Mcp Gene Research Articles

Protective immunity against infectious spleen and kidney necrosis virus induced by immunization with DNA plasmid containing mcp gene in Chinese perch Siniperca chuatsi

Infectious spleen and kidney necrosis virus (ISKNV) is the causative agent of a disease leading to high mortality and economic losses in Chinese perch, Siniperca chuatsi. There is an urgent need to develop an effective vaccine against this fatal disease. In this study, the mcp gene encoding the major capsid protein, the predominant structural component of the iridovirus particles, was cloned into a eukaryotic expression vector pcDNA3.1+, and the recombinant plasmid, designated as pcMCP, was constructed. Expression of the mcp gene was confirmed in transfected cells and muscle tissues of vaccinated fish by RT-PCR, immunodot blot and western blot. Immune response was induced by intramuscular injection of Chinese perch with pcMCP added QCDC adjuvant. The expression levels of type I IFN system genes including IRF-7, IRAK1, Mx and Viperin were up-regulated at 6 h, and reached a peak at 48 h. In addition, there was a second peak of the expression levels of IRF-7 and Mx gene on the 21st day post-vaccination. Before the 21st day post-vaccination, the levels of IgM did not show a significant difference among all groups, but there was a remarkable increase on the 28th day post-vaccination. The relative percent survival (RPS) of Chinese perch vaccinated with pcMCP added QCDC adjuvant was 80% in a challenge trial on the 28th day post-vaccination. Moreover, real-time PCR demonstrated that the levels of viral load in the dead fish of the vaccinated group were significantly higher than those in mock-vaccinated fish. Together, these results indicate that pcMCP is a potential candidate DNA vaccine against ISKNV disease.

Unveiling of the Diversity of Prasinoviruses (Phycodnaviridae) in Marine Samples by Using High-Throughput Sequencing Analyses of PCR-Amplified DNA Polymerase and Major Capsid Protein Genes

Viruses strongly influence the ecology and evolution of their eukaryotic hosts in the marine environment, but little is known about their diversity and distribution. Prasinoviruses infect an abundant and widespread class of phytoplankton, the Mamiellophyceae, and thereby exert a specific and important role in microbial ecosystems. However, molecular tools to specifically identify this viral genus in environmental samples are still lacking. We developed two primer sets, designed for use with polymerase chain reactions and 454 pyrosequencing technologies, to target two conserved genes, encoding the DNA polymerase (PolB gene) and the major capsid protein (MCP gene). While only one copy of the PolB gene is present in Prasinovirus genomes, there are at least seven paralogs for MCP, the copy we named number 6 being shared with other eukaryotic alga-infecting viruses. Primer sets for PolB and MCP6 were thus designed and tested on 6 samples from the Tara Oceans project. The results suggest that the MCP6 amplicons show greater richness but that PolB gave a wider coverage of Prasinovirus diversity. As a consequence, we recommend use of the PolB primer set, which will certainly reveal exciting new insights about the diversity and distribution of prasinoviruses at the community scale.

Analysis of the genetic diversity of the lymphocystis virus and its evolutionary relationship with its hosts

Lymphocystis disease virus (LCDV) is the causative agent of lymphocystis disease. In this study, the mcp gene of LCDV and the cyt b gene of the host fish were selected as molecular markers, and the phylogenetic relationships between LCDV and its host were analyzed. The 25 LCDV isolates examined in this study were attributed to seven LCDV genotypes: genotype I (LCDV-1), genotype II (LCDV-cn, etc.), genotype III (LCDV-rf), genotype IV (LCDV-rc and LCDV-sb), genotype V (LCDV-cb), genotype VI (LCDV-tl), and genotype VII (LCDV-sa). Genotype VII is a new genotype. LCDV1 was found to have differentiated first, followed by LCDV-rf; then LCDV-tl; LCDV-cb; and then LCDV-sa; and by LCDV-rc and LCDV-sb; and finally by LCDV-cn, LCDV-C, and LCDV-jf. From the host evolutionary perspective, Rachycentron canadum was found to have differentiated first, followed by Trichogaster leeri, Chanda baculis, and Sebastes schlegeli, Lateolabrax sp., Sparus aurata, Platichthys flesus, and Paralichthys olivaceus. Comparison of the phylogenies of the host fish species and LCDVs revealed no significant evidence of cospeciation between LCDVs and their host fish. In-depth studies of the genetic variation in LCDVs can enhance our understanding of the mechanism of LCDV infection, which may provide important insights into the prevention and treatment of lymphocystis disease.

