1605779
Complement Inflamm 1990;7:261-268
© 1990 S. Karger AG, Basel 1012-8204/90/0076-0261 $2.75/0
C4 Nomenclature Statement (1990) 1,2 G. Mauffa, C.A. Alperb, R. Dawkinsc, G. Doxiadisd, C.M. Giles*, G. Hauptmann{, C. Rittnerz, P.M. Schneider* “Institut für Medizinische Mikrobiologie und Hygiene, University of Cologne, FRG; b Center for Blood Research, Boston, Mass., USA;c Department of Clinical Immunology, Royal Perth Hospital, Australia; d Institut für Immungenetik, University of Essen, FRG; 'Rheumatology Unit, Department of Medicine, Royal Postgraduate Medical School, London, UK; fCentre de Recherche en Hématologie et Immunologie, Strasbourg, France; «Institut für Rechtsmedizin, University of Mainz, FRG
Key Words. Complement genetics • C4 • Nomenclature • Gene duplications • Non-expressed alleles • RFLP Abstract. A common and revised nomenclature of the allotypes of the fourth component (C4) of human complement has been proposed. It is based on the results of the C4 Reference Typing of the Vlth Complement Genetics Workshop and Conference, Mainz, FRG, 1989, the previous C4 nomenclature and the guidelines for human gene nomenclature (ISGN). The designation of allotypes derives from their relative electrophoretic mobility, the distinction between C4A and C4B proteins from their relative hemolytic activity. Common alleles retain their single digit numeric designation, intermediate variants their two- or three-digit desig nations; newly discovered alleles should not interfere with already described variants. At least 13 C4A alleles, 16 C4B alleles as well as non-expressed genes at each C4 locus are presently known. There are also duplicated loci of each C4 gene; they should be designated by repetition of the locus symbol at the haplotype or genotype level. As a phenotype they will be placed in parenthesis without repetition of the locus symbol. Aberrant allotypes or hybrid genes should be explained by a special suffix. No special nomenclature is recommended for restriction fragment length polymorphisms. Their designation should follow the general rules of the ISGN.
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l This statement was discussed at the Vlth Complement Genetics Workshop and Conference, Mainz, on July 29, 1989, and has been agreed upon by: M. Abbai (Toulouse), F. Christiansen (Perth), M. Cuccia-Belvedere (Pavia), E. du Toit (Cape Town), G. Geserick (Berlin), M. Hobart (Cambridge), M.L. Lokki (Helsinki), R. McLean (Baltimore, Md.), S. Nakamura (Tokyo), G.J. O’Neill (Miami, Fla.), J. Partanen (Helsinki), O.G. Segurado (Madrid), I. Siemens (Köln), K. Suzuki (Osaka), J. Taborda de Messias (Curitiba), K. Tokunaga (Tokyo), B. Uring-Lambert (Strasbourg), N. Usher (Liverpool). 2 Abbreviations used in this statement are explained on p. 174 of this issue.
Mauff/Alper/Dawkins/Doxiadis/Giles/Hauptmann/Rittner/Schneider
Introduction
A common designation for the allotypes of the fourth component (C4) of human complement was proposed in a nomencla ture statement in 1983 [1] based on an ear lier suggestion by Awdeh and Alper [2], Since then the genetic basis of C4 polymor phism has in principle been resolved al though a number of experimental findings remain unexplained. There are two kinds of C4 protein in the plasma of most individu als, C4A and C4B, with different antigenic ity [3] and numerous allotypes for each [1], C4A and C4B are encoded at two separate but closely linked loci [4] within the major histocompatibility complex (MHC) on the short arm of chromosome 6 [5]. Duplica tions and deletions or non-expressions are common for both loci [6-9]. Some individu als carry hybrid C4 genes which are thought to code for unusual, aberrant proteins, some times with reversed antigenicity. Besides progress in the definition of allo typic characteristics of the C4 proteins sub stantial knowledge has come from molecular investigations at the DNA level [see e.g. 10, 11]. Nevertheless, due to a multitude of de tectable variants of the expressed proteins as opposed to limited variability at the DNA level, being more closely related to epitopic differences of C4 [11], the nomenclature of the protein allotypes has continued to be in use over the years. Obviously, for the desig nation of phenotypes, allotypes, alleles, genes, and genotypes the present principle of C4 variant designation will remain to be the basis for communication in the foreseeable future. However, the identification of aber rant C4 proteins, recognition of duplicated C4 genes as part of defined haplotypes with other MHC genes, and an increasing number
of newly reported, in part identical, variants call for amendments in the nomenclature of C4 allotypes. Therefore, at a public nomen clature session of the Vlth Complement Ge netics Workshop and Conference on July 29, 1989, at Mainz, FRG, a revision of the present nomenclature was discussed and adopted. It was based on preworkshop refer ence typing which included the majority of known C4 variants. Detailed accounts of the reference typing will be found in other con tributions to this issue [12-16], To avoid redundancy, the following rules will only repeat the main principles of the 1983 C4 Nomenclature for better under standing and specify new or revised parts; all other unchanged details will be found in the 1983 nomenclature statement [1],
General Rules for C4 Nomenclature and Technology of C4 Typing
In principle, the revised C4 nomenclature follows the terminology of the Guidelines for Human Gene Nomenclature (ISGN, 1987) [17], and also the general rules of the 1983 nomenclature statement. Since the C4B pro tein has been associated with Chido 4 and high hemolytic activity without exception, it is therefore suggested that relative hemolytic activity in an overlay on agarose gel electro phoresis containing sheep red blood cells and C4-deficient guinea pig serum is applied to distinguish between C4A and C4B pro teins [2], C4B showing about a four times higher activity than C4A [18]. Therefore, the basic typing procedure consists of immunofixation with C4-specific antisera after sepa ration of samples in agarose gel electropho resis and simultaneous hemolytic overlay as outlined in detail in table 1 in the 1983 stateDownloaded by: University of Exeter 144.173.6.94 - 5/6/2020 5:42:21 PM
262
C4 Nomenclature Statement (1990)
263
Table 1. Recognized C4 alleles Allele designation
RM
new
previous
mean
A Q0 A8 A7 A6 A 58 A 55 A5 A 45 A4 A3 A2 A 12 A1 A 91
A Q0 A7 A7 A6 A6 A6 A5 A5 A 4/35 A3 A2 A 12/A1 A1 A 91/A90/A1/B31
silent allele 197.2 183.9 138.2 136.0 127.8 119.4 116.9 107.9 98.9 82.6 64.0 54.2 47.8
BQ0
BQ0
B6 B5 B 45 B4 B 35 B3 B 22 B2 B 13 B 12 B 11 B 1 B 92 B 94 B 95 B 96
B 6/B7 B 5/B6 B5 B4 B3 B 29/3/4 B 22 B2 B 12 B 12 B 12/A 95 B 1 B 92 B 94 B 95 B9
silent allele 72.6 67.3 59.2 46.6 43.5 39.2 33.2 29.4 12.9 9.5 7.7
Lysis type
MAB
1
A A A A A A A A A A A A A
n.d. A A A A A A A A A A/B A/B B
B B B B B B B B B B B B B B B B
B/A A/B A A/B B A/B B B A A A B B B n.d. B
range
136.3-139.4
4
127.5-128.0 116.1-121.1
2
1 4
1 1 1 1 59.6-67.8 53.6-54.8 45.3-49.8
6
72.2-73.0 62.5-70.4
2 8
46.3-47.2 42.6-44.3 37.0-42.5
3
3 6
1 2
9
1 27.6-31.2 12.7-13.2 8.7-10.0
2
3 3
1 1
0.0
-20.7 (- 40.7)a n.d. (-92.6)a
n
-19.7-21.1
2
1 1
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RM = Relative migration value (mean value if several samples were tested; 3-10 determinations per indi vidual sample), the range represents the lowest and highest individual allotype value, n = number of tested samples; MAB = monoclonal antibody reactivity anti-C4A/Rgl,2, anti-C4B/Chl [12, 13]. n.d. = not deter mined. Only estimates.
Mauff/Alper/Dawkins/Doxiadis/Giles/Hauptmann/Rittner/Schneider
ment, with the exception that before neur aminidase (NANA’se), treatment with CARB-B [ 19] should be used if the number of expressed alleles and the relative migra tion distance are to be determined. Additional techniques may be necessary or helpful to define or subtype allotypes and haplotypes, such as Western blots with poly clonal or monoclonal antibodies after pro longed agarose-gel electrophoresis, Rodgers/ Chido typing, C4 a- and ß-chain typing [9, 20-22], or DNA TaqI RFLP for the recogni tion of gene size and duplications [6].
