- Email: [email protected]
The immunogenetics of insulin-dependent (Type I) diabetes
like peptides of malignancy. ogy 1988; 123:2709.
Endocrinol-
Suv;a LJ, Wirrxh
Rice BF, Pouthier RL, Miller MC: Hypercalcemia and neoplasia: a model system. Endocrinology 1971; 88:1210. Rodan SB, Insogna KL, Vignery AMC, et al.: Factors associated with humoral hypercalcemia of malignancy stimulate adenylate cyclase in osteoblastic cells. J Clin Invest 1983; 72:lSll. Rodda CP, Kubota M, Heath JA, et al.: Evidence for a novel parathyroid hormonerelated protein in fetal lamb parathyroid glands and sheep placenta: comparisons with a similar protein implicated in humoral hypercalcemia of malignancy. J Endocrinol 1988; 117:261. Simpson EL, Mundy GR, D’Souza SM, Ibbotson KJ, Bockman R, Jacobs JW: Absence of parathyroid hormone messenger RNA in nonparathyroid tumors associated with hypercalcemia. N Engl J Med 1983; 309:325. Stewart AF, Horst R, Deftos LJ, Cadman EC, Land R, Broadus AE: Biochemical evaluation of patients with cancer-associated hypercalcemia. Evidence for humoral and nonhumoral groups. N Engl J Med 1980; 303:1377. Stewart AF, Vignery A, Silvergate A, et al.: Quantitative bone histomorphometry in humoral hypercalcemia of malignancy: uncoupling of bone cell activity. J Clin Endocrinol Metab 1982; 55:219. Stewart AF, Insogna KL, Goltzman D, Broadus AE: Identification of adenylate cyclase-stimulating activity and cytochemical glucose-6-phosphate dehydrogenase-stimulating activity in extracts of tumors from patients with humoral hypercalcemia of malignancy. Proc Nat1 Acad Sci USA 1983; 80:1454. Stewart AF, Mangin M, Wu T, et al.: Synthetic human parathyroid hormone-like protein stimulates bone resorption and causes hypercalcemia in rats. J Clin Invest 1988; 81:596. Stewart AF, Wu T, Goumas D, Burtis WJ, Broadus AE: N-terminal amino acid sequence of two novel tumor-derived adenylate cyclase-stimulating proteins: identification of parathyroid hormone-like and hormone-unlike domains. parathyroid Biochem Biophys Res Commun 1987; 1461672. Strewler GJ, Williams RD, Nissenson RA: Human renal carcinoma cells produce hypercalcemia in the nude mouse and a novel protein recognized by parathyroid hormone receptors. J Clin Invest 1983; 711769. Strewler GJ, Stern PH, Jacobs JW, et al.: Parathyroid hormone-like protein from human renal carcinoma cells. Structural and functional homology with parathyroid hormone. J Clin Invest 1987; 80:1803.
44
0 1989.
GA/\, Wcttcnhall
RI%,
ct
al.: A parathyroid hormone-related protein implicated in malignant hypercalccmia: cloning and expression. Science 1987; 237:893. Thiede MA, Strewler GJ, Nissenson RA, Rosenblatt M, Rodan GA: Human renal carcinoma expresses two messages encoding a parathyroid hormone-like peptide: evidence for the alternative splicing of a single-copy gene. Proc Nat1 Acad Sci USA 1988; 85:4605. Thiede MA, Rodan GA: Expression of a calcium-mobilizing parathyroid hormonelike peptide in lactating mammary tissue. Science 1988; 242:278. Thorikay M, Kramer S, Reynolds FH, et al.: Synthesis of a gene-encoding parathyroid hormone-like protein (1-141): purification and biological characterization of the cx-
pressed 124:lil.
protein.
Endocrinology
I YXY:
Yasuda T, Banville D, Hcndy GN, Goltzman D: Characterization of the human parathyroid hormone-like peptide gene. Functional and evolutionary aspects. J Biol Chem 1989; 26417720. Yasuda T, Banville D, Rabbani SA, Hendv GN, Goltzman D: Rat parathyroid hormone-like peptide: Comparison with the human homologue and expression in malignant and normal tissue. Mel Endocrinol 1989; 3:518. Yates AJP, Guttierrez GE, Smolens P, et al.: Effects of a synthetic peptide of a parathyroid hormone-related protein on calcium homeostasis, renal tubular calcium reabsorption, and bone metabolism in viva and in vitro in rodents. J Clin Invest 1988; 81~932.
The Immunogenetics of InsulinDependent (Type I) Diabetes J.A. Fletcher and A.H. Barn&t
Insulin-dependent diabetes develops when a genetically predisposed individual is exposed to an as-yet-unknowfl environmental insult. A major part of‘genetic susceptibility to the disorder is encoded close to or within the HLA-DQ region, but non-HLA-linked genes are also implicated.
major forms of diabetes are recognized. The first creates an absolute dependence by (Type I) patients on exogenous insulin for survival; in the second; most compliant noninsulin-dependent (Type II) subjects will respond to dietary measures, alone or in combination with oral hypoglycemic drugs. Insulin-dependent diabetes (IDDM) is thought to have an autoimmune pathogenesis; evidence for this includes the finding, at diagnosis, of infiltration of the islets of Langerhans with inflammatory cells, circulating islet cell-specilic antibodies, and alterations in circulating T cell subsets (reviewed in Vergani 1987 and Wilkin 1987). In contrast, there is no evidence for autoimTwo
munity as a causative factor in noninsulin-dependent diabetes (NIDDM). Confirmation of the existence of an important genetic component of the etiology of IDDM came from studies in the mid-1970s that established that certain HLA antigens are increased in frequency in insulin-dependent diabetics compared to the healthy nondiabetic population (see below). The lack of HLA associations in noninsulin-dependent subjects (Nerup et al. 1974) confirmed the distinct etiology of IDDM and NIDDM. Studies of diabetic identical twins (Tattersall and Pyke 1972) have, however, suggested the existence of an environmental agent that triggers IDDM in genetically predisposed individuals. ??
J.A. Fletcher and A.H. Barnett are at the Department of Medicine, University of Birmingham and East Birmingham Hospital, Birmingham, B15 2TH, UK.
Elscvier
Science
Publishing
Co., Inc.
1043.2760/89/$3.50
HLA
and IDDM
The HLA system, or major histocompatibility complex (MHC), of man is located on the short arm of chromo-
TEM SrpterdwrlOctohev
Class
DP
Class
II
DQ
DR
Class
III
C2K4
c
Figure 1. The HLA region
on short arm of chromosome 6. The class 1 (HLA-A, -B and X), class II (HLA-DP, -DQ and -DR) and class III (complement) gents are shown. The circic at the left of the figure rcprcsents the ccntromcre.
some 6. Within this region are encoded three principal classes of molecule (figure 1): Class I molecules arc further into subclasses subdivided three (HLA-A, -B and -C), as are class II molecules (HLA-DP, -DQ and -DR). Class III genes include those encoding three complement proteins (C2, C4, and factor B). The HLA genes arc characterized by variable amounts of polymorphism, i.e., the existence in a population of multiple peptide products derived from a single genetic locus. Relations between the specific variants (alleles) found on a given chromosome arc nonrandom. For example, in Caucasian populations, the alleles HLA-DR3 and -B8 tend to cooccur on the same chromosome at a far greater frequency than could be due to chance alone. This phenomenon, known as “linkage disequilibrium,” occurs throughout most of the HLA region, resulting in well-recognized combinations of alleles at the class I-III genes; these tend to occur together on the same chromosome (haplotypes). Early studies of HLA and diabetes employed serological and cellular immune-typing methods. In Caucasian populations, positive associations with the class I antigens B8, B15, and B18 are well-recognized (reviewed in Tiwari and Terasaki 1985), but the strongest have been defined for class Many studies of Caucasian populations have shown that DR3 and DR4 are positively associated with IDDM, to the extent that over 90% of patients have either or both of these markers (Tiwari and Terasaki 1985). Weak positive associations of DRl and DRw8 have also been identified (Thomson et al. 1988). DR2 is markedly reduced in frequency in patients with IDDM, and a weak “protective” effect
I
has also been shown for DR5 (Thomson et al. 1988). The DR314 heterozygous state has consistently been shown to be associated with a higher relative risk for IDDM than has either DR313 or DR414 homozygosity; this suggests that the DR3- and DR4-associated diseasesusceptibility alleles are different (Rotter et al. 1983). Although DR3 and DR4 are found with great frequency in diabetics, they are also found in approximately 50% of healthy Caucasian subjects. This percentage is markedly higher than that of the IDDM-susceptibility genotype in the population, which suggests that DR3 and DR4 do not correspond directly to the “diabetogenic genes.” Recent research on diabetes immunogenetics has thus attempted to define subsets of DR3- and DR4-bearing chromosomes with more specific (i.e., stronger) associations with IDDM, in the hope that analysis of the HLA alleles present on these haplotypes will identify primary-susceptibility elements for the disease.
reactivity with a DQ-specific monoclonal antibody (TAlO); only the TAlOnegative subset (now known as DQw8) is positively associated with IDDM (Tait and Boyle 1986), whereas DR4DQw7 (TAlO-positive) is reduced in frequency in IDDM. Conversely, cellular typing methods have defined subsets of DR2 haplotypes that differ in their associations with IDDM (Bach et al. 1985). The use of serological and cellular immunological methods to investigate the immunogenetics of HLA-linked disorders is limited, however, by the i’ange of available typing reagents. This review will concentrate on the use of DNA studies.
??
DNA Studies
The close associations of DR antigens with IDDM have led workers in this field to concentrate on the genes of the class II region. In the early phases of the immune response, the HLA-D molecules are involved in presenting processed foreign antigen to T helper cells, and they also play an important role in inducing self-tolerance during T cell development. Structurally, they are transmembrane glycopeptides consisting of a 34,000-Da a! chain and a 29,000Da p chain, which are noncovalently associated (figure 2). The class II HLA genetic region is shown in more detail in figure 3. The genes encoding class II HLA (r and p peptides are now designated “A” and “B,” respectively.
associations
II markers.
TEM Srpren~hc~lOctohet
A
??
More Specific Markers for IDDM
The search for more specific markers for IDDM susceptibility has involved several experimental approaches. For example, DR4 haplotypes have been subdivided into two subsets according to
0 1989,Elscvicr
Sciencr
Publishing
Cu., Inc.
1043.2760/891$3.50
Early DNA studies of HLA polymorphism and IDDM used the technique of restriction-fragment-length polymorphism (RFLP) analysis (Southern 1975). This uses restriction endonuclease enzymes to cut DNA at specific nucleotide sequences. After separation by agarose gel electrophoresis, the DNA fragments are transferred and bound, in singlestranded form, to a nylon membrane by capillary (Southern 1975) blotting. Radio-labeled, single-stranded DNA cor-
4s
of DR3 is inconclusive,
Figure 2. The structure of the HLA class II molecule and its transmembrane position are shown in schematic form. The two extracellular domains of the cy chain (crl and cu2) and of the p chain (01 and p2) are shown. The intracellular portions of the OLand /3 chains are represented by zig-zag lines.
responding to the gene of interest (“gene probe”) is then used to detect gene sequences on the nylon membrane. Many authors have used this approach to compare class II HLA DNA sequences in patients with IDDM with those in control subjects. The most from studies
consistent findings derive that used a DQB probe
(Owerbach et al. 1983, 1984; CohenHaguenauer et al. 1985; Bohme et al. 1986; Festenstein et al. 1986; Nepom et al. 1986; Bruserud et al. 1987; Henson et al. 1987). These showed that DR4bearing chromosomes are associated with one of two DQB RFLP patterns in healthy individuals. Approximately 70% of DR4-positive control subjects possess one pattern, which is DQw8related, while the remaining the DQw7 pattern. Among
30% have Caucasian
insulin-dependent diabetics, however, DQw8 is found in over 90% of DR4-positive subjects. DQw7 is present in the remainder of DR4-positive patients, but is significantly less common than in the healthy population. A similar analysis
since
only
determining
one
DZ
‘6a0d
a
IDDM. This suggests that the DR2related IDDM-protective factor is found only on the DR2-DQw6 chromosome. Naturally, these RFLP studies have
lotype (DRA is nonpolymorphic), but that the DQA sequence was that usually found on the Caucasian DR4 haplotype. The implication of both the DQA and DQB genes as predisposing factors for IDDM provides a possible explanation for the apparent interaction between the DR3- and DR4-related components of susceptibility. Nepom et al. (1987) have shown that, in DR314 heterozygous IDDM patients, “hvbrid” DQ hcterodimers consisting of the (DQw2)-cu
had the effect of focusing attention on the DQ subregion, particularly the DQB gene, as a potential primary-susceptibility determinant for IDDM. This possible role of DQB was supported by a recent study of class II HLA DNA sequences from IDDM-associated chromosomes (Todd et al. 1987). DQB sequences from several haplotypes with signilicant negative associations with IDDM were found to share the common characteristic of encoding aspartic acid at amino acid position 57 of the HLADOB peptidc chain (Asp-57). Todd et al. (1987) speculated that the amino acid at position 57 determines a critical function of the DQ molecule in IDDM. According to this hypothesis, full susceptibility to IDDM would be conferred by two Asp-57.negative DQB alleles, whereas the possession of two Asp-57positive DQB alleles would provide almost complete resistance to IDDM. There arc certain exceptions, however-the Caucasian DR7-DQw2 haplotype is Asp-57-negative, but does not predispose to IDDM; conversely, an Asp-57-positive haplotype found in the Japanese (DR4-DQw4) shows a positive association with IDDM (Aparicio et al. 1988). These exceptions suggest that if the DQ molecule is directly involved in
46
DO DX I3
other-amino
acid residues, including those on the DQu chain, must also be involved. Recently, we have lound direct evidence for the involvement 01 DQA as a component ofsusceptibility (Todd et al. 1989). In this study, class II HLA alleles from a Black DR7 haplotype, which is unique in showing a significant positive association with IDDM, were sequenced. It was shown that the DRB and DQB DNA sequences were the same as those found previously on the nonIDDM-associated Caucasian DR7 hap-
Figure 3. The class II HLA region. The genes encoding the 01 and /3 chains of the class II molecules are shown. Under the new nomenclature, the genes encoding the 01 and /3 chains are designated “A” and “B”, respectively.
DP
susceptibility,
DQB RFLP pattern (DQw2) is found in the great majority of DR3-positive diabetic patients and control subjects. DQB RFLP analysis can also resolve DR2 chromosomes into two subsets that differ in their associations with IDDM (Bohme et al. 1986; Cohen ct al. 1986). The negative association of DR2 with IDDM is due to a subset, DQw6, that is markedly reduced in frequency in IDDM patients, whereas the other subset (DQw5) has a neutral, or even association with slightly positive,
DQ
Rot ncc
chain from the DR3 haplotype and the (DQw8)-p chain from the DR4 haplotype can be formed. Such hybrid molecules might account for the relatively high risk for IDDM conferred by DR314 hcterozygosity. ??
Non-HLA
Susceptibility
There is growing evidence that an HLAlinked predisposition does not fully explain the genetic basis of IDDM (Niven and Hitman 1987). Several non-HLAlinked loci, including the immunoglobulin heavy chain genes (reviewed in Field and McArthur 1987), T cell receptor genes (Hoover et al. 1986), and the insulin gene (reviewed in Niven and Hitman 1987), have been proposed as factors contributing to IDDM predisposition. The evidence, however, is conflicting, and no conclusion can yet be drawn. ??
IDDM in 1989
An enormous effort has been made to understand the etiology of IDDM. However, it is fair to say that none 01
DR ‘BI RII 0111a’
8 1989, Elsrvicr Science Publishing Co., Inc. 1043.2760/8Y/$3.50
TEM SeptemheriOc_~oher
RG: The genetics of to insulin-dependent dia-
the central questions regarding the causation of IDDM has received a definitive answer. We still do not know the relative contributions of environmental and
Field
genetic
Fletcher J, Mijovic C, Odugbcsan 0, Jenkins D, Bradwell AR, Barn&t AH: Trans.racial studies implicate HLA-DQ as a component of genetic susceptibility to Type 1 (insulin-dependent) diabetes. Diabetologia 1988; 31:864.
factors,
involved, and DQB dates for but other involved. emphasize
the
number
of genes
nor their identity. The DQA genes are promising candisusceptibility determinants, loci within HLA may also be Regarding future research, we the following points:
(1) The most direct
typing methods yield the most useful data: the USC of the polymerase chain reaction (Saiki et al. 1985) to amplify DNA for direct sequencing and lor hybridization studies seems likely to supersede older, immunologically based techniques. 01 the HLA-linked (2) A full understanding component of IDDM susceptibility will probably depend on the complete mapping of the HLA region and the identification 01 all the genes therein. of the immunogenetics of (3) The analvsis IDDM in several racial groups will help to distinguish disease associations that are secondary to linkage disequilibrium from those of primary pathogcnetic importance (Fletcher st al. 1988).
