Sequence heterogeneity of murine complementary DNA clones related to the C4 and C4-Slp isoforms of the fourth complement component

Tosi, Mario; Lévi‐Strauss, Matthieu; Duponchel, Christiane; Meo, Tommaso

doi:10.1098/rstb.1984.0099

Cited by 23 publications

(10 citation statements)

References 19 publications

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“…In fact, the limited sequence comparison (nucleotide residues 460-816 of Fig. 1) that we have made between pMC4/7 (C4) and pMC4/201 (C4-Slp) revealed only five nucleotide replacements, of which three are silent (17). Since selective arguments can be rejected because of the extensive divergence between mouse and human C4 in this region, this observation supports the notion that the mouse C4 duplication is indeed a recent event.…”

Section: Methodssupporting

confidence: 66%

“…1). Interestingly, in the same region of C4-a4 we have also found an exceptional cluster of nucleotide substitutions (unpublished data) between the sequence of pMC4/7 and that of pMC4/201, a different murine C4 cDNA clone previously related to the C4-Slp isotype (17). Noticeably, the amino acid sequences deduced from the murine cDNA clones fail to reveal any correspondence between the human and murine isotypes.…”

Section: Methodsmentioning

confidence: 87%

“…Four sites where amino acid replacements distinguish the human C4A and C4B protein (15) are marked: * and o indicate identity of the murine residues with C4A and C4B, respectively. (17). We have determined the nucleotide sequences of cosmid 10.8 around this HindIII site, and we found exonic sequences corresponding to those of cDNA clone pMC4/7 (data not shown).…”

Section: Methodsmentioning

confidence: 99%

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Multiple duplications of complement C4 gene correlate with H-2-controlled testosterone-independent expression of its sex-limited isoform, C4-Slp.

Lévi‐Strauss

Tosi

Steinmetz

et al. 1985

Proc. Natl. Acad. Sci. U.S.A.

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Mouse liver cDNA clones related to the C4 and C4-Slp isoforms of the fourth component of complement differ by few nucleotide changes within a region of substantial divergence from human C4. It is suggested that the mouse C4 gene duplication is an evolutionarily recent event with respect to the time of mammalian radiation. This conclusion is reinforced by the presence of a single C4 gene in the Syrian hamster. Most H-2 haplotypes, including those characterized by an undetectable C4-Slp protein, possess two C4 gene copies which, in contrast to the neighboring factor B, show a marked restriction site polymorphism. The genetic variation of this region is emphasized by the presence in the mouse of a rare "polymorphism" for C4 gene number. Multiple C4-related gene copies characterize those exceptional wild-derived H-2 haplotypes, H-2w7, H-2wl6, and H-2wl9, that determine the expression of the C4-Slp protein in female animals.The S region of the mouse H-2 gene complex was defined by two nonallelic genes encoding the isotypic forms C4 and C4-Slp (sex-limited protein) § of the fourth component of the complement system (1-3).Prior to the discovery of their biological function and the recognition that discrete genes encode the C4 and C4-Slp forms, the two proteins were collectively referred to as the mouse Ss (serum substance) protein, detected by means of xenoantisera (see ref. 4 for a recent review). C4-Slp, the form that is identified by an allotypic antiserum, does not possess all the functional properties of C4 and in fact is hemolytically inactive. The two proteins display considerable structural and quantitative genetic variation (5) and are also distinguished by the regulation and site of expression of their genes. C4-Slp is normally produced only in hepatocytes and its expression is controlled by at least two genetically dissectable mechanisms. By definition, this protein displays sexual dimorphism in that it is androgen controlled and normally undetectable (although testosterone inducible) in females of the standard C4-Slp-positive strains. However, some exceptional wild-derived H-2 haplotypes-namely, H_2w7, H-2w16, and H-2w`9-determine the expression of C4-Slp in both male and female mice (6, 7). In these mice, the protein is produced not only by the liver but also by resident peritoneal macrophages, a cell type that in all strains is the second normal site of C4 expression. This allelism for an androgen-controlled or androgen-independent C4-Slp expression was shown by segregation analysis to involve cis-acting regulatory elements genetically inseparable from the structural gene (8). Alternatively, C4-Slp can be expressed in female mice of certain strains by a second type of androgen-and androgen receptor-independent mechanism, which, however, is not linked to the H-2 complex. This latter phenotype appears to be controlled by transacting and as yet unmapped regulator genes (rsl) able to mimic testosterone action (9). On the other hand, the genetic bases of the lack of C4-Slp even in the males of strains...

