Abst rac ts 45

I-9 ANNA FOGDELL AND OLLE OLERUP

Center for BioTechnology, Karolinska Institute, NOVUM, S-141 57 Huddinge, Sweden and Department of Clinical Immunology, Karolinska Institute at Huddinge Hospital, Huddinge, Sweden

The DQAI*OI04 allele is c a r r i e d by DR10 (DRBI*IOOI) and DR14 (DRBI*1401) hap lo types in Caucas i ans , b lack Afr icans and Or ien ta l s

Introduction. The DQAI*OI04 and DQAI*OIOI alleles only differ by a single base pair in the first exon. DQAI*OI04 has a guanine as the fifth nucleotide, whereas DQAI*OIOI and all other sequenced DQAl alleles has an adenine at this position.

DQAI*O104 was originally described in black Americans with the DRBI*I2,DQAI*0104,DQBI*0501; DRBI*I2,DQAI*0IO4,DQBI*0605 and DRBI*I4,DQAI*OIO4,DQBI*0503 haplotypes (i). When we dave- loped DQAI typlng by PCR amplification using sequence-speci- fic primers (PCR-SSP), we observed that all DR10- and DRI4- positive samples (including the DRBl*140I-homozygous cell line - 31227ABO) were DQAI*OI04- and not DQAl*OlOl-positive as previously described (2).

The aim of the present study was to confirm this finding in a larger number of samples representing the major differ- ent ethnic groups.

Matarials. Six cell lines of the Xth Histocompatibility Workshop (3 DR1- and 3 DRl4-positive), 32 Caucasian samples (11 DR1-, I0 DR10- and ll DRl4-positive), 18 African samples (8 DR1-, 8 DRIO- and 2 DRl4-positive), ii Japanese samples (5 DR1- and 6 DRl4-positive), and finally one Chinese DRIO- positive sample were studied.

Mathode. DR-DQ typlng was performed by TagI DRB-DQA-DQB RFLP analysis. DQAI and DQBI typings as well as DRBI*OI and DRBI*I4 subtypings were performed by PCR-SSP (2).

Rasulte. All DRIO- and DRl4-positive samples carried the DQAI*OI04 allele, whereas DRl-positive (DRBI*010I-DRBI*0103) samples carried the DQAI*OIOI allele. DR14 PCR-SSP subtyping showed that all DR14 samples were DRBl*140i-positive. The associated DQBI specificity was DQBI*0501 and DQBI*0503 in DRIO and DR14 haplotypes, respectively.

Conolusioms. In contrast to previously published data (3), the DRBI*1001 and DRBI*I401 haplotypes carry the DQAI*0104 and not the DQAl*OlOl allele in the three major ethnic groups studied. Thus, the DQ ~-~ heterodimer is different in DRBI*I001 and DRBI*OI haplotypes, DQAI*0104-DQBI*0501 and DQAI*OIOI-DQBI*0501, respectively.

Since the DQAI*0104 allele occurred in all DRBI*I001 and DRBI*I401 haplotypes in Africans, Caucasians and Orientals, the point mutation in the second codes of the DQa chain probably occurred prlor to the separation of the two haplo- types.

R e f e r ~ n c e ~ i. Lee KW e~ al. Tissue An tlgens 1991: 38: 231. 2. Olerup e= al. Tissue Antigens 1993 (~n press}. 3. Bodmer JG e= al. Nomenclature for factors of the HLA system, 1991.

Tissue ~n=igens 1992: 39: 161

1-11 A novel HLA DRBI *01 allele

*GUIGNIER F. ** MERCIER B, *ROZ P. "*FEREC C, °*SALEUN JP, "CHATELAIN P

* Centre R6glonal de Transfusion Sanguine. 8 boulevard Marechal de Lattre de Tassigny. BP 1542, 21034 - DIJON - FRANCE "* Centre Depanemental de Transfusion Sanguine, Centre de Biogenetique, 46 n~e Felix Le Dantec, 29275 - BREST - FRANCE

Up to now 3 HLA DRBI*01 alleles have been described (DRBI*0101. 0102 and 0103 ). In the present report we describe an additional novel DRB]*01 allele which was identified by oligotyping in a Caucasian patiem. Indeed the positive signal with one probe indicated the presence of a DRBI*01 allele, but negative signals with other probes eliminate the DRB 1 *0101, 0102 and 0103 alleles. Thus we decided to sequence this probable novel allele

The analysis of the sequencing data shows that the novel sequence results of a probable genie conversion between DRBI*0101 or 0102 and DRB3*0201 or 0301. Indeed the nucleotidic sequence from AA number 13 to 76 is identical to the sequence of the DRBI*0101 or 0102 alleles. From AA number 77 to 88 the sequence is identical to the sequence of the DRBI "0301 or 0301 alleles

This result was con~nned both by hybridisation with specific probes and with PCR-SSP. We also performed a family study in order to characterise the haplotyp¢. The analysis allowed us to identify the haplntype : A l l , B35, DRBI*0], DRB6, D Q B ] ' 0 f DQAI*01, DPBI'030I. This allele seems to be very rare in Caucasian population (it was identified once =Lmong 400 typings).

This work was supported by "Ligue Nationale Contre le Cancer" (Saone and Loire Comrmttee)

I-lO MARTINEZ-LASO J.*, BENMAMAR D.', VARELA P.*, MARTINEZ-QUILES N.", BELHAN1 F.@, BF.KI(HOUCHA F.$, MORALES P.', CORELL A.*. MARTIN-VILI~ JM.* AND ARNAJZ- VI LLENA A~*

• Department o[ Instructor'. Hospaa112 de L~tuDre. Unn~rmdad COmplu¢l~,¢. 28041. Madrid. Spain # Service a' Hema,,~ogse. Hoptutl Um~mm]~ de Bcm-Mcmx=. AI81¢rs. Ats©na $ Um~: d'lmmun~ogl¢. Hopaai C.¢nLral de 1' Armee. AJglers. AJgena

EVOLUTIONARY RELATIONSHIPS OF HLA-DR8 AIJ.gLES AND

DESCRIPTION OF A NEW SUBTYPE (DRBI*0806) IN THE

ALGERIAN POPULATION

A new HLA-DR8 allele found in the Algerian population (HLA-DRBI*0806) is described in the present work. This allele has an exon-2 nucleotide sequence identical to that of the HLA-DRBI*080I allele, except for a GGT to GTG change in codon 86,

yielding an aminoacid substitution (Glycine to Valine); this change is also found in

other HIA-DRB1 and HLA-DRB3 evolutionary related allele palm. HLA-DRBI=0806 is

the most frequent HLA-DR8 allele found in this population (3 out of 8 HLA-DR8 positive individuals) and is found associated within the HLA-DOAI*0102-DQBI*0602 haplotype (HLA-DO6 semtype); this combination of HLA-DO alleles is sporadically found

associated to other HLA-DR8 alleles in other ethnic groups, ie: HLA-DRBI=0804 in

Bushmen and in North American Negroids and HLA-DRBI*0803 in African Negroids.

Also, the HLA-DRBI*0806-DQ6 haplotype only bears one HLA-DRB gene copy, a

common characteristic of the HLA-DR8 haplotypes family. An gxon-2 HLA-DR8

dendmgram has also been constructed with the available sequence data. indicating that

this new allele does not seem to be placed at a distance significantly different from the origin to that of HLA-DRBI*0804 (proposed to be the eldest of HLA-DR8 allele diversification pathway). A possible HLA-DR8 evolutive pathway is postulated according to the available exon-2 HLA-DR8 allele sequences.

1-12 M G Guttridge, E A Bidwell and P T Klouda

UK Transplant Support Service, Sou~, Bristol BSI0 5hD, England

HIA-A3.3: A New Variant of HIA-A3 ~=~scted hy ID-IEF

The serological specificity A3 is ccmprlsed of t~D alleles, A*0301 and A*0302. The alleles differ by three nucleotide substitutions in exon 3 that encode am/no acid changes at positions 152 (Glu to Val) and 156 (Leu to Glu). The molecules are indistinguishable by serology, but are identified uslng cellular and biocbamical techniques. In this s~dy, a new varzant of HIA-A3, termed A3.3, is described. HIA-A3.3 w~s detected by c~e-dimensic~al isoelectric focusing (ID-IEF) in a European caucasoid mxlivi~,~1, JC, typzng by serology as A3, 30; BS, 60; Cw3, w7; DR1, 17. NO unusual serological r~ctlons identified for the A3.3 molecule. A sD]dy of ID-IEF band patterns of A3 molecules indicated that A3.3 focused clearly acidic to A*0301 and A*0302, and slightly basic to BS. HIA class I typlng by serology and ID-IEF, and class II typlng by DNA-RFLP of the donor's family showed A3.3 was on a haplotype HIA-A3.3, B8, Cw7, DR3 (w17), DQw2 (Table). In a preliminary investigatlon to d e ~ the nucleotide sequence of the /%3.3 gene, segments of HIA-A genes ~ amplified fr~n genomlc £~qA by the PCR using primers designed to amplify b~ 134-246 of exon 2 (erect/rig amLno acids 46-82), and bp 26-199 of exon 3 (encoding am/z~ acids 100-156). The PCR pro~*cts were clcmed in E. cell cells uszng the Ml3mpl8 vector and sequenced uslng the dideoxy chain termom~tlon method. The results indicated that A3.3 had a nucleotide sequence indist/nguishable from At0301 in the reglons of exon 2 and 3 ~gated. This suggests that the amino acid substitution a~ting for the differences in pl of A3.3 and A*0301 is not located between asH_no acids 46-82 or 100-156 of the HLA-A3.3 molecule.

Table: HIA Phenotypes for Family C.

Donor Relation haplorype

DC Father b

PC ~ _E_c d

MC Sibling 1 c

JC Sibling 2 b

A B C DR DQ

2.2 15.3 3 4 8 3.3 8 7 17 2

1 8 7 7 2 30.1 60 3 1 5

2.2 15.3 3 4 8 i 8 7 7 2

3.3 8 7 17 2 30.1 60 3 1 5