References Aparicio JMR, Wakisaka A, Takada A, Matsuura N. Aizawa M: HLA-DQ system and insulin-dependent diabctcs mellitus in Japanese: does it contribute to the development of IDDM as it does in Caucasians? Immunogenetics 1988; 28:240.
LL,
McArthur
susceptibility
betes mellitus-possible Clin Invest Med 1987;
new 10:437.
markers.
Henson V, Maclaren N, Riley W, Wakeland Eli: Polymorphisms of DQB genes in HLADR4 haplotypes from healthy and diabetic individuals. Immunogenetics 1987; 25: 152. Hoover ML, Angelini G, Ball E ct al.: HLADQ and T-cell receptor genes in insulindependent diabetes mcllitus. Cold Spring Harbor Symp Quant Biol 1986; 51:803. Nepom BS, Palmer J, Kim SJ, Hansen JA, Holbeck SL, Nepom GT: Specific genomic markers for the HLA-DQ subregion discriminate between DR4’ insulin-dependent diabetes mellitus and DR4’ seropositivc juvenile rheumatoid arthritis. J Exp Med 1986; 164:345. Nepom BS, Schwartz D, Palmer JP, Nepom GT: HLA-DQaand P-chains produce hybrid molecules in DR314 hetcroxygotcs. Diabetes 1987; 36:114. Nerup J, Platz P, Anderson antigens and diabetes 1974; 21864.
00, et al.: HLAmcllitus. Lancet
Niven MJ, Hitman GA: Non-HLA associations in type 1 (insulin-dependent) diabetes mellitus, i,l Barnett AH (cd): Immunogenetics of Insulin-Dependent Diabetes. Lancaster, England, MTP Press, 1987.
Bach FH, Rich SS, Barbom J, Sefall M: InsLllirl-dcpcndcrlt cliabetcs-aasociatc~l HLA-D region encoded dctcrminants. Hum Immunol IYR5; t 2:3Y
Owerbach D, Lernmark A, Platz P. et al.: HLA-D region P-chain DNA endonuclease fragments differ between HLA-DR identical healthy and insulin-dependent diabetic individuals. Nature 1983; 303:815.
Bohmc J, Carlsson B. Wallin J, ct al.: Only one DO-p restriction lragment pattern of each DR specilicity is associated with insulin-dependent diabetes. J Immunol 1986; 137:941_
Owerbach D, Hagglof B, Lernmark A, Molmgren G: Susceptibility to insulindependent diabetes detined by restriction enzyme polymorphism of HLA-D region genomic DNA. Diabetes 1984; 33:958.
Bruserud 0, Paulsen G, Markussen G, Lundin K, Thoreacn AB, Thorsby E: Genomic HLA-DQP polymorphism associated with insulin-dependent mellitus. diabetes Stand J Immunol 1987; 25:235.
Rotter
JI, Anderson
Cohen N, Brautbar C, Font M-P, Dausset J, Cohen D: HLA-DRZ-associated Dw sublypes correlate with RFLP clusters: most DR2 IDDM patients belong to one of these clusters. Immunogenetics 1986; 23:84.
Festenstein H, Awad J. Hitman GA et al.: New DNA polymorphisms associated with autoimmune diseases. Nature 1986; 322: 64.
0 IYSY,
JE, Con&ton
betes Saiki
1983; 32:169. RK. Scharf
S, Faloona
F, et al.: Enzy-
matic amplification of /!-globin genomic sequences and restriction site analysis for diagnosis of sickle cell anemia. Science 1985; 230: 1350. Southern EM: Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol 1975; 98503. Tait BD, Boyle AJ: DR4 and susceptibility to type I diabetes mellitus: discrimination of high risk and low risk DR4 haplotypes on the basis of TAlO typing. Tissue Antigens 1987; 28:65. Tattersall twins. Thomson
RB, Pyke DA: Diabetes Lancet 1972; 2: 1120. G, Robinson
WP,
Elsevic~
Scienw
Publishing
Co.. Inc.
1043.2760/8Y/$3.50
in identical
Kuhner
MK, et
al.: Genetic heterogeneity, modes of inheritance, and risk estimates for a joint study of caucasians with insulin-dependent diabetes mellitus. Am J Hum Gcnet 1988; 43:799. Tiwari JL, Terasaki PI: Juvenile diabetes mellitus (insulin-dependent). HLA and Disease Associations. Berlin-Heidelberg, Springer-Verlag. 1985; p 185. Todd JA, Belt Jl, McDe\itt HO: gene contributes to susceptibility sistance to insulin-dependent mellitus. Nature 1987; 329:599.
HLA-DQB and rediabetes
Todd JA, Mijovic C, Fletcher J, Jenkins D, Bradwcll AR, Barnctt AH: Identifcation of susceptibility loci lor insulin-dependent diabetes metlitus by trans-racial gene mapping. Nature 1989; 338:587. Vergani
D: Cell mediated
immunity,
nett AH (ed): Immunogenetics Dependent Diabetes. Lancaster, MTP Press, 1987.
in Barof InsulinEngland,
Wilkin T: Autoimmunity and autoantibodies in type 1 (insulin-dependent) diabetes.
in
Barnett AH (cd): Immunogenetics of Insulin-Dependent Diabetes. Lancaster, England, MTP Press, 1987. TEM
Reprints of articles in TEM are available (minimum order: 100). Please contact the TEM Editorial Office: 655 Avenue of the Americas New York, New York 10010 TEL. (2 12)633-3922 FAX (2 12)633-39 13
Cohen-Haguenauer 0, Robbins E, Massart C ct al.: A systematic study 01 class 11-p DNA restriction fragments in insulin-dependent diabetes mellitus. Proc Nat1 Acad Sci USA 1985; 82:3335.