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Section: Methodssupporting

confidence: 66%

Section: Methodsmentioning

confidence: 87%

Section: Methodsmentioning

confidence: 99%

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Multiple duplications of complement C4 gene correlate with H-2-controlled testosterone-independent expression of its sex-limited isoform, C4-Slp.

Lévi‐Strauss

Tosi

Steinmetz

et al. 1985

Proc. Natl. Acad. Sci. U.S.A.

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“…DNA from cosmid clones was 32P-labeled by nick-translation (7) and hybridized to blots of electrophoretically separated liver poly(A)+ RNA. To localize the hybridizing sequences, restriction endonuclease fragments of cosmid clones were separated on agarose gels, blotted as described above, and hybridized with oligo(dT)-primed, 32P-labeled cDNA (2) (6,9,10). Starting from these clones, we isolated and linked by overlap-hybridization methods a number of S-region cosmids.…”

mentioning

confidence: 99%

Liver mRNA probes disclose two cytochrome P-450 genes duplicated in tandem with the complement C4 loci of the mouse H-2S region.

Amor¹,

Tosi²,

Duponchel³

et al. 1985

Proc. Natl. Acad. Sci. U.S.A.

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A search for uncharacterized genes of the S region of the murineH-2 major histocompatibility complex was undertaken; a series of cosmid clones previously aligned by overlap hybridizations were used as radiolabeled probes. Sequences hybridizing with liver poly(A)+ RNA were found within a cosmid covering a region 3' to the C4-Slp gene (the gene encoding the hemolytically inactive isoform of the fourth component of serum complement). Radiolabeled, short cDNA complementary to liver poly(A)+ RNA was used to establish the transcriptional polarity of the newly detected gene and to define fragments containing its 3' end. DNA sequence analyses and comparisons with porcine peptides established that the gene encodes the enzyme steroid 21-hydroxylase (EC 1.14.99.10), a cytochrome P-450 often referred to as P-450(C21), whose major site of expression is the adrenal gland. Two copies of the P-450(C21) gene, very similar yet distinguishable by restriction endonuclease analysis, were found individually associated with C4 and C4-Slp, genes that encode isoforms of mouse fourth component of complement. One of the P450(C21) genes is coamplified with C4-Slp in H-2 7, a haplotype carrying a rare elongation of the S region. Comparisons with other members of the P450 gene family show that the P-450(C21) genes encode peptides of extraordinary evolutionary conservation. The detection of a liver transcript of P-450(C21) raises the issue of the specific metabolic role of this enzyme in this organ and may have implications for the interpretation of human congenital adrenal hyperplasia.The S region of the murine H-2 major histocompatibility complex is commonly viewed as composed of loci related to the complement system, or class III genes. Although it represents the largest genetic segment (0.4 centimorgan) of the H-2 complex, surprisingly few marker genes have been definitely mapped in this region.To extend the functional characterization of this H-2 segment, we sought coding DNA sequences in cloned genomic fragments by using mRNA hybridization and overlapping cosmid clones. We now report the finding of DNA sequences that hybridize with a 20S liver mRNA and that lie immediately adjacent to the gene (C4) encoding the fourth component of complement (C4) and to the gene (C4-Slp) encoding the nonhemolytic isoform of C4, sex-limited protein (Slp). By sequencing genomic DNA, we have established that these sequences correspond to two copies of the gene encoding steroid 21-hydroxylase [21-OHase; steroid 21-monooxygenase; steroid,hydrogen-donor:oxygen oxidoreductase (21-hydroxylating), EC 1.14.99.10], a member of the cytochrome P-450 isozyme superfamily (1). MATERIALS AND METHODSPreparation of Liver DNA and RNA. Genomic DNA was isolated (2) from liver nuclei prepared by the citric acid method (3). RNA was isolated essentially as described (4) and precipitated in 2.7 M lithium chloride. The polyadenylylated RNA fraction was bound to poly(U)-Sephadex (Bethesda Research Laboratories) according to the instructions of the manufacturer but was elut...

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“…Seven probes were used in this study: 1) pAG64c, a general mouse class I clone of 0.52 kb from the 3' end of an H-2 d cDNA clone (Brickell et al 1983(Brickell et al , 1985; 2) pGRC1.7, a 1.7 kb rat clone from a DNA cosmid clone that maps to the grc + region and is deleted in the grc-strains (Cortese ; 3) A~, a 6 kb Eco RI fragment from the 41.1 kb BALB/c genomic cosmid clone that contains the entire A~ gene from the first exon to an Eco RI site in the 3" untranslated region (UT; Steinmetz et al 1982;Malissen et al 1983); 4) C4, a 1.9 kb Bam HI fragment of the mouse pMC4/7 cDNA clone subcloned into pUC8 that encodes the carboxy terminal half of the ecchain and almost the entire length of the -y-chain and that detects only C4-related genes (Tosi et al 1984;Levi-Strauss et al 1985); 5) pTL3', a 641 base pair (bp) Pst I/Hind III fragment from the T13 c gene of the BALB/c mouse that is a thymus leukemia (TL)-specific probe (Pontarotti et al 1986); 6) Hsp 70, a 2.3 kb Barn HI~Hind III genomic DNA fragment that encodes the entire human heat shock protein (Hunt and Morimoto 1985; ATCC, Rockville, MD); and a mouse TNF a probe (unpublished). The probes were labeled by random priming Vogelstein 1983, 1984) using commercial kits to an activity of 7.5 x 105 cpm/ml of the hybridization solution, and 0.15 ml of the solution was used per 1 cm 2 of membrane.…”

Section: Methodsmentioning

confidence: 99%

Physical mapping of the MHC and grc by the pulse field electrophoresis

et al. 1992

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The development of the physical map of the major histocompatibility complex of the rat was undertaken using pulse field gel electrophoresis of fragments of genomic DNA from the BIL/2 (grc+) and BIL/1 (grc-) strains obtained primarily from single and double digests with the enzymes Mlu I, Not I, and Sfi I and hybridized with a variety of mouse, rat, and human probes. Both strains are maintained by inbreeding the BIL heterozygote (forced heterozygosity; F31); hence, their differences lie almost entirely in the MHC-grc regions. The MHC-grc region was contained in five fragments of DNA comprising 3000-3200 kilobases (kb); thus, its size appears to be closer to that of the human MHC than to that of the mouse MHC. This distance may be an underestimate of the size of the entire region, however, because the cluster of class I loci in the RT1. A region could not be defined in detail in this study. The most striking difference between the BIL/2 strain, which has normal growth and reproductive characteristics, and the BIL/1 strain, which has growth and reproductive defects and an enhanced susceptibility to chemical carcinogens, is a deletion of approximately 70 kb in the latter strain. The studies on grc+ and grc- strains suggest that the phenotypic defects of the grc- strains may be due to the loss of genes that are normally present in this deleted region.

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Sequence heterogeneity of murine complementary DNA clones related to the C4 and C4-Slp isoforms of the fourth complement component

Cited by 23 publications

References 19 publications

Multiple duplications of complement C4 gene correlate with H-2-controlled testosterone-independent expression of its sex-limited isoform, C4-Slp.

Multiple duplications of complement C4 gene correlate with H-2-controlled testosterone-independent expression of its sex-limited isoform, C4-Slp.

Liver mRNA probes disclose two cytochrome P-450 genes duplicated in tandem with the complement C4 loci of the mouse H-2S region.

Physical mapping of the MHC and grc by the pulse field electrophoresis

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