Designation of C4 Allotypes
The allotypes are to be designated ac cording to their relative electrophoretic mo bility by increasing arabic numbers from
cathode to anode for both proteins (fig. 1,2, table 1). Allotypes with migration positions cathodal of A 1 or B 1 should be preceded by the number 9 (e.g. A 91, A 92, etc., B 91, B 92, etc.). The two most common allotypes in all ethnic groups A 3 and B 1, as well as the majority of the less common variants will retain their single-digit numeric desig nation [1], with the exception of C4B 5, B 6, and B 7. A redesignation of this region was found to be necessary since the most com mon variant ‘B6’, almost invariably associ ated with C4A 4, HLA-B55 and DR 1 or W6(14), has repeatedly been published as C4B 5. Thus, to avoid any further misun derstanding, the variant designated B 5 in the 1983 nomenclature will now become B 45, B 6 will be B 5, and B 7 will be B 6. The deleted or nonexpressed C4A will be C4A Q0, the deleted or nonexpressed C4B will be C4B Q0. Downloaded by: University of Exeter 144.173.6.94 - 5/6/2020 5:42:21 PM
264
C4 Nomenclature Statement (1990)
265
Fig. 1. C4 allotypes. Prolonged immunofixation agarose-gel electrophoresis (CARB-B and NANA’se treatment) of standard and rare allotypes submitted to the Vlth Complement Genetics Workshop Reference Typing Phenotype a
b
c
1: 3: 5: 7: 9: 11: 13:
WOO RIT MAU DUT MAU ALP ALP
14: 16: 18: 20: 22: 24:
RIT TOK ALP DOX MAU TAB
25: DUT 27: TOK
001/1
AB 006/5 002/2
015/1 004 007/2 HDW CWT2 007/1 001/1
006/4 001/7 002/2
CWT6
C4A
B
Phenotype
8 6
1 1 1
2: 4: 6: 8: 10: 12:
CUC TAB ALP MAU MAU ALP
15: 17: 19: 21: 23:
RIT DOX DAW MAU HAU
55, 2 45, 12 3 12, 91 91
(92)
1 Q0 Q0
4, 3, 2 4 91 3 4 3, 2
6 45, 2 35 35, 22, 1 5, 3, 13, 1
45,12 4
92 2, 1,96
C4A 001/1 001/2
001
037/1 006/5 006
KHH
7, 3 58, 3 5,3 4 55, 2 3, 1
B
1 3, 1
1 2
1 1 5 35, 22, 1 3, 1
002/2
4, 3, 2 3 3 4 3, 2
001/1
6, 3
1, 94
001/1
Rob 037/1
2 12, 1
11, 1
26: PAR
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★: A 91 B 35 in sample 18 could not electrophoretically be separated but were distinguished by lysis and segregation analysis. The classified allotypes are in bold face (standard separation conditions). For explanation of abbreviations see page 174 of this issue.
Mauff/Alper/Dawkins/Doxiadis/Giles/Hauptmann/Rittner/Schneider
266
--
1 90
-
180
-
170
-
160
-
1 50
-
140
-
130
-
1 20
-
110100 90 80 70 60 50 40
C4B
C4A
RM 200
8
6 58555 45'' 4
A 3
-
2
-
6 5 45-
12-
-
4
91-
3-35J 22-
-
30
-
20
-
10
-
2--- ■
B 1
13-, 12-iï=
o -10
-20 -30
-
94-,
-40 -50 -60
92-,
-
95
-70 -80
-
-90
-
-100
96
-
—
Fig. 2. Relative migration (RM) distances of C4 allotypes seen during the VIth Complement Genetics Workshop reference typing (C4A 3, B 1 with minor band positions).
they should receive a one-digit designation if they migrate outside the range of the known variants, two or three digits if they migrate between known allotypes. Allotype classifications may be estimated according to the Downloaded by: University of Exeter 144.173.6.94 - 5/6/2020 5:42:21 PM
As before, allotypes with intermediate migration between already described aliotypes will receive a two (or three) digit designation. Newly discovered allotypes should not interfere with already described variants;
C4 Nomenclature Statement (1990)
C4 Reference Variants
No provision for reference laboratories has been made for the future. But a small number of standard reference variants will be available on request for a limited period of time with the senior author. They are exclusively intended for the classification of internal laboratory standards.
Amendments
Any modifications of the present C4 no menclature which might become necessary subsequently should be discussed in public at international meetings (such as the Inter national Complement or Complement Ge netics Workshops) after prior invitation or announcement in the program.
Acknowledgements The authors gratefully acknowledge the editorial and technical assistance of Mrs. Martina Brenden, Margot Braun-Stilwell, Hilde Hasbach, Dr. Georg Plum, Gudrun Riethmeister, Beate Stradmann-Bellinghausen, and the untiring secretarial and photo graphic assistance of Mrs. Renate Thelen and Sigrid Glanschneider.
References 1 Mauff G, Alper CA, Awdeh Z, Batchelor JR, Ber trams J, et al: Statement on the nomenclature of human C4 allotypes. Immunobiology 1983; 164: 184-191. 2 Awdeh ZL, Alper CA: Inherited structural poly morphism of the fourth component of human complement. Proc Natl Acad Sci 1980;77:35763580. 3 Tilley CA, Romans DG, Crookston MC: Localiza Downloaded by: University of Exeter 144.173.6.94 - 5/6/2020 5:42:21 PM
major or minor bands after NANA’se treat ment. A more reliable classification will re sult from the combination of the measure ments of relative migration distances after CARB-B treatment, using the distance be tween C4B 1 and C4A 3 as 100 (see table 1) and electrophoretic side by side comparison; details of the method have been described in Mauff et al. [23]. Of the variants described in the 1983 no menclature statement, examples of C4A 51, 14, 13, 11, C4B 51, 31, 29, 21, 91, 93 were not submitted or seen during this workshop. C4B 31 is now considered to be identical with C4A 91, and B 29 with B 3. Others may have received new designations or positions such as C4A 51, A 91, B 51 or B 4, or possi bly belong to heterogeneous groups not yet recognized, such as A 14, A 13, 11 or B 21. Their classification will have to wait for fu ture reference typings. Duplicated loci, if written as haplotype or genotype, are expressed by repetition of the locus symbol: e.g. C4A*3 A*2; they will be placed in parentheses if written as phenotype or allotypes without repetition of the locus symbol: e.g. C4A (3, 2). Aberrant allotypes or products of hybrid genes, if discovered by additional methods, e.g. Rodgers and Chido typing, Western blot with monoclonal antibodies, C4-chain typ ing, gene size or RFLPs will not receive a special nomenclature. But if subtypes are described they should be designated and ex plained by a special suffix to the main allo type. For RFLPs a special nomenclature was not thought to be necessary for the time being. Designations should include the probe, restriction enzymes and the size of the detectable fragments and possibly their relative intensities, but otherwise follow the general rules of the ISGN [17).
267
4
5
6
7
8
9
10
11
12
13
Mauff/Alper/Dawkins/Doxiadis/Giles/Hauptmann/Rittner/Schneider
tion of Chido and Rodgers determinants to the C4d fragment of human C4. Nature 1978;276: 713. O’Neill GJ, Yang SY, Dupont B: Two HLA-linked loci controlling the fourth component of human complement. Proc Natl Acad Sci 1978;75:5165— 5169. Rittner C, Hauptmann G, Grosse-Wilde H, Grosshans E, Tongio MM, Mayer S: Linkage be tween HLA (major histocompatibility complex) and genes controlling the synthesis of the fourth component of complement; in Kissmeyer-Nielsen F (ed): Histocompatibility Testing 1975. Copen hagen, Munksgard, 1975, p 945. Schneider PM, Carroll MC, Alper CA, Rittner C, Whitehead S, et al: Polymorphism of the human complement C4 and 21 steroid hydroxylase genes. J Clin Invest 1967;78:650-657. Giles CM, Uring-Lambert B, Boksch W, Braun M, Goetz J, et al: The study of a French family with two duplicated C4A haplotypes. Hum Genet 1987;77:359-365. Yu CY, Campbell RD, Porter RR: A structural model for the location of the Rodgers and the chido antigenic determinants and their correla tion with the human complement component C4A/C4B isotypes. Immunogenetics 1988;27: 399-405. Steuer M, Mauff G, Adam C, Baur MP, Bender K, Goetz J, et al: An estimate on the frequency of duplicated haplotypes and silent alleles of human C4 protein polymorphism. I. Investigation in healthy Caucasoid families. Tissue Antigens 1989; 33:501-510. Carroll MC, Campbell RD, Bentley DR, Porter RR: A molecular map of human major histocom patibility complex class III region linking comple ment genes C4, C2 and factor B. Nature 1984;307: 237-241. Yu CY, Belt KT, Giles C, Campbell RC, Porter R: Structural basis of the polymorphism of human complement components C4A and C4B: Gene size, reactivity antigenicity. EMBO J 1986;5: 2873-2881. Doxiadis G, Grosse-Wilde H: C4 allotyping by prolonged gel electrophoresis and immunoblotting using monoclonal and polyclonal antibodies. Complement Inflamm 1990;7:269-276. Giles CM: C4: Rodgers and Chido typing. Com plement Inflamm 1990;7:213-217.
14 Mauff G, Brenden M, Braun-Stilwell M, Doxiadis G, Giles C, Hauptmann G, Rittner C, Schneider PM, Stradmann-Bellinghausen B, Uring-Lambert B: C4 reference typing report. Complement In flamm 1990;7:193-212. 15 Rittner C, Stradmann-Bellinghausen B: C4 alphachain reference typing report. Complement Inflamm 1990;7:225-229. 16 Schneider PM: C4 DNA RFLP reference typing report. Complement Inflamm 1990;7:218-224. 17 Shows TB, McAlpine PJ, Boucheix C, Collins FS, Coneally PM, Frezal J, et al: Guidelines for hu man gene nomenclature. An international system for human gene nomenclature (ISGN, 1987). Cytogenet Cell Genet 1987;46:11-28. 18 Isenman DE, Young JR: The molecular basis for the difference in immune hemolysis activity of the Chido and Rodgers isotypes of human comple ment component C4. J Immunol 1984; 132:3019— 3027. 19 Sim E, Cross SJ: Phenotyping of human comple ment C4, a class-III HLA antigen. Biochem J 1986;239:763-767. 20 Doxiadis G, Doxiadis I, Frenz G, Vogeler U, Grosse-Wilde H: Relevance of complotyping and subtyping of MHC class I gene products in haplo type definition for allogeneic bone marrow trans plantation. Bone Marrow Transplant 1989;4:17— 22.
21 Giles CM: Partial inhibition of anti-Rg and antiCh reagents. II. Demonstration of separable anti bodies for different determinants. Vox Sang 1985; 48:167-173. 22 Roos MH, Mollenhauer E, Demant P, Rittner C: A molecular basis for the two locus models of human complement component C4. Nature 1982; 298:856. 23 Mauff G, Brenden M, Braun-Stilwell M: Relative electrophoretic migration distances for the classi fication of C4 allotypes. Complement Inflamm 1990;7:277-281.
Gottfried Mauff Institut für Medizinische Mikrobiologie und Hygiene Universität zu Köln Goldenfelsstrasse 21 D-W-5000 Köln 41 (FRG)
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268
Complement Inflamm 1990;7:261-268
© 1990 S. Karger AG, Basel 1012-8204/90/0076-0261 $2.75/0
C4 Nomenclature Statement (1990) 1,2 G. Mauffa, C.A. Alperb, R. Dawkinsc, G. Doxiadisd, C.M. Giles*, G. Hauptmann{, C. Rittnerz, P.M. Schneider* “Institut für Medizinische Mikrobiologie und Hygiene, University of Cologne, FRG; b Center for Blood Research, Boston, Mass., USA;c Department of Clinical Immunology, Royal Perth Hospital, Australia; d Institut für Immungenetik, University of Essen, FRG; 'Rheumatology Unit, Department of Medicine, Royal Postgraduate Medical School, London, UK; fCentre de Recherche en Hématologie et Immunologie, Strasbourg, France; «Institut für Rechtsmedizin, University of Mainz, FRG
Key Words. Complement genetics • C4 • Nomenclature • Gene duplications • Non-expressed alleles • RFLP Abstract. A common and revised nomenclature of the allotypes of the fourth component (C4) of human complement has been proposed. It is based on the results of the C4 Reference Typing of the Vlth Complement Genetics Workshop and Conference, Mainz, FRG, 1989, the previous C4 nomenclature and the guidelines for human gene nomenclature (ISGN). The designation of allotypes derives from their relative electrophoretic mobility, the distinction between C4A and C4B proteins from their relative hemolytic activity. Common alleles retain their single digit numeric designation, intermediate variants their two- or three-digit desig nations; newly discovered alleles should not interfere with already described variants. At least 13 C4A alleles, 16 C4B alleles as well as non-expressed genes at each C4 locus are presently known. There are also duplicated loci of each C4 gene; they should be designated by repetition of the locus symbol at the haplotype or genotype level. As a phenotype they will be placed in parenthesis without repetition of the locus symbol. Aberrant allotypes or hybrid genes should be explained by a special suffix. No special nomenclature is recommended for restriction fragment length polymorphisms. Their designation should follow the general rules of the ISGN.
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l This statement was discussed at the Vlth Complement Genetics Workshop and Conference, Mainz, on July 29, 1989, and has been agreed upon by: M. Abbai (Toulouse), F. Christiansen (Perth), M. Cuccia-Belvedere (Pavia), E. du Toit (Cape Town), G. Geserick (Berlin), M. Hobart (Cambridge), M.L. Lokki (Helsinki), R. McLean (Baltimore, Md.), S. Nakamura (Tokyo), G.J. O’Neill (Miami, Fla.), J. Partanen (Helsinki), O.G. Segurado (Madrid), I. Siemens (Köln), K. Suzuki (Osaka), J. Taborda de Messias (Curitiba), K. Tokunaga (Tokyo), B. Uring-Lambert (Strasbourg), N. Usher (Liverpool). 2 Abbreviations used in this statement are explained on p. 174 of this issue.
Mauff/Alper/Dawkins/Doxiadis/Giles/Hauptmann/Rittner/Schneider
Introduction
A common designation for the allotypes of the fourth component (C4) of human complement was proposed in a nomencla ture statement in 1983 [1] based on an ear lier suggestion by Awdeh and Alper [2], Since then the genetic basis of C4 polymor phism has in principle been resolved al though a number of experimental findings remain unexplained. There are two kinds of C4 protein in the plasma of most individu als, C4A and C4B, with different antigenic ity [3] and numerous allotypes for each [1], C4A and C4B are encoded at two separate but closely linked loci [4] within the major histocompatibility complex (MHC) on the short arm of chromosome 6 [5]. Duplica tions and deletions or non-expressions are common for both loci [6-9]. Some individu als carry hybrid C4 genes which are thought to code for unusual, aberrant proteins, some times with reversed antigenicity. Besides progress in the definition of allo typic characteristics of the C4 proteins sub stantial knowledge has come from molecular investigations at the DNA level [see e.g. 10, 11]. Nevertheless, due to a multitude of de tectable variants of the expressed proteins as opposed to limited variability at the DNA level, being more closely related to epitopic differences of C4 [11], the nomenclature of the protein allotypes has continued to be in use over the years. Obviously, for the desig nation of phenotypes, allotypes, alleles, genes, and genotypes the present principle of C4 variant designation will remain to be the basis for communication in the foreseeable future. However, the identification of aber rant C4 proteins, recognition of duplicated C4 genes as part of defined haplotypes with other MHC genes, and an increasing number
of newly reported, in part identical, variants call for amendments in the nomenclature of C4 allotypes. Therefore, at a public nomen clature session of the Vlth Complement Ge netics Workshop and Conference on July 29, 1989, at Mainz, FRG, a revision of the present nomenclature was discussed and adopted. It was based on preworkshop refer ence typing which included the majority of known C4 variants. Detailed accounts of the reference typing will be found in other con tributions to this issue [12-16], To avoid redundancy, the following rules will only repeat the main principles of the 1983 C4 Nomenclature for better under standing and specify new or revised parts; all other unchanged details will be found in the 1983 nomenclature statement [1],
General Rules for C4 Nomenclature and Technology of C4 Typing
In principle, the revised C4 nomenclature follows the terminology of the Guidelines for Human Gene Nomenclature (ISGN, 1987) [17], and also the general rules of the 1983 nomenclature statement. Since the C4B pro tein has been associated with Chido 4 and high hemolytic activity without exception, it is therefore suggested that relative hemolytic activity in an overlay on agarose gel electro phoresis containing sheep red blood cells and C4-deficient guinea pig serum is applied to distinguish between C4A and C4B pro teins [2], C4B showing about a four times higher activity than C4A [18]. Therefore, the basic typing procedure consists of immunofixation with C4-specific antisera after sepa ration of samples in agarose gel electropho resis and simultaneous hemolytic overlay as outlined in detail in table 1 in the 1983 stateDownloaded by: University of Exeter 144.173.6.94 - 5/6/2020 5:42:21 PM
262
C4 Nomenclature Statement (1990)
263
Table 1. Recognized C4 alleles Allele designation
RM
new
previous
mean
A Q0 A8 A7 A6 A 58 A 55 A5 A 45 A4 A3 A2 A 12 A1 A 91
A Q0 A7 A7 A6 A6 A6 A5 A5 A 4/35 A3 A2 A 12/A1 A1 A 91/A90/A1/B31
silent allele 197.2 183.9 138.2 136.0 127.8 119.4 116.9 107.9 98.9 82.6 64.0 54.2 47.8
BQ0
BQ0
B6 B5 B 45 B4 B 35 B3 B 22 B2 B 13 B 12 B 11 B 1 B 92 B 94 B 95 B 96
B 6/B7 B 5/B6 B5 B4 B3 B 29/3/4 B 22 B2 B 12 B 12 B 12/A 95 B 1 B 92 B 94 B 95 B9
silent allele 72.6 67.3 59.2 46.6 43.5 39.2 33.2 29.4 12.9 9.5 7.7
Lysis type
MAB
1
A A A A A A A A A A A A A
n.d. A A A A A A A A A A/B A/B B
B B B B B B B B B B B B B B B B
B/A A/B A A/B B A/B B B A A A B B B n.d. B
range
136.3-139.4
4
127.5-128.0 116.1-121.1
2
1 4
1 1 1 1 59.6-67.8 53.6-54.8 45.3-49.8
6
72.2-73.0 62.5-70.4
2 8
46.3-47.2 42.6-44.3 37.0-42.5
3
3 6
1 2
9
1 27.6-31.2 12.7-13.2 8.7-10.0
2
3 3
1 1
0.0
-20.7 (- 40.7)a n.d. (-92.6)a
n
-19.7-21.1
2
1 1
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RM = Relative migration value (mean value if several samples were tested; 3-10 determinations per indi vidual sample), the range represents the lowest and highest individual allotype value, n = number of tested samples; MAB = monoclonal antibody reactivity anti-C4A/Rgl,2, anti-C4B/Chl [12, 13]. n.d. = not deter mined. Only estimates.
Mauff/Alper/Dawkins/Doxiadis/Giles/Hauptmann/Rittner/Schneider
ment, with the exception that before neur aminidase (NANA’se), treatment with CARB-B [ 19] should be used if the number of expressed alleles and the relative migra tion distance are to be determined. Additional techniques may be necessary or helpful to define or subtype allotypes and haplotypes, such as Western blots with poly clonal or monoclonal antibodies after pro longed agarose-gel electrophoresis, Rodgers/ Chido typing, C4 a- and ß-chain typing [9, 20-22], or DNA TaqI RFLP for the recogni tion of gene size and duplications [6].
Designation of C4 Allotypes
The allotypes are to be designated ac cording to their relative electrophoretic mo bility by increasing arabic numbers from
cathode to anode for both proteins (fig. 1,2, table 1). Allotypes with migration positions cathodal of A 1 or B 1 should be preceded by the number 9 (e.g. A 91, A 92, etc., B 91, B 92, etc.). The two most common allotypes in all ethnic groups A 3 and B 1, as well as the majority of the less common variants will retain their single-digit numeric desig nation [1], with the exception of C4B 5, B 6, and B 7. A redesignation of this region was found to be necessary since the most com mon variant ‘B6’, almost invariably associ ated with C4A 4, HLA-B55 and DR 1 or W6(14), has repeatedly been published as C4B 5. Thus, to avoid any further misun derstanding, the variant designated B 5 in the 1983 nomenclature will now become B 45, B 6 will be B 5, and B 7 will be B 6. The deleted or nonexpressed C4A will be C4A Q0, the deleted or nonexpressed C4B will be C4B Q0. Downloaded by: University of Exeter 144.173.6.94 - 5/6/2020 5:42:21 PM
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C4 Nomenclature Statement (1990)
265
Fig. 1. C4 allotypes. Prolonged immunofixation agarose-gel electrophoresis (CARB-B and NANA’se treatment) of standard and rare allotypes submitted to the Vlth Complement Genetics Workshop Reference Typing Phenotype a
b
c
1: 3: 5: 7: 9: 11: 13:
WOO RIT MAU DUT MAU ALP ALP
14: 16: 18: 20: 22: 24:
RIT TOK ALP DOX MAU TAB
25: DUT 27: TOK
001/1
AB 006/5 002/2
015/1 004 007/2 HDW CWT2 007/1 001/1
006/4 001/7 002/2
CWT6
C4A
B
Phenotype
8 6
1 1 1
2: 4: 6: 8: 10: 12:
CUC TAB ALP MAU MAU ALP
15: 17: 19: 21: 23:
RIT DOX DAW MAU HAU
55, 2 45, 12 3 12, 91 91
(92)
1 Q0 Q0
4, 3, 2 4 91 3 4 3, 2
6 45, 2 35 35, 22, 1 5, 3, 13, 1
45,12 4
92 2, 1,96
C4A 001/1 001/2
001
037/1 006/5 006
KHH
7, 3 58, 3 5,3 4 55, 2 3, 1
B
1 3, 1
1 2
1 1 5 35, 22, 1 3, 1
002/2
4, 3, 2 3 3 4 3, 2
001/1
6, 3
1, 94
001/1
Rob 037/1
2 12, 1
11, 1
26: PAR
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★: A 91 B 35 in sample 18 could not electrophoretically be separated but were distinguished by lysis and segregation analysis. The classified allotypes are in bold face (standard separation conditions). For explanation of abbreviations see page 174 of this issue.
Mauff/Alper/Dawkins/Doxiadis/Giles/Hauptmann/Rittner/Schneider
266
--
1 90
-
180
-
170
-
160
-
1 50
-
140
-
130
-
1 20
-
110100 90 80 70 60 50 40
C4B
C4A
RM 200
8
6 58555 45'' 4
A 3
-
2
-
6 5 45-
12-
-
4
91-
3-35J 22-
-
30
-
20
-
10
-
2--- ■
B 1
13-, 12-iï=
o -10
-20 -30
-
94-,
-40 -50 -60
92-,
-
95
-70 -80
-
-90
-
-100
96
-
—
Fig. 2. Relative migration (RM) distances of C4 allotypes seen during the VIth Complement Genetics Workshop reference typing (C4A 3, B 1 with minor band positions).
they should receive a one-digit designation if they migrate outside the range of the known variants, two or three digits if they migrate between known allotypes. Allotype classifications may be estimated according to the Downloaded by: University of Exeter 144.173.6.94 - 5/6/2020 5:42:21 PM
As before, allotypes with intermediate migration between already described aliotypes will receive a two (or three) digit designation. Newly discovered allotypes should not interfere with already described variants;
C4 Nomenclature Statement (1990)
C4 Reference Variants
No provision for reference laboratories has been made for the future. But a small number of standard reference variants will be available on request for a limited period of time with the senior author. They are exclusively intended for the classification of internal laboratory standards.
Amendments
Any modifications of the present C4 no menclature which might become necessary subsequently should be discussed in public at international meetings (such as the Inter national Complement or Complement Ge netics Workshops) after prior invitation or announcement in the program.
Acknowledgements The authors gratefully acknowledge the editorial and technical assistance of Mrs. Martina Brenden, Margot Braun-Stilwell, Hilde Hasbach, Dr. Georg Plum, Gudrun Riethmeister, Beate Stradmann-Bellinghausen, and the untiring secretarial and photo graphic assistance of Mrs. Renate Thelen and Sigrid Glanschneider.
References 1 Mauff G, Alper CA, Awdeh Z, Batchelor JR, Ber trams J, et al: Statement on the nomenclature of human C4 allotypes. Immunobiology 1983; 164: 184-191. 2 Awdeh ZL, Alper CA: Inherited structural poly morphism of the fourth component of human complement. Proc Natl Acad Sci 1980;77:35763580. 3 Tilley CA, Romans DG, Crookston MC: Localiza Downloaded by: University of Exeter 144.173.6.94 - 5/6/2020 5:42:21 PM
major or minor bands after NANA’se treat ment. A more reliable classification will re sult from the combination of the measure ments of relative migration distances after CARB-B treatment, using the distance be tween C4B 1 and C4A 3 as 100 (see table 1) and electrophoretic side by side comparison; details of the method have been described in Mauff et al. [23]. Of the variants described in the 1983 no menclature statement, examples of C4A 51, 14, 13, 11, C4B 51, 31, 29, 21, 91, 93 were not submitted or seen during this workshop. C4B 31 is now considered to be identical with C4A 91, and B 29 with B 3. Others may have received new designations or positions such as C4A 51, A 91, B 51 or B 4, or possi bly belong to heterogeneous groups not yet recognized, such as A 14, A 13, 11 or B 21. Their classification will have to wait for fu ture reference typings. Duplicated loci, if written as haplotype or genotype, are expressed by repetition of the locus symbol: e.g. C4A*3 A*2; they will be placed in parentheses if written as phenotype or allotypes without repetition of the locus symbol: e.g. C4A (3, 2). Aberrant allotypes or products of hybrid genes, if discovered by additional methods, e.g. Rodgers and Chido typing, Western blot with monoclonal antibodies, C4-chain typ ing, gene size or RFLPs will not receive a special nomenclature. But if subtypes are described they should be designated and ex plained by a special suffix to the main allo type. For RFLPs a special nomenclature was not thought to be necessary for the time being. Designations should include the probe, restriction enzymes and the size of the detectable fragments and possibly their relative intensities, but otherwise follow the general rules of the ISGN [17).
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11
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tion of Chido and Rodgers determinants to the C4d fragment of human C4. Nature 1978;276: 713. O’Neill GJ, Yang SY, Dupont B: Two HLA-linked loci controlling the fourth component of human complement. Proc Natl Acad Sci 1978;75:5165— 5169. Rittner C, Hauptmann G, Grosse-Wilde H, Grosshans E, Tongio MM, Mayer S: Linkage be tween HLA (major histocompatibility complex) and genes controlling the synthesis of the fourth component of complement; in Kissmeyer-Nielsen F (ed): Histocompatibility Testing 1975. Copen hagen, Munksgard, 1975, p 945. Schneider PM, Carroll MC, Alper CA, Rittner C, Whitehead S, et al: Polymorphism of the human complement C4 and 21 steroid hydroxylase genes. J Clin Invest 1967;78:650-657. Giles CM, Uring-Lambert B, Boksch W, Braun M, Goetz J, et al: The study of a French family with two duplicated C4A haplotypes. Hum Genet 1987;77:359-365. Yu CY, Campbell RD, Porter RR: A structural model for the location of the Rodgers and the chido antigenic determinants and their correla tion with the human complement component C4A/C4B isotypes. Immunogenetics 1988;27: 399-405. Steuer M, Mauff G, Adam C, Baur MP, Bender K, Goetz J, et al: An estimate on the frequency of duplicated haplotypes and silent alleles of human C4 protein polymorphism. I. Investigation in healthy Caucasoid families. Tissue Antigens 1989; 33:501-510. Carroll MC, Campbell RD, Bentley DR, Porter RR: A molecular map of human major histocom patibility complex class III region linking comple ment genes C4, C2 and factor B. Nature 1984;307: 237-241. Yu CY, Belt KT, Giles C, Campbell RC, Porter R: Structural basis of the polymorphism of human complement components C4A and C4B: Gene size, reactivity antigenicity. EMBO J 1986;5: 2873-2881. Doxiadis G, Grosse-Wilde H: C4 allotyping by prolonged gel electrophoresis and immunoblotting using monoclonal and polyclonal antibodies. Complement Inflamm 1990;7:269-276. Giles CM: C4: Rodgers and Chido typing. Com plement Inflamm 1990;7:213-217.
14 Mauff G, Brenden M, Braun-Stilwell M, Doxiadis G, Giles C, Hauptmann G, Rittner C, Schneider PM, Stradmann-Bellinghausen B, Uring-Lambert B: C4 reference typing report. Complement In flamm 1990;7:193-212. 15 Rittner C, Stradmann-Bellinghausen B: C4 alphachain reference typing report. Complement Inflamm 1990;7:225-229. 16 Schneider PM: C4 DNA RFLP reference typing report. Complement Inflamm 1990;7:218-224. 17 Shows TB, McAlpine PJ, Boucheix C, Collins FS, Coneally PM, Frezal J, et al: Guidelines for hu man gene nomenclature. An international system for human gene nomenclature (ISGN, 1987). Cytogenet Cell Genet 1987;46:11-28. 18 Isenman DE, Young JR: The molecular basis for the difference in immune hemolysis activity of the Chido and Rodgers isotypes of human comple ment component C4. J Immunol 1984; 132:3019— 3027. 19 Sim E, Cross SJ: Phenotyping of human comple ment C4, a class-III HLA antigen. Biochem J 1986;239:763-767. 20 Doxiadis G, Doxiadis I, Frenz G, Vogeler U, Grosse-Wilde H: Relevance of complotyping and subtyping of MHC class I gene products in haplo type definition for allogeneic bone marrow trans plantation. Bone Marrow Transplant 1989;4:17— 22.
21 Giles CM: Partial inhibition of anti-Rg and antiCh reagents. II. Demonstration of separable anti bodies for different determinants. Vox Sang 1985; 48:167-173. 22 Roos MH, Mollenhauer E, Demant P, Rittner C: A molecular basis for the two locus models of human complement component C4. Nature 1982; 298:856. 23 Mauff G, Brenden M, Braun-Stilwell M: Relative electrophoretic migration distances for the classi fication of C4 allotypes. Complement Inflamm 1990;7:277-281.
Gottfried Mauff Institut für Medizinische Mikrobiologie und Hygiene Universität zu Köln Goldenfelsstrasse 21 D-W-5000 Köln 41 (FRG)
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