TEM Septew~herlOctohr~
CE, Rubin
PI, Terasaki PI, Rimoin DL: HLA genotypic study of insulin-dependent diabetes. The excess of DR314 heterozygotes allows rejection of the recessive hypothesis. Dia-
Endocrinol-
Suv;a LJ, Wirrxh
Rice BF, Pouthier RL, Miller MC: Hypercalcemia and neoplasia: a model system. Endocrinology 1971; 88:1210. Rodan SB, Insogna KL, Vignery AMC, et al.: Factors associated with humoral hypercalcemia of malignancy stimulate adenylate cyclase in osteoblastic cells. J Clin Invest 1983; 72:lSll. Rodda CP, Kubota M, Heath JA, et al.: Evidence for a novel parathyroid hormonerelated protein in fetal lamb parathyroid glands and sheep placenta: comparisons with a similar protein implicated in humoral hypercalcemia of malignancy. J Endocrinol 1988; 117:261. Simpson EL, Mundy GR, D’Souza SM, Ibbotson KJ, Bockman R, Jacobs JW: Absence of parathyroid hormone messenger RNA in nonparathyroid tumors associated with hypercalcemia. N Engl J Med 1983; 309:325. Stewart AF, Horst R, Deftos LJ, Cadman EC, Land R, Broadus AE: Biochemical evaluation of patients with cancer-associated hypercalcemia. Evidence for humoral and nonhumoral groups. N Engl J Med 1980; 303:1377. Stewart AF, Vignery A, Silvergate A, et al.: Quantitative bone histomorphometry in humoral hypercalcemia of malignancy: uncoupling of bone cell activity. J Clin Endocrinol Metab 1982; 55:219. Stewart AF, Insogna KL, Goltzman D, Broadus AE: Identification of adenylate cyclase-stimulating activity and cytochemical glucose-6-phosphate dehydrogenase-stimulating activity in extracts of tumors from patients with humoral hypercalcemia of malignancy. Proc Nat1 Acad Sci USA 1983; 80:1454. Stewart AF, Mangin M, Wu T, et al.: Synthetic human parathyroid hormone-like protein stimulates bone resorption and causes hypercalcemia in rats. J Clin Invest 1988; 81:596. Stewart AF, Wu T, Goumas D, Burtis WJ, Broadus AE: N-terminal amino acid sequence of two novel tumor-derived adenylate cyclase-stimulating proteins: identification of parathyroid hormone-like and hormone-unlike domains. parathyroid Biochem Biophys Res Commun 1987; 1461672. Strewler GJ, Williams RD, Nissenson RA: Human renal carcinoma cells produce hypercalcemia in the nude mouse and a novel protein recognized by parathyroid hormone receptors. J Clin Invest 1983; 711769. Strewler GJ, Stern PH, Jacobs JW, et al.: Parathyroid hormone-like protein from human renal carcinoma cells. Structural and functional homology with parathyroid hormone. J Clin Invest 1987; 80:1803.
44
0 1989.
GA/\, Wcttcnhall
RI%,
ct
al.: A parathyroid hormone-related protein implicated in malignant hypercalccmia: cloning and expression. Science 1987; 237:893. Thiede MA, Strewler GJ, Nissenson RA, Rosenblatt M, Rodan GA: Human renal carcinoma expresses two messages encoding a parathyroid hormone-like peptide: evidence for the alternative splicing of a single-copy gene. Proc Nat1 Acad Sci USA 1988; 85:4605. Thiede MA, Rodan GA: Expression of a calcium-mobilizing parathyroid hormonelike peptide in lactating mammary tissue. Science 1988; 242:278. Thorikay M, Kramer S, Reynolds FH, et al.: Synthesis of a gene-encoding parathyroid hormone-like protein (1-141): purification and biological characterization of the cx-
pressed 124:lil.
protein.
Endocrinology
I YXY:
Yasuda T, Banville D, Hcndy GN, Goltzman D: Characterization of the human parathyroid hormone-like peptide gene. Functional and evolutionary aspects. J Biol Chem 1989; 26417720. Yasuda T, Banville D, Rabbani SA, Hendv GN, Goltzman D: Rat parathyroid hormone-like peptide: Comparison with the human homologue and expression in malignant and normal tissue. Mel Endocrinol 1989; 3:518. Yates AJP, Guttierrez GE, Smolens P, et al.: Effects of a synthetic peptide of a parathyroid hormone-related protein on calcium homeostasis, renal tubular calcium reabsorption, and bone metabolism in viva and in vitro in rodents. J Clin Invest 1988; 81~932.
The Immunogenetics of InsulinDependent (Type I) Diabetes J.A. Fletcher and A.H. Barn&t
Insulin-dependent diabetes develops when a genetically predisposed individual is exposed to an as-yet-unknowfl environmental insult. A major part of‘genetic susceptibility to the disorder is encoded close to or within the HLA-DQ region, but non-HLA-linked genes are also implicated.
major forms of diabetes are recognized. The first creates an absolute dependence by (Type I) patients on exogenous insulin for survival; in the second; most compliant noninsulin-dependent (Type II) subjects will respond to dietary measures, alone or in combination with oral hypoglycemic drugs. Insulin-dependent diabetes (IDDM) is thought to have an autoimmune pathogenesis; evidence for this includes the finding, at diagnosis, of infiltration of the islets of Langerhans with inflammatory cells, circulating islet cell-specilic antibodies, and alterations in circulating T cell subsets (reviewed in Vergani 1987 and Wilkin 1987). In contrast, there is no evidence for autoimTwo
munity as a causative factor in noninsulin-dependent diabetes (NIDDM). Confirmation of the existence of an important genetic component of the etiology of IDDM came from studies in the mid-1970s that established that certain HLA antigens are increased in frequency in insulin-dependent diabetics compared to the healthy nondiabetic population (see below). The lack of HLA associations in noninsulin-dependent subjects (Nerup et al. 1974) confirmed the distinct etiology of IDDM and NIDDM. Studies of diabetic identical twins (Tattersall and Pyke 1972) have, however, suggested the existence of an environmental agent that triggers IDDM in genetically predisposed individuals. ??
J.A. Fletcher and A.H. Barnett are at the Department of Medicine, University of Birmingham and East Birmingham Hospital, Birmingham, B15 2TH, UK.
Elscvier
Science
Publishing
Co., Inc.
1043.2760/89/$3.50
HLA
and IDDM
The HLA system, or major histocompatibility complex (MHC), of man is located on the short arm of chromo-
TEM SrpterdwrlOctohev
Class
DP
Class
II
DQ
DR
Class
III
C2K4
c
Figure 1. The HLA region
on short arm of chromosome 6. The class 1 (HLA-A, -B and X), class II (HLA-DP, -DQ and -DR) and class III (complement) gents are shown. The circic at the left of the figure rcprcsents the ccntromcre.
some 6. Within this region are encoded three principal classes of molecule (figure 1): Class I molecules arc further into subclasses subdivided three (HLA-A, -B and -C), as are class II molecules (HLA-DP, -DQ and -DR). Class III genes include those encoding three complement proteins (C2, C4, and factor B). The HLA genes arc characterized by variable amounts of polymorphism, i.e., the existence in a population of multiple peptide products derived from a single genetic locus. Relations between the specific variants (alleles) found on a given chromosome arc nonrandom. For example, in Caucasian populations, the alleles HLA-DR3 and -B8 tend to cooccur on the same chromosome at a far greater frequency than could be due to chance alone. This phenomenon, known as “linkage disequilibrium,” occurs throughout most of the HLA region, resulting in well-recognized combinations of alleles at the class I-III genes; these tend to occur together on the same chromosome (haplotypes). Early studies of HLA and diabetes employed serological and cellular immune-typing methods. In Caucasian populations, positive associations with the class I antigens B8, B15, and B18 are well-recognized (reviewed in Tiwari and Terasaki 1985), but the strongest have been defined for class Many studies of Caucasian populations have shown that DR3 and DR4 are positively associated with IDDM, to the extent that over 90% of patients have either or both of these markers (Tiwari and Terasaki 1985). Weak positive associations of DRl and DRw8 have also been identified (Thomson et al. 1988). DR2 is markedly reduced in frequency in patients with IDDM, and a weak “protective” effect
I
has also been shown for DR5 (Thomson et al. 1988). The DR314 heterozygous state has consistently been shown to be associated with a higher relative risk for IDDM than has either DR313 or DR414 homozygosity; this suggests that the DR3- and DR4-associated diseasesusceptibility alleles are different (Rotter et al. 1983). Although DR3 and DR4 are found with great frequency in diabetics, they are also found in approximately 50% of healthy Caucasian subjects. This percentage is markedly higher than that of the IDDM-susceptibility genotype in the population, which suggests that DR3 and DR4 do not correspond directly to the “diabetogenic genes.” Recent research on diabetes immunogenetics has thus attempted to define subsets of DR3- and DR4-bearing chromosomes with more specific (i.e., stronger) associations with IDDM, in the hope that analysis of the HLA alleles present on these haplotypes will identify primary-susceptibility elements for the disease.
reactivity with a DQ-specific monoclonal antibody (TAlO); only the TAlOnegative subset (now known as DQw8) is positively associated with IDDM (Tait and Boyle 1986), whereas DR4DQw7 (TAlO-positive) is reduced in frequency in IDDM. Conversely, cellular typing methods have defined subsets of DR2 haplotypes that differ in their associations with IDDM (Bach et al. 1985). The use of serological and cellular immunological methods to investigate the immunogenetics of HLA-linked disorders is limited, however, by the i’ange of available typing reagents. This review will concentrate on the use of DNA studies.
??
DNA Studies
The close associations of DR antigens with IDDM have led workers in this field to concentrate on the genes of the class II region. In the early phases of the immune response, the HLA-D molecules are involved in presenting processed foreign antigen to T helper cells, and they also play an important role in inducing self-tolerance during T cell development. Structurally, they are transmembrane glycopeptides consisting of a 34,000-Da a! chain and a 29,000Da p chain, which are noncovalently associated (figure 2). The class II HLA genetic region is shown in more detail in figure 3. The genes encoding class II HLA (r and p peptides are now designated “A” and “B,” respectively.
associations
II markers.
TEM Srpren~hc~lOctohet
A
??
More Specific Markers for IDDM
The search for more specific markers for IDDM susceptibility has involved several experimental approaches. For example, DR4 haplotypes have been subdivided into two subsets according to
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Early DNA studies of HLA polymorphism and IDDM used the technique of restriction-fragment-length polymorphism (RFLP) analysis (Southern 1975). This uses restriction endonuclease enzymes to cut DNA at specific nucleotide sequences. After separation by agarose gel electrophoresis, the DNA fragments are transferred and bound, in singlestranded form, to a nylon membrane by capillary (Southern 1975) blotting. Radio-labeled, single-stranded DNA cor-
4s
of DR3 is inconclusive,
Figure 2. The structure of the HLA class II molecule and its transmembrane position are shown in schematic form. The two extracellular domains of the cy chain (crl and cu2) and of the p chain (01 and p2) are shown. The intracellular portions of the OLand /3 chains are represented by zig-zag lines.
responding to the gene of interest (“gene probe”) is then used to detect gene sequences on the nylon membrane. Many authors have used this approach to compare class II HLA DNA sequences in patients with IDDM with those in control subjects. The most from studies
consistent findings derive that used a DQB probe
(Owerbach et al. 1983, 1984; CohenHaguenauer et al. 1985; Bohme et al. 1986; Festenstein et al. 1986; Nepom et al. 1986; Bruserud et al. 1987; Henson et al. 1987). These showed that DR4bearing chromosomes are associated with one of two DQB RFLP patterns in healthy individuals. Approximately 70% of DR4-positive control subjects possess one pattern, which is DQw8related, while the remaining the DQw7 pattern. Among
30% have Caucasian
insulin-dependent diabetics, however, DQw8 is found in over 90% of DR4-positive subjects. DQw7 is present in the remainder of DR4-positive patients, but is significantly less common than in the healthy population. A similar analysis
since
only
determining
one
DZ
‘6a0d
a
IDDM. This suggests that the DR2related IDDM-protective factor is found only on the DR2-DQw6 chromosome. Naturally, these RFLP studies have
lotype (DRA is nonpolymorphic), but that the DQA sequence was that usually found on the Caucasian DR4 haplotype. The implication of both the DQA and DQB genes as predisposing factors for IDDM provides a possible explanation for the apparent interaction between the DR3- and DR4-related components of susceptibility. Nepom et al. (1987) have shown that, in DR314 heterozygous IDDM patients, “hvbrid” DQ hcterodimers consisting of the (DQw2)-cu
had the effect of focusing attention on the DQ subregion, particularly the DQB gene, as a potential primary-susceptibility determinant for IDDM. This possible role of DQB was supported by a recent study of class II HLA DNA sequences from IDDM-associated chromosomes (Todd et al. 1987). DQB sequences from several haplotypes with signilicant negative associations with IDDM were found to share the common characteristic of encoding aspartic acid at amino acid position 57 of the HLADOB peptidc chain (Asp-57). Todd et al. (1987) speculated that the amino acid at position 57 determines a critical function of the DQ molecule in IDDM. According to this hypothesis, full susceptibility to IDDM would be conferred by two Asp-57.negative DQB alleles, whereas the possession of two Asp-57positive DQB alleles would provide almost complete resistance to IDDM. There arc certain exceptions, however-the Caucasian DR7-DQw2 haplotype is Asp-57-negative, but does not predispose to IDDM; conversely, an Asp-57-positive haplotype found in the Japanese (DR4-DQw4) shows a positive association with IDDM (Aparicio et al. 1988). These exceptions suggest that if the DQ molecule is directly involved in
46
DO DX I3
other-amino
acid residues, including those on the DQu chain, must also be involved. Recently, we have lound direct evidence for the involvement 01 DQA as a component ofsusceptibility (Todd et al. 1989). In this study, class II HLA alleles from a Black DR7 haplotype, which is unique in showing a significant positive association with IDDM, were sequenced. It was shown that the DRB and DQB DNA sequences were the same as those found previously on the nonIDDM-associated Caucasian DR7 hap-
Figure 3. The class II HLA region. The genes encoding the 01 and /3 chains of the class II molecules are shown. Under the new nomenclature, the genes encoding the 01 and /3 chains are designated “A” and “B”, respectively.
DP
susceptibility,
DQB RFLP pattern (DQw2) is found in the great majority of DR3-positive diabetic patients and control subjects. DQB RFLP analysis can also resolve DR2 chromosomes into two subsets that differ in their associations with IDDM (Bohme et al. 1986; Cohen ct al. 1986). The negative association of DR2 with IDDM is due to a subset, DQw6, that is markedly reduced in frequency in IDDM patients, whereas the other subset (DQw5) has a neutral, or even association with slightly positive,
DQ
Rot ncc
chain from the DR3 haplotype and the (DQw8)-p chain from the DR4 haplotype can be formed. Such hybrid molecules might account for the relatively high risk for IDDM conferred by DR314 hcterozygosity. ??
Non-HLA
Susceptibility
There is growing evidence that an HLAlinked predisposition does not fully explain the genetic basis of IDDM (Niven and Hitman 1987). Several non-HLAlinked loci, including the immunoglobulin heavy chain genes (reviewed in Field and McArthur 1987), T cell receptor genes (Hoover et al. 1986), and the insulin gene (reviewed in Niven and Hitman 1987), have been proposed as factors contributing to IDDM predisposition. The evidence, however, is conflicting, and no conclusion can yet be drawn. ??
IDDM in 1989
An enormous effort has been made to understand the etiology of IDDM. However, it is fair to say that none 01
DR ‘BI RII 0111a’
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TEM SeptemheriOc_~oher
RG: The genetics of to insulin-dependent dia-
the central questions regarding the causation of IDDM has received a definitive answer. We still do not know the relative contributions of environmental and
Field
genetic
Fletcher J, Mijovic C, Odugbcsan 0, Jenkins D, Bradwell AR, Barn&t AH: Trans.racial studies implicate HLA-DQ as a component of genetic susceptibility to Type 1 (insulin-dependent) diabetes. Diabetologia 1988; 31:864.
factors,
involved, and DQB dates for but other involved. emphasize
the
number
of genes
nor their identity. The DQA genes are promising candisusceptibility determinants, loci within HLA may also be Regarding future research, we the following points:
(1) The most direct
typing methods yield the most useful data: the USC of the polymerase chain reaction (Saiki et al. 1985) to amplify DNA for direct sequencing and lor hybridization studies seems likely to supersede older, immunologically based techniques. 01 the HLA-linked (2) A full understanding component of IDDM susceptibility will probably depend on the complete mapping of the HLA region and the identification 01 all the genes therein. of the immunogenetics of (3) The analvsis IDDM in several racial groups will help to distinguish disease associations that are secondary to linkage disequilibrium from those of primary pathogcnetic importance (Fletcher st al. 1988).
References Aparicio JMR, Wakisaka A, Takada A, Matsuura N. Aizawa M: HLA-DQ system and insulin-dependent diabctcs mellitus in Japanese: does it contribute to the development of IDDM as it does in Caucasians? Immunogenetics 1988; 28:240.
LL,
McArthur
susceptibility
betes mellitus-possible Clin Invest Med 1987;
new 10:437.
markers.
Henson V, Maclaren N, Riley W, Wakeland Eli: Polymorphisms of DQB genes in HLADR4 haplotypes from healthy and diabetic individuals. Immunogenetics 1987; 25: 152. Hoover ML, Angelini G, Ball E ct al.: HLADQ and T-cell receptor genes in insulindependent diabetes mcllitus. Cold Spring Harbor Symp Quant Biol 1986; 51:803. Nepom BS, Palmer J, Kim SJ, Hansen JA, Holbeck SL, Nepom GT: Specific genomic markers for the HLA-DQ subregion discriminate between DR4’ insulin-dependent diabetes mellitus and DR4’ seropositivc juvenile rheumatoid arthritis. J Exp Med 1986; 164:345. Nepom BS, Schwartz D, Palmer JP, Nepom GT: HLA-DQaand P-chains produce hybrid molecules in DR314 hetcroxygotcs. Diabetes 1987; 36:114. Nerup J, Platz P, Anderson antigens and diabetes 1974; 21864.
00, et al.: HLAmcllitus. Lancet
Niven MJ, Hitman GA: Non-HLA associations in type 1 (insulin-dependent) diabetes mellitus, i,l Barnett AH (cd): Immunogenetics of Insulin-Dependent Diabetes. Lancaster, England, MTP Press, 1987.
Bach FH, Rich SS, Barbom J, Sefall M: InsLllirl-dcpcndcrlt cliabetcs-aasociatc~l HLA-D region encoded dctcrminants. Hum Immunol IYR5; t 2:3Y
Owerbach D, Lernmark A, Platz P. et al.: HLA-D region P-chain DNA endonuclease fragments differ between HLA-DR identical healthy and insulin-dependent diabetic individuals. Nature 1983; 303:815.
Bohmc J, Carlsson B. Wallin J, ct al.: Only one DO-p restriction lragment pattern of each DR specilicity is associated with insulin-dependent diabetes. J Immunol 1986; 137:941_
Owerbach D, Hagglof B, Lernmark A, Molmgren G: Susceptibility to insulindependent diabetes detined by restriction enzyme polymorphism of HLA-D region genomic DNA. Diabetes 1984; 33:958.
Bruserud 0, Paulsen G, Markussen G, Lundin K, Thoreacn AB, Thorsby E: Genomic HLA-DQP polymorphism associated with insulin-dependent mellitus. diabetes Stand J Immunol 1987; 25:235.
Rotter
JI, Anderson
Cohen N, Brautbar C, Font M-P, Dausset J, Cohen D: HLA-DRZ-associated Dw sublypes correlate with RFLP clusters: most DR2 IDDM patients belong to one of these clusters. Immunogenetics 1986; 23:84.
Festenstein H, Awad J. Hitman GA et al.: New DNA polymorphisms associated with autoimmune diseases. Nature 1986; 322: 64.
0 IYSY,
JE, Con&ton
betes Saiki
1983; 32:169. RK. Scharf
S, Faloona
F, et al.: Enzy-
matic amplification of /!-globin genomic sequences and restriction site analysis for diagnosis of sickle cell anemia. Science 1985; 230: 1350. Southern EM: Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol 1975; 98503. Tait BD, Boyle AJ: DR4 and susceptibility to type I diabetes mellitus: discrimination of high risk and low risk DR4 haplotypes on the basis of TAlO typing. Tissue Antigens 1987; 28:65. Tattersall twins. Thomson
RB, Pyke DA: Diabetes Lancet 1972; 2: 1120. G, Robinson
WP,
Elsevic~
Scienw
Publishing
Co.. Inc.
1043.2760/8Y/$3.50
in identical
Kuhner
MK, et
al.: Genetic heterogeneity, modes of inheritance, and risk estimates for a joint study of caucasians with insulin-dependent diabetes mellitus. Am J Hum Gcnet 1988; 43:799. Tiwari JL, Terasaki PI: Juvenile diabetes mellitus (insulin-dependent). HLA and Disease Associations. Berlin-Heidelberg, Springer-Verlag. 1985; p 185. Todd JA, Belt Jl, McDe\itt HO: gene contributes to susceptibility sistance to insulin-dependent mellitus. Nature 1987; 329:599.
HLA-DQB and rediabetes
Todd JA, Mijovic C, Fletcher J, Jenkins D, Bradwcll AR, Barnctt AH: Identifcation of susceptibility loci lor insulin-dependent diabetes metlitus by trans-racial gene mapping. Nature 1989; 338:587. Vergani
D: Cell mediated
immunity,
nett AH (ed): Immunogenetics Dependent Diabetes. Lancaster, MTP Press, 1987.
in Barof InsulinEngland,
Wilkin T: Autoimmunity and autoantibodies in type 1 (insulin-dependent) diabetes.
in
Barnett AH (cd): Immunogenetics of Insulin-Dependent Diabetes. Lancaster, England, MTP Press, 1987. TEM
Reprints of articles in TEM are available (minimum order: 100). Please contact the TEM Editorial Office: 655 Avenue of the Americas New York, New York 10010 TEL. (2 12)633-3922 FAX (2 12)633-39 13
Cohen-Haguenauer 0, Robbins E, Massart C ct al.: A systematic study 01 class 11-p DNA restriction fragments in insulin-dependent diabetes mellitus. Proc Nat1 Acad Sci USA 1985; 82:3335.
TEM Septew~herlOctohr~
CE, Rubin
PI, Terasaki PI, Rimoin DL: HLA genotypic study of insulin-dependent diabetes. The excess of DR314 heterozygotes allows rejection of the recessive hypothesis. Dia-