The Nomenclature of Properdin Factor B Allotypes

Z. Immun.-Forsch. vol. 154, pp. 115-120 (1978) Hygiene.Institut, Universitat Kaln, Federal Republic of Germany Centre de Transfusion Sanguine, Strasbourg, France Africa Institute of South Africa, Pretoria, South Africa Zentrallaboratorium des Blutspendedienstes SRK, Bern, Switzerland

The Nomenclature of Properdin Factor B Allotypes 1) G. MAUFF2 ), G. HAUPTMANN, H.

and R.

W.

HITZEROTH 2 ),

F.

GAUCHEL,

SCHERZ

With 3 Figures R,eceived May 26, 1977 . Accepted in Revised Form December 17, 1977

Abstract In a comparative study the presently known eleven allotypes of properdin factor B (Bf) were examined. Bf polymorphism consists of the two common alleles F and S, the two less common alleles F 1 and S 1 and seven further rare alleles. A variant designation has been proposed according to their relative electrophoretic mobility in comparison to the migration difference between the Sand F 1 band. Three rare variant alleles were redesignated: F 1.55,80.45 and 80.7, which previously had been described as F 1.6, S 0.8 and S 1, respectively. Conversion studies did neither reveal variant mobility in the Bb nor in the Ba fragment of factor B in three of the rare alleles. This finding confirms t,he earlier report on one of the variants, possibly suggesting the existence of a so far unknown third clearing fragment.

Of recently discovered blood protein polymorphisms the study of properdin factor B, first described by ALPER et al. (2), has been one of the most rewarding. Varied distribution of allotypes in different populations and close immunogenetic linkage to the major histocompatibility region in man on chromosome 6 has led many workers to start studies in the field. It is the purpose of this report to present data on the rare variants of human factor B (Bf) and to introduce a designation for rare Bf phenotypes previously proposed (5). 2 common allotypes, F and S, and 2 less common allotypes, F 1 and S 1, were described by ALPER et al. (2). They show an equidistant 5-banded pattern on examination with barbital buffer in immuno-fixation agarose-gel electrophoresis (IAGE) with 1 major band, 3 anodal and 1 cathodal minor band. F and S differ from one another ') Presented in part at the EMBO Workshop on the genetics of the complement system, Cambridge, England, 1977. 2) Supported through a grant to G. M. by the Georg and Agnes BlumenthalStiftung, Berlin, and a research fellowship (1975/76) awarded to H. ·W. H. by the Alexander von Humboldt-Stiftung, Bonn-Bad Godesberg.

116 . MAUFF, HAUPTMANN, HITZEROTH, GAUCHEL, and SCHERZ

by the difference of 1 minor band position whereas F 1 seemed at first to differ by the distance of 2 minor bands anodal from 8, and 8 I by 1 minor band cathodal from 8. Further studies revealed that F 1 or 81 actually migrate at the distance of 212 respectively 112 minor bands from 8 (5). Besides these allotypes a rare allotype faster than F 1 was reported in the serum of a German individual (5) and allotypes migrating between F and F 1 in a French person (3), in South African Bantus and a Colored person (6). Furthermore, rare 8 allotypes could be observed in a 80uth African Colored person (6), in a French man as well as in Shouth American Karaja-Bananal-Goias Indians from Brazil (4) and recently in a Swiss woman. A rare phenotype not included in tIns study was also seen by HOBART and LACHMANN (personal communication) using iso-electric focusing in polyacrylamide gel.

Materials and Methods With the exception of the latter phenotype we have compared all so far reported rare Bf.phenotypes in a combined study, including the Swiss variant which is reported here for the fu'st time. The method of ALPER and co-workers (2) or its modification (6, 3) was applied. By measuring relative migration on charts of densitometric evaluations of the stained electrophoretic plates and using the distance S -+ F 1 as a reference, we applied the designation already introduced by ALPER et al. (1) for rare C3-variants and accepted at the Bonnworkshop (7). The distance of S -+ F and the distance between major and minor bands was found to amount to 40% of S ->- F 1 and was also used as a reference. Mean values of at least 10 determinations 'w ere taken before attributing a final designation. As in C3 polymorphism for the reason of minimal discrimination no designation smaller than 0.05 was chosen. With this technique all rare allotypes were classified (Figure 1).

Results The first rare phenotype to be discovered besides F 1 and 8 1 was found to possess approximately 1.57 (ill = 1.568 ± 0.074,1 a) times the distance of 8 -+- F 1 between its 2 bands. The distance 8 -+- F 1 was determined to be significantly shorter than the distance F -+-S 1 by comparison of absolute differences in the t-test, S 1 being 1.1 times as far apart from F than F 1 from S (5). Consequently, the distance 8-+-81 amounted to 1.1-0.4 = 0.7. The 80uth African allotypes, previously named Fnl , Fn2 and 8n were determined as F 0.65 (ill = 0.66 ± 0.05, 1 a), F 0.85 (ill = 0.83 ± 0.07, 1 a) and 80.3. Less than 10 determinations were done "with the latter type because ofinsufficient amount of serum and relative migration was estimated from the minor band position.

The Nomenclature of Properdin Factor B Allotypes . 117 Rv • F1S0.7'

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Fig. 1. Determination of rare Bf allotypes, example of diagram from one electrQphoretic plate. Distance in mm from chart of densiometric evaluation, Rv = relative variant designation. Reference values: F1S0 .7 = 1.7; F1.55S = 1.55; FSO.7 = 1.1; F1S = 1.0; FIF = 0.6; FS and first anodal resp. cathodal minor bands = 0.4. Determined variants: FnlSO.7; Fn.S; FnlS; Fn.F. Mean values from 10 such det erminations were taken, therefore values from this'chart do not correspond ei'actly to final designation (see also fig. 2).

F 0.85 and S 0.3 nearly coincided with the most pronounced anodal minor band of F or S 1, respectively. Therefore, distinction from normal homozygous F or rare S 1 was found to be difficult and confirmation of these variant phenotypes must await family studies, although the existence of the F 0.85 allele was proven by its combination with S in 2 Negroid individuals. The variants of the 2 French individuals were designated as F 0.55 (ill = 0.57 ± 0.05, 1 0') and 80.8 in 2 laboratories independently by 2 of the authors (G. H. and G. M.). 80.8 was first measured with the distance S.-?S 1 as reference. Using 8.-?F 1, a relative migration of 80.56 was calculated. In the second laboratory 8 0.45 (ill = 0.46 ± 0.04, 1 0') was determined. The difference might be due to inclusion of 2. serum samples from the Brazil Indians which at first also were considered to be identical with Bf «80.8» but later in a more extensive study were seen to differ slightly. The third rare S allotype recently came to our attention. It was found in a 8wiss woman together with common 8 and in three of

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Fig. 3. lmmunofixation agarose-gel electrophoresis with anti-Bf-serum (antiHuman-C3-Aktivator, Behringwerke, Marburg, Germany; Bb-specific). No.1 Bf S, unconverted, nos. 2 to IS; sera converted with 3.33 mg/ml zymosan (lCN Pharmaceuticals, Cleveland, Ohio, USA) at 37°C for IS hours. No.2 Bf S, no. 3 Bf FS, no. 4 Bf F, no. 5 Bf FlS, no. 6 Bf FSO.7*), no. 7 Bf FIF, no. S Bf SSO.7*), no. 9 Bf F1.55S*), no. 10 Bf FO.55S, no. II Bf SS0.45, no. 12 Bf FO.S5S, no. 13 Bf FO.65S, no. 14 Bf FO.S5F, no. 15 Bf FO.65S0.7*), no. 16 Bf FISO.7*), no. 17 BfSO.7*), no. IS BfFl. *) new nomenclature

The Nomenclature of Properdin Fact,or B Allotypes . 119

her children, who possessed FSvar as well as SSvar. From the determination of the distance F -+Svar a relative migration of S 0.25 (ill = 0.23 ± 0.03, 1 a) was calculated. Discussion Although the mode of inheritance has not been checked in all reported variants it can be assumed from electrophoretic migration that at present at least 11 different allotypes exist for factor B. The discovery of further variants is to be expected in the future. At the EMBO workshop on the genetics of the complement system (CAMBRIDGE, 1977) agreement was reached among investigators in the field to adopt the nomenclature ofrelative electrophoretic mobility as applied to C 3 polymorphism for all present and future Bf phenotypes which are not common F or S, using S -+F I as reference distance. Simultaneously, a reference laboratory for the determination of all rare Bf allotypes was established at the Hygiene-Institut at the University of Cologne. Although the designation according to relative mobility will be unsatisfactory to the molecular biologist, until the molecular basis of variation in factor B genetics becomes known a numerical nomenclature for practical purposes will make results compatible from laboratories all over the world. This ma,kes necessary redesignation of 3 of the known allotypes (Figure 2): F 1.6 S, which incidentally was classified with too high a value in the first report, will become F 1.55 S, S 0.8 will become S 0.45, and S 1 will become S 0.7. Studies of the Ba and Bb fragments of some of the rare allotypes have been reported before (2,5,3). In the Bb-fragment of the molecule polymorphism was also confirmed in this study for most allotypes, except for F 0.55 S, F 0.85 F and F 0.65 S 0.7 (Figure 3). Lack of variant banding in F 0.55 S for the Bb and Ba fragment was previously found by one of the authors (3). The difficulties encountered with F 0.85 underline the necessity for family studies of rare Bf phenotypes. No conversion products could be generated in the F 0.65 S .07 (F 0.65 S 1, old nomenclature) serum due to natural ageing. In the Ba fragment common S-, FS- and S-types in accordance with findings by ALPER et al. (2) were visible as polymorphic bands. Again, the F 0.85 F-type appeared as normal F. Double banding was also seen in the FS 0.7-, F 1.55S-, F 0.85 S-, and F 0.65S-types which all were polymorphic also in the Bb fragment. This could mean moleculare variation in these variants in the Ba as well as in the Bb portion of the native factor B molecule. Studies are under way in different laboratories to solve the questions still unanswered, such as polymorphic nature, antigenicity and electro-

120 .

M..~UFF,

HAUPTMANN, HITZEROTH, GAUCHEL, and SCHERZ

phoretic mobility of a hypothetical third fragment or the appearance of intermediate Ba bands during insufficient conversion. Undoubtedly, besides its immunogenetic significance factor B polymorphism will serve as a valuable marker for anthropological studies. References 1. ALPEiR, C. A., and R. P. PROPP. 1965. Genetic polymorphism of the third component of human complement (C3). J. clin. Invest. 47: 21S1. 2. ALPER, C. A., T. BOENISCH, and L. WATSON. 1972. Genetic polymorphism in human glycine-rich beta-glycoprotein. J. expo Med. 135: 6S. 3. HAUPTMANN, G., M. M. TONGIO, and S. MAYER. 1976. Bfpolymorphism: Study of a new variant (FO.55). Hum. Genet. 33: 275. 4. HAUPTMANN, G., E. WERTHEIMER, M. M. TONGIO, and S. MAYER. 1977. Bf polymorphism: Another variant (SO.S). Hum. Genet; 36: 109. 5. MAUFF, G., K. HUMMEL, and G. PULVERER. 1975. Porperdin factor B (glycinerich beta-glycoprotein or C3 proactivator)-polymorphism: Genetic and biochemical aspects. First application to paternity cases. Z. Immun.-Forsch. 150: 327. 6. MAUFF, G., F. D. GAUCHEL, and H. W. HITZEROTH. 1976. Polymorphism of properdin factor B in South African Negroid, Indian and Colored POpulfLtions. Hum. Genet. 33: 319. 7. RITTNER, CH. (Ed.) 1973. First international symposium and workshop on the polymorphism of the third component of tJ;te human complement system. Vox Sang. (Basel) 25: 9. Priv.-Doz. Dr. med. G. MAUFF, Hygiene-Institut del' Universitat, Goldenfelsstr. 21, D - 5000 Kaln 41, Federal Republic of Germany.