- Email: [email protected]
A novel HLA DRB1∗01 allele
Abstracts
45
I-9
I-lO
ANNA FOGDELL AND OLLE OLERUP C e n t e r for B i o T e c h n o l o g y , K a r o l i n s k a Institute, NOVUM, S-141 57 Huddinge, S w e d e n and D e p a r t m e n t of C l i n i c a l Immunology, K a r o l i n s k a I n s t i t u t e at H u d d i n g e Hospital, Huddinge, S w e d e n
DQAI*OI04 allele is c a r r i e d by DR10 (DRBI*IOOI) a n d DR14 (DRBI*1401) h a p l o t y p e s in C a u c a s i a n s , b l a c k A f r i c a n s a n d O r i e n t a l s
The
Introduction. T h e DQAI*OI04 and DQAI*OIOI a l l e l e s only d i f f e r by a s i n g l e b a s e pair in the first exon. DQAI*OI04 has a g u a n i n e as the fifth n u c l e o t i d e , w h e r e a s DQAI*OIOI and all o t h e r s e q u e n c e d DQAl a l l e l e s has an a d e n i n e at this position. DQAI*O104 was o r i g i n a l l y d e s c r i b e d in black A m e r i c a n s w i t h the D R B I * I 2 , D Q A I * 0 1 0 4 , D Q B I * 0 5 0 1 ; DRBI*I2,DQAI*0IO4,DQBI*0605 and D R B I * I 4 , D Q A I * O I O 4 , D Q B I * 0 5 0 3 h a p l o t y p e s (i). When we daveloped DQAI t y p l n g by PCR a m p l i f i c a t i o n using s e q u e n c e - s p e c i fic p r i m e r s (PCR-SSP), we o b s e r v e d that all DR10- and DRI4positive samples ( i n c l u d i n g the DRBl*140I-homozygous cell line - 31227ABO) w e r e DQAI*OI04- and not DQAl*OlOl-positive as p r e v i o u s l y d e s c r i b e d (2). The aim of the p r e s e n t s t u d y was to c o n f i r m this f i n d i n g in a l a r g e r n u m b e r of s a m p l e s r e p r e s e n t i n g the major d i f f e r ent e t h n i c groups. M a t a r i a l s . Six cell lines of the Xth H i s t o c o m p a t i b i l i t y W o r k s h o p (3 DR1- and 3 D R l 4 - p o s i t i v e ) , 32 C a u c a s i a n s a m p l e s (11 DR1-, I0 D R 1 0 - and ll D R l 4 - p o s i t i v e ) , 18 A f r i c a n s a m p l e s (8 DR1-, 8 D R I O - and 2 D R l 4 - p o s i t i v e ) , ii J a p a n e s e s a m p l e s (5 DR1- and 6 D R l 4 - p o s i t i v e ) , and f i n a l l y one C h i n e s e DRIOp o s i t i v e s a m p l e w e r e studied. Mathode. D R - D Q t y p l n g w a s p e r f o r m e d by TagI DRB-DQA-DQB R F L P analysis. DQAI and DQBI t y p i n g s as well as DRBI*OI and DRBI*I4 s u b t y p i n g s w e r e p e r f o r m e d by P C R - S S P (2). R a s u l t e . All DRIO- and D R l 4 - p o s i t i v e s a m p l e s c a r r i e d the DQAI*OI04 allele, w h e r e a s D R l - p o s i t i v e (DRBI*010I-DRBI*0103) s a m p l e s c a r r i e d the DQAI*OIOI allele. DR14 P C R - S S P s u b t y p i n g s h o w e d that all DR14 s a m p l e s were DRBl*140i-positive. The a s s o c i a t e d DQBI s p e c i f i c i t y was DQBI*0501 and DQBI*0503 in D R I O and DR14 h a p l o t y p e s , r e s p e c t i v e l y . C o n o l u s i o m s . In c o n t r a s t to p r e v i o u s l y p u b l i s h e d data (3), the DRBI*1001 and DRBI*I401 h a p l o t y p e s carry the DQAI*0104 and not the DQAl*OlOl a l l e l e in the t h r e e major e t h n i c g r o u p s studied. Thus, the DQ ~-~ h e t e r o d i m e r is d i f f e r e n t in DRBI*I001 and DRBI*OI h a p l o t y p e s , DQAI*0104-DQBI*0501 and DQAI*OIOI-DQBI*0501, r e s p e c t i v e l y . Since the DQAI*0104 a l l e l e o c c u r r e d in all DRBI*I001 and DRBI*I401 h a p l o t y p e s in A f r i c a n s , C a u c a s i a n s and Orientals, the p o i n t m u t a t i o n in the s e c o n d codes of the DQa chain p r o b a b l y o c c u r r e d p r l o r to the s e p a r a t i o n of the two h a p l o types. Refer~nce~
i. Lee KW 2. Olerup 3. Bodmer Tissue
e~ al. Tissue An tlgens 1991: 38: 231. e= al. Tissue Antigens 1993 (~n press}. JG e= al. Nomenclature for factors of the HLA system, 1991. ~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 o f 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 =Lmong400 typings).
MARTINEZ-LASO J.*, BENMAMAR D.', VARELA P.*, MARTINEZ-QUILES N.", BELHAN1 F.@,
BF.KI(HOUCHA F.$, MORALES P.', CORELL A.*. MARTIN-VILI~ JM.* AND ARNAJZVILLENA A~*
• Department o[ Instructor'. Hospaa112de L~tuDre. Unn~rmdadCOmplu¢l~,¢.28041. Madrid.Spain # Servicea' Hema,,~ogse.HoptutlUm~mm]~ de Bcm-Mcmx=. AI81¢rs.Ats©na $ Um~:d'lmmun~ogl¢.Hopaai C.¢nLralde 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 G G T to G T G 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 Also, the HLA-DRBI*0806-DQ6 haplotype common characteristic of the HLA-DR8 dendmgram has also been constructed with
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, S o u ~ ,
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 a r e 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
DC
Father
PC
~
MC
Sibling 1
C
DR
DQ
b
2.2 3.3
15.3 8
3 7
4 17
8 2
_E_c d
1 30.1
8 60
7 3
7 1
2 5
15.3 8
3 7
4 7
8 2
8 60
7 3
17 1
2 5
haplorype
c
This work was supported by "Ligue Nationale Contre le Cancer" (Saone and Loire Comrmttee)
and HLA-DRBI*0803 in African Negroids. only bears one HLA-DRB gene copy, a haplotypes family. An gxon-2 HLA-DR8 the available sequence data. indicating that
JC
Sibling 2
b
A
2.2 i 3.3 30.1
B
45
I-9
I-lO
ANNA FOGDELL AND OLLE OLERUP C e n t e r for B i o T e c h n o l o g y , K a r o l i n s k a Institute, NOVUM, S-141 57 Huddinge, S w e d e n and D e p a r t m e n t of C l i n i c a l Immunology, K a r o l i n s k a I n s t i t u t e at H u d d i n g e Hospital, Huddinge, S w e d e n
DQAI*OI04 allele is c a r r i e d by DR10 (DRBI*IOOI) a n d DR14 (DRBI*1401) h a p l o t y p e s in C a u c a s i a n s , b l a c k A f r i c a n s a n d O r i e n t a l s
The
Introduction. T h e DQAI*OI04 and DQAI*OIOI a l l e l e s only d i f f e r by a s i n g l e b a s e pair in the first exon. DQAI*OI04 has a g u a n i n e as the fifth n u c l e o t i d e , w h e r e a s DQAI*OIOI and all o t h e r s e q u e n c e d DQAl a l l e l e s has an a d e n i n e at this position. DQAI*O104 was o r i g i n a l l y d e s c r i b e d in black A m e r i c a n s w i t h the D R B I * I 2 , D Q A I * 0 1 0 4 , D Q B I * 0 5 0 1 ; DRBI*I2,DQAI*0IO4,DQBI*0605 and D R B I * I 4 , D Q A I * O I O 4 , D Q B I * 0 5 0 3 h a p l o t y p e s (i). When we daveloped DQAI t y p l n g by PCR a m p l i f i c a t i o n using s e q u e n c e - s p e c i fic p r i m e r s (PCR-SSP), we o b s e r v e d that all DR10- and DRI4positive samples ( i n c l u d i n g the DRBl*140I-homozygous cell line - 31227ABO) w e r e DQAI*OI04- and not DQAl*OlOl-positive as p r e v i o u s l y d e s c r i b e d (2). The aim of the p r e s e n t s t u d y was to c o n f i r m this f i n d i n g in a l a r g e r n u m b e r of s a m p l e s r e p r e s e n t i n g the major d i f f e r ent e t h n i c groups. M a t a r i a l s . Six cell lines of the Xth H i s t o c o m p a t i b i l i t y W o r k s h o p (3 DR1- and 3 D R l 4 - p o s i t i v e ) , 32 C a u c a s i a n s a m p l e s (11 DR1-, I0 D R 1 0 - and ll D R l 4 - p o s i t i v e ) , 18 A f r i c a n s a m p l e s (8 DR1-, 8 D R I O - and 2 D R l 4 - p o s i t i v e ) , ii J a p a n e s e s a m p l e s (5 DR1- and 6 D R l 4 - p o s i t i v e ) , and f i n a l l y one C h i n e s e DRIOp o s i t i v e s a m p l e w e r e studied. Mathode. D R - D Q t y p l n g w a s p e r f o r m e d by TagI DRB-DQA-DQB R F L P analysis. DQAI and DQBI t y p i n g s as well as DRBI*OI and DRBI*I4 s u b t y p i n g s w e r e p e r f o r m e d by P C R - S S P (2). R a s u l t e . All DRIO- and D R l 4 - p o s i t i v e s a m p l e s c a r r i e d the DQAI*OI04 allele, w h e r e a s D R l - p o s i t i v e (DRBI*010I-DRBI*0103) s a m p l e s c a r r i e d the DQAI*OIOI allele. DR14 P C R - S S P s u b t y p i n g s h o w e d that all DR14 s a m p l e s were DRBl*140i-positive. The a s s o c i a t e d DQBI s p e c i f i c i t y was DQBI*0501 and DQBI*0503 in D R I O and DR14 h a p l o t y p e s , r e s p e c t i v e l y . C o n o l u s i o m s . In c o n t r a s t to p r e v i o u s l y p u b l i s h e d data (3), the DRBI*1001 and DRBI*I401 h a p l o t y p e s carry the DQAI*0104 and not the DQAl*OlOl a l l e l e in the t h r e e major e t h n i c g r o u p s studied. Thus, the DQ ~-~ h e t e r o d i m e r is d i f f e r e n t in DRBI*I001 and DRBI*OI h a p l o t y p e s , DQAI*0104-DQBI*0501 and DQAI*OIOI-DQBI*0501, r e s p e c t i v e l y . Since the DQAI*0104 a l l e l e o c c u r r e d in all DRBI*I001 and DRBI*I401 h a p l o t y p e s in A f r i c a n s , C a u c a s i a n s and Orientals, the p o i n t m u t a t i o n in the s e c o n d codes of the DQa chain p r o b a b l y o c c u r r e d p r l o r to the s e p a r a t i o n of the two h a p l o types. Refer~nce~
i. Lee KW 2. Olerup 3. Bodmer Tissue
e~ al. Tissue An tlgens 1991: 38: 231. e= al. Tissue Antigens 1993 (~n press}. JG e= al. Nomenclature for factors of the HLA system, 1991. ~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 o f 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 =Lmong400 typings).
MARTINEZ-LASO J.*, BENMAMAR D.', VARELA P.*, MARTINEZ-QUILES N.", BELHAN1 F.@,
BF.KI(HOUCHA F.$, MORALES P.', CORELL A.*. MARTIN-VILI~ JM.* AND ARNAJZVILLENA A~*
• Department o[ Instructor'. Hospaa112de L~tuDre. Unn~rmdadCOmplu¢l~,¢.28041. Madrid.Spain # Servicea' Hema,,~ogse.HoptutlUm~mm]~ de Bcm-Mcmx=. AI81¢rs.Ats©na $ Um~:d'lmmun~ogl¢.Hopaai C.¢nLralde 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 G G T to G T G 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 Also, the HLA-DRBI*0806-DQ6 haplotype common characteristic of the HLA-DR8 dendmgram has also been constructed with
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, S o u ~ ,
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 a r e 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
DC
Father
PC
~
MC
Sibling 1
C
DR
DQ
b
2.2 3.3
15.3 8
3 7
4 17
8 2
_E_c d
1 30.1
8 60
7 3
7 1
2 5
15.3 8
3 7
4 7
8 2
8 60
7 3
17 1
2 5
haplorype
c
This work was supported by "Ligue Nationale Contre le Cancer" (Saone and Loire Comrmttee)
and HLA-DRBI*0803 in African Negroids. only bears one HLA-DRB gene copy, a haplotypes family. An gxon-2 HLA-DR8 the available sequence data. indicating that
JC
Sibling 2
b
A
2.2 i 3.3 30.1
B