reciprocally graded distributions in bovine nasal epidermis. Proc Natl Acad Sci USA 93:7701–5 Schweizer J, Furstenberger G, Winter H (1987) Selective suppression of two postnatally acquired 70 kD and 65 kD keratin proteins during continuous treatment of adult mouse tail epidermis with vitamin A. J Invest Dermatol 89:125–31 Schweizer J, Marks F (1977) A developmental study of the distribution and frequency of Langerhans cells in relation to formation of patterning in mouse tail epidermis. J Invest Dermatol 69:198–204 Smith C, Zhu K, Merritt A, Picton R, Youngs D, Garrod D et al. (2004) Regulation of desmocollin gene expression in the epidermis: CCAAT/enhancer-binding proteins modulate early and late events in keratinocyte differentiation. Biochem J 380:757–65 Srivastava AK, Pispa J, Hartung AJ, Du Y, Ezer S, Jenks T et al. (1997) The Tabby phenotype is caused by mutation in a mouse homologue of the EDA gene that reveals novel mouse and human exons and encodes a protein (ectodysplasin-A) with collagenous domains. Proc Natl Acad Sci USA 94:13069–74
www.jidonline.org
247