Manifestation of atypical hemolytic uremic syndrome caused by novel mutations in MCP

Atypical hemolytic uremic syndrome (aHUS) is a rare disease characterized by microangiopathic hemolytic anemia, thrombocytopenia, and acute renal failure. Mutations in genes encoding regulators of the alternative complement pathway (CFH, MCP, C3, CFI, CFB, THBD, and CFHR1-5) are connected with this disease. Polymorphisms (SNPs) in these genes might also influence the manifestation of aHUS. We have analyzed the genes of CFH, CFI, MCP, and C3 in a cohort of 10 unrelated Czech patients with clinically diagnosed familial aHUS. Surprisingly, 4 patients had mutations only in MCP, without mutations in any of the other genes that cause aHUS. Mutations, as yet unpublished, were widely distributed over the gene (SCR2 domain, signal peptide, and cytoplasmic region). The phenotype of the patients and their close relatives (14 individuals) was also investigated. Functional examination of MCP was also provided and proved lower expression on granulocytes in all mutations. Severity of disease varied, but onset was never earlier than 5 years of age. Penetrance of disease was 50% among carriers. We found that the severity and recurrence of the disease within families varied and might also be dependent on SNPs. Mutations in the MCP gene seems to be a common etiology of aHUS in Czech patients.

Establishment, characterization of a new cell line from heart of half smooth tongue sole (Cynoglossus semilaevis)

A new cell line was established from the heart of a cultured marine fish, half smooth tongue sole (Cynoglossus semilaevis), designated as CSH (Cynoglossus semilaevis heart cell line). The CSH cells grow over 400 days in minimum essential medium (MEM) supplemented with 10% fetal bovine serum (FBS) and 2 ng/ml basic fibroblast growth factor (bFGF). The suitable temperature for the cell growth was 24-30°C with the optimum growth at 24°C and a reduced growth at 12 and 30°C. FBS and bFGF concentration were the two important components for CSH cells proliferation. Twenty percent FBS in the medium was found to be the optimum concentration and bFGF promoted the growth of CSH cells. The double time of the cells at 24°C was determined to 73.39 h. Chromosome analysis revealed that 44% of the cells maintained a normal diploid chromosome number (2n=42) in the CSH cells at Passage 58. The fluorescent signals were observed in CSH after the cells were transfected with green fluorescent protein (GFP) reporter plasmids. CSH cells showed the cytopathic effect (CPE) after infection with lymphosystis disease virus (LCDV). Moreover, the LCDV particles can be observed in the cytoplasm of virus-infected cells by electron microscopy, and a segment of MCP gene for major capsid protein of LCDV was found by PCR amplification DNA of virus-infected cells.

Distribution and expression in vitro and in vivo of DNA vaccine against lymphocystis disease virus in Japanese flounder (Paralichthys olivaceus)

Lymphocystis disease, caused by the lymphocystis disease virus (LCDV), is a significant worldwide problem in fish industry causing substantial economic losses. In this study, we aimed to develop the DNA vaccine against LCDV, using DNA vaccination technology. We evaluated plasmid pEGFP-N2-LCDV1.3 kb as a DNA vaccine candidate. The plasmid DNA was transiently expressed after liposome transfection into the eukaryotic COS 7 cell line. The distribution and expression of the DNA vaccine (pEGFP-N2-LCDV1.3kb) were also analyzed in tissues of the vaccinated Japanese flounder by PCR, RT-PCR and fluorescent microscopy. Results from PCR analysis indicated that the vaccine-containing plasmids were distributed in injected muscle, the muscle opposite the injection site, the hind intestine, gill, spleen, head, kidney and liver, 6 and 25 days after vaccination. The vaccine plasmids disappeared 100 d post-vaccination. Fluorescent microscopy revealed green fluorescence in the injected muscle, the muscle opposite the injection site, the hind intestine, gill, spleen, head, kidney and liver of fish 48 h post-vaccination, green fluorescence did not appear in the control treated tissue. Green fluorescence became weak at 60 days post-vaccination. RT-PCR analysis indicated that the mcp gene was expressed in all tested tissues of vaccinated fish 6–50 days post-vaccination. These results demonstrate that the antigen encoded by the DNA vaccine is distributed and expressed in all of the tissues analyzed in the vaccinated fish. The antigen would therefore potentially initiate a specific immune response. the plasmid DNA was injected into Japanese flounder (Paralichthys olivaceus) intramuscularly and antibodies against LCDV were evaluated. The results indicate that the plasmid encoded DNA vaccine could induce an immune response to LCDV and would therefore offer immune protection against LCD. Further studies are required for the development and application of this promising DNA vaccine.

Mutations affecting chemotaxis of Agrobacterium vitis strain E26 also alter attachment to grapevine roots and biocontrol of crown gall disease

Crown gall in grapevine, caused by tumorigenic Agrobacterium vitis strains, can cause severe losses in most viticulture regions worldwide. The only effective means of control is through cultivation practices. One non‐tumorigenic A. vitis strain, E26, can prevent crown gall infection when applied to wounds on grapevine prior to or simultaneous with inoculation of tumorigenic strains. ME19, a Tn5 mutant of strain E26, was impaired in terms of its ability to be chemo‐attracted by grapevine root tissue extracts and its attachment to grapevine roots, and had reduced biological control activity; it did not significantly differ from the wild‐type strain of E26 in phenotypes of agrocin production, growth in minimum medium or swarming activity. Complementation of ME19 with the cosmid clone of CP1543 from an E26 DNA library restored the chemotaxis, attachment, and biocontrol phenotypes. A 7·3‐kb KpnI fragment from CP1543 was cloned and sequenced, and sequence analysis revealed that the Tn5 insertion occurred in a region that shares a significant homology with genes coding for methyl‐accepting chemotaxis proteins (MCPs) in many bacteria. Complementation of the mcp gene mutants restored the affected phenotypes to the level of wild‐type E26. An in‐frame deletion mutant of the mcp gene was generated and was determined to have the same phenotypes as the original Tn5 mutant.

Transcriptional analysis of a major capsid protein gene from Spodoptera exigua ascovirus 5a

The major capsid protein (mcp) gene of Spodoptera exigua ascovirus 5a (SeAV-5a) was confirmed by aphidicolin viral DNA replication inhibition analysis to be a late gene. The 5' and 3' ends of mcp gene transcripts have been mapped. Primer extension analyses indicated that transcription of the mcp gene initiates from a cytosine 25 nucleotides (nt) upstream of the translation start codon. Two independent approaches by 3' rapid amplification of cDNA ends (3' RACE) and oligo (dT) cellulose binding assay suggested that SeAV-5a mcp mRNA is polyadenylated. Analyses by 3' RACE also revealed that mcp transcripts terminate at a U, either at 26 or 38 nt downstream of the translation stop codon. The putative 5' transcription control region of the SeAV-5a mcp gene shares similarities with other ascoviruses and Chilo iridescent virus (CIV), containing a conserved TATA-box-like motif (TAATTAAA) and an ATTTGATCTT motif upstream of it. The 3' downstream regions of the mcp gene of all the ascoviruses examined and CIV can form a stem-loop structure, and the ends of the mcp gene transcripts of SeAV-5a are within the predicted stem-loop region. This suggests that the stem-loop structure of the mcp gene might be involved in transcription termination.

Detection of major bacterial and viral pathogens in trash fish used to feed cultured flounder in Korea

The possibility of trash fish food-direct pathogen infection to cultured flounder is a very important concern of fish farmers in Korea. A total of 26 groups of trash fish and shrimp samples were collected from 11 hatcheries on Jeju Island. Four groups and two groups of homogenized frozen trash fish samples were directly isolated and confirmed by PCR assays with S. iniae and S. parauberis, respectively. One iridovirus PCR positive result was obtained from one group of mixed trash fish samples. Phylogenetic analysis based on the MCP gene from one iridovirus strain was genetically closely related to seabass, red sea bream, rock bream, turbot, and Korean flounder iridovirus. Betanodaviruses were not detected from the trash fish samples. Based on these results, trash fish can be one of the sources or carriers of bacterial and viral pathogens for feeding cultured flounder in Korea.

Unusual Clinical Severity of Complement Membrane Cofactor Protein–Associated Hemolytic-Uremic Syndrome and Uniparental Isodisomy

Atypical hemolytic-uremic syndrome (aHUS; OMIM 235400) is genetically and clinically heterogeneous. Mutations in membrane cofactor protein (MCP; CD46), a widely expressed complement regulator, predispose to recurrent forms of the disease. Patients carrying MCP mutations have a favorable clinical outcome in comparison to those with factor H (CFH) or factor I (IF) mutations, which lead in most cases to end-stage renal failure. We identified 1 patient who presented at 1 year of age with a first episode of aHUS requiring dialysis therapy. After 2 recurrences of the disease, the patient developed end-stage renal failure. No mutation in the CFH and IF genes was found. A novel homozygous mutation (IVS10+2 T-->C) in the splice-donor of exon 10 encoding the transmembrane region of the MCP gene was associated with dramatically decreased cell-surface expression of MCP. Because the nucleotide substitution was inherited from the patient's father, but not her mother, a large deletion or uniparental disomy was suspected. Both karyotyping and cytogenetic analysis of chromosome 1q32 were performed, for which MCP maps showed no abnormalities. Subsequent genotype analysis using microsatellite markers spanning chromosome 1 showed that the affected child was homozygous for the entire series of markers tested and that all alleles originated from the father. Complete paternal uniparental isodisomy of chromosome 1 is a novel mechanism resulting in severe deficiency of MCP expression. The outcome of the disease reported here indicates that MCP mutation and complete paternal uniparental disomy of chromosome 1 could have an additive effect in determining the severity of the HUS phenotype.

Study on the distribution and expression of a DNA vaccine against lymphocystis disease virus in Japanese flounder (Paralichthys olivaceus)

The distribution and expression of lymphocystis disease virus (LCDV) vaccine, on the basis of DNA vaccine (pEGFP-N2-LCDV0.6 kb) construction, were analyzed in tissues of the Japanese flounder by PCR, RT-PCR and fluorescent microscopy. Results from PCR studies indicated that the vaccine-containing plasmids were distributed in injected muscle, muscle located opposite the injection site, hind intestine, gill, spleen, head kidney, liver and gonad 7 days after vaccination. However, these vaccine-containing plasmids disappeared by 90 days following vaccination. Fluorescent microscopy observations revealed that green fluorescence appeared in muscle, muscle located at the opposite side of the injection site, hind intestine, gill, spleen, head kidney and liver of fish 36 h after vaccination, and that green fluorescence did not appear in control tissue. The green fluorescence became weaker at 60 days post-vaccination, however, it remained detectable in the spleen 90 days post-vaccination. Results from RT-PCR studies indicated that the Mcp gene is expressed in all tissues of vaccinated fish 7–20 days after vaccination. These results demonstrate that the DNA vaccine is distributed and expressed in different tissues of vaccinated fish, and therefore, may have provided an antigen producing specific immune response.

Molecular characterization of three Rana grylio virus (RGV) isolates and Paralichthys olivaceus lymphocystis disease virus (LCDV-C) in iridoviruses

Three Rana grylio virus (RGV) isolates and lymphocystis disease virus (LCDV-C) were molecularly characterized by antigenicity comparison, Western blot detection of viral polypeptides, restriction fragment length polymorphism analysis of viral genomes, and MCP sequence analysis. Significant antigenicity differences existed among the three RGV isolates and LCDV-C. Western blot detection indicated that the viral polypeptides of three RGV isolates could be recognized by the anti-RGV9807 serum, whereas no bands were observed in the LCDV-C, and significant differences exist among the band patterns of three RGV isolates. Restriction fragment length polymorphism (RFLP) analysis was performed by digesting genomic DNA of the four iridovirus isolates with restriction endonucleases HindIII, KpnI, XbaI and BamHI. On the whole, obvious discrepancies existed between LCDV-C and RGV isolates, and some significant band pattern differences were also revealed between RGV9808 and RGV9506 (or RGV9807) in the profiles of restriction endonucleases XbaI, KpnI and BamHI. PCR amplification and sequence analysis of MCP gene sequence further revealed their phylogenetic relationship among the three RGV isolates, LCDV-C and other iridoviruses. RGV9506, RGV9807 and RGV9808 are clustered together with other ranaviruses, such as FV3, BIV, TFV and ENHV, although the RGV9808 is more close to EHNV than to other ranaviruses. Additionally, LCDV-C is clustered with LCDV-1, the type species of genus Lymphocystisvirus. The current study provides clear evidence that significant genetic difference exists among the three RGV isolates. Therefore, further work on comparative genomic studies will contribute significantly to understanding of their taxonomic position and pathological mechanism.

Breast Cancer Progression and Host Polymorphisms in the Chemokine System: Role of the Macrophage Chemoattractant Protein-1 (MCP-1) −2518 G Allele

Interaction between tumor cells and stroma is essential for tumor growth. Tumor cells stimulate the formation of stromal tissue, which excretes a variety of growth factors, cytokines, and proteases. Tumor-associated macrophages (TAMs) are one of the major components of tumor stromal tissue and are capable of eliciting diverse aspects of tumor growth as either a positive or negative regulator (1). In breast carcinoma, large numbers of infiltrating T cells and TAMs are often observed. The leukocyte infiltrate is found within the tumor stromal areas as well as in the epithelial areas that constitute the tumor mass (2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13). Recent reports suggest that the inflammatory reaction at the breast tumor site affects tumor growth and progression. Whereas lymphocytes have been shown to have divergent effects on development of breast cancer (2)(10)(11)(12)(13), it is widely accepted that high concentrations of TAMs are correlated with poor prognosis (2)(3)(4)(5)(6)(7)(8)(9)(10). Macrophage infiltration into tumors is regulated by several cytokines and chemokines, in particular macrophage chemoattractant protein-1 (MCP-1). MCP-1 is a member of the C-C chemokine family and possesses chemotactic activity for monocytes and T lymphocytes (14)(15)(16)(17). MCP-1 is produced not only by tumor cells, but also by stromal cells such fibroblasts, endothelial cells, and monocytes. MCP gene transfer enhances the metastatic potential of cancer cells with increased neovascularization, whereas MCP-1 itself activates monocyte cytostatic function against tumor cells (18)(19). Recently, several studies have focused on other chemokines and chemokine receptors in the susceptibility and progression of cancer (20), in particular …

Predisposition to atypical hemolytic uremic syndrome involves the concurrence of different susceptibility alleles in the regulators of complement activation gene cluster in 1q32

The efficiency of the complement system as an innate immune defense mechanism depends on a fine control that restricts its action to pathogens and prevents non-specific damage to host tissues. Genetic and functional analyses have shown that this critical control of complement activation may be impaired in atypical hemolytic uremic syndrome (aHUS) patients. Mutations in HF1, MCP or FI have been found in aHUS patients, but incomplete penetrance of the disease in individuals carrying these mutations is relatively frequent and no genetic defect has yet been found in a majority of aHUS patients. We report here the identification of a specific SNP haplotype block, spanning the MCP gene in the regulators of complement activation gene cluster, which is over-represented in aHUS patients and strongly associates with the severity of the disease. Linkage disequilibrium analyses suggest that this SNP haplotype also includes the CR1, DAF and C4BP genes. Initial studies identified two SNPs in the haplotype that influence the transcription activity of the MCP promoter in transient transfection experiments. Notably, the SNP haplotype block was found to be particularly frequent among patients who carry mutations in HF1, MCP or FI. These findings and the identification of aHUS patients carrying mutations in two complement regulatory genes provide an important insight into the etiology of aHUS. Together, they suggest that complement regulatory molecules act as a protein network and that multiple hits, involving plasma- and membrane-associated complement regulatory proteins, are necessary to impair protection to host tissues and to confer significant predisposition to aHUS.

Rhizobium leguminosarum methyl-accepting chemotaxis protein genes are down-regulated in the pea nodule.

Regulation of methyl-accepting chemotaxis protein (MCP) genes of Rhizobium leguminosarum was studied under symbiotic conditions. Transcriptional fusions using both beta-galactosidase and beta-glucuronidase genes within two different mcp genes demonstrated that mcp expression decreased significantly during nodulation. Immunoblots using an anti-MCP antibody detected MCPs in free-living cells but not in bacteroids. Down-regulation during nodulation was not dependent upon known regulatory proteins involved in induction of expression of genes involved in nitrogen fixation. Environmental conditions found in the bacteroid that may trigger down-regulation were investigated by growing free-living cultures under a variety of growth conditions. Growth under low oxygen concentration or using succinate as a sole carbon source did not lower expression of the mcp gene fusions.

Transgenic pigs expressing human decay-accelerating factor regulated by porcine MCP gene promoter.

Porcine membrane cofactor protein (pMCP) is abundantly expressed throughout the body with particularly strong expression on the vascular endothelia. Previous studies demonstrated that the promoter of the pMCP gene induced efficient expression of a human complement regulatory protein, decay-accelerating factor (DAF; CD55), in transgenic mice. In the present study, we tried to produce transgenic pigs with two hybrid genes, 0.9/hDAF and 5.4/hDAF, which were composed of human DAF (hDAF) gene regulated under pMCP promoters of different lengths (0.9 and 5.4 kb). Five live founder transgenic pigs were obtained only with the 0.9/hDAF construct. Although, four founder pigs transmitted the transgene to the second generation, the transmission rates varied among founders. We examined the expression of hDAF in tissues of descendants of two lines (Dm1 and Dm4). Human DAