DNA polymorphism of the human complement component C7 gene in familial deficiencies

Summary A C7 cDNA probe detecting a TaqI restriction fragment length polymorphism has been used to examine the segregation of the silent allele (C7^*Q0) in two familial deficiencies. Carrier diagnosis in healthy children is possible when both parents are heterozygotes. Only one of these two families was informative. The silent allele is linked to different TaqI alleles in both families. This suggests that at least two different C7^*Q0 alleles are present in our population. This paper gives a protocol for genetic studies of hereditary traits in which the C7 gene and other genes tightly linked to it are involved.     

Relationships between the gene and protein structure in human complement component C9

Human complement component C9 is a multidomain protein for which a large number of surface topographical features have been determined. We have analyzed the exon-intron boundaries of the human C9 gene and find a good correlation between splice sites and surface features of the protein but little correlation with the putative protein domain structure, even in the cysteine-rich sequence homology with the low-density lipoprotein (LDL) receptor which is likely to be an independently folded structural motif. This is surprising because in the LDL receptor the same sequence is precisely bounded by introns, and it has been assumed that this sequence is present in both proteins as a result of exon shuffling. We deduce that substantial rearrangement of the exon-intron structure of the C9 gene must have occurred before the exchange of cysteine-rich domains, possibly linked to the process of exon duplication which was required to generate the repeats in the LDL receptor.     

Genetic polymorphism of human complement component C81 in the Japanese population

Summary Genetic polymorphism of human C81 has been investigated using polyacrylamide gel isoelectric focusing (PAGIEF) in the presence of 3.1 M urea followed by electroblotting with enzyme immunoassay. In 448 individuals phenotypes of C81 were classified into three common and four rare patterns, and these were considered to be controlled by two common alleles, C81^*A and C81^* B, and three rare alleles which were tentatively designated C81^*A1J and C81^*A2J for acidic variants and C81^*B1J for the basic variant. The alleles of C81^*A2J and C81^*B1J are new rare alleles, but C81^*A1J might correspond to C81^*A1 in the former studies. Family data were in accordance with the hereditary rules. The gene frequencies were estimated as C81^*A is 0.6228, C81^*B is 0.3672, C81^*A1J is 0.0078, C81^*A2J is 0.0011, and C81^*B1J is 0.0011, respectively. The gene frequencies of the two common alleles agreed approximately with other ethnic groups. PAGIEF of neuraminidase-treated plasma samples followed by electroblotting with enzyme immunoassay is applicable to the study of heterogeneity of C81.     

Genetic polymorphism of complement component C8

Summary Extensive genetic polymorphism of complement component C8 was demonstrated by isoelectric focusing of serum or plasma samples followed by immunoblotting procedures. Using these methods, we could detect both - (C81) and (C82) chain polymorphisms in the same gel. Two-dimensional (2D) electrophoresis of C8 immunoprecipitates was used to obtain further information of the C8 patterns. Evidence was obtained that the C81 polymorphism resides in the structural gene of the C8 chain. Both C8 systems show autosomal, chiefly codominant inheritance, and the distribution of phenotypes agrees with the Hardy-Weinberg equilibrium. Our findings suggest at least five different alleles in the C81 system; the gene frequencies of the two most common ones, C81 ^*A and C81 ^*B being 0.59 and 0.39, respectively. In C82 we found evidence for at least three codominant alleles, the gene frequencies for the two most common ones, C82 ^*B and C82 ^*A being 0.94 and 0.05, respectively. In addition, family studies disclosed the existence of a null allele, C82 ^* Q0.     

Genetic polymorphism of the second component of human complement (C2)

Summary Proteins were separated by prolonged isoelectric focusing in polyacrylamide gels, whereupon C2 bands were detected by a specific hemolytic assay. This was performed by treating the gel with iodine to increase C2 activity, and then developing C2 bands with an agarose gel overlay containing sensitized sheep cells and diluted human serum as a complement source deficient in functional C2. The gene frequencies observed in a material of 122 unrelated adults were: C2¹:0.97 and C2²:0.03.C2 linkage relations and C2 haplotype associations have been examined a family material. It is concluded that C2 is very closely linked to HLA loci.     

The sequence and topology of human complement component C9.

A partial nucleotide sequence of human complement component C9 cDNA representing 94% of the coding region of the mature protein is presented. The amino acid sequence predicted from the open reading frame of this cDNA concurs with the amino acid sequence at the amino-terminal end of three proteolytic fragments of purified C9 protein. No long stretches of hydrophobic residues are present, even in the carboxy-terminal half of the molecule which reacts with lipid-soluble photoaffinity probes. Monoclonal antibody epitopes have been mapped by comparing overlapping fragments of C9 molecule to which the antibodies bind on Western blots. Several of these epitopes map to small regions containing other surface features (e.g., proteolytic cleavage sites and N-linked oligosaccharide). The amino-terminal half of C9 is rich in cysteine residues and contains a region with a high level of homology to the LDL receptor cysteine-rich domains. A model for C9 topology based on these findings is proposed.     

Molecular analysis of human complement component C5: localization of the structural gene to chromosome 9

A human C5 clone (pC5HG2) was isolated from a cDNA library constructed from Hep G2 mRNA. The DNA sequence showed that the pC5HG2 insert was comprised of 3309 base pairs of pro-C5 coding sequence and 404 base pairs of 3'-untranslated sequence. The derived amino acid sequence contained the entire coding sequence of the C5 alpha-chain, the beta-alpha-chain junction region, and 100 amino acids (approximately 50%) of the beta-chain. Protein sequences of four C5 tryptic peptides were aligned exactly to this sequence and demonstrated that C5 synthesized and secreted by Hep G2 cells is probably identical with plasma-derived C5. Coding sequence alignment of the human C5 sequences with those of murine C5 indicated that 80% of the nucleotides and 79% of the amino acids were placed identically in the two species. Amino acid sequence alignment of the homologous family members C3, C4, and alpha 2-macroglobulin with that of C5 demonstrated 27%, 25%, and 19% identity, respectively. As was found in murine C5, the corresponding thiol ester region of human C5 contained several conserved amino acids, but the critical cysteine and glutamine residues which give rise to the intramolecular thiol ester bond in C3, C4, and alpha 2-macroglobulin were absent in C5, having been replaced by serine and alanine, respectively. With the use of a panel of hamster-human somatic cell hybrids, the C5 gene was mapped to human chromosome 9. In situ chromosomal hybridization studies employing metaphase cells further localized the gene to bands 9q32-34, with the largest cluster of grains at 9q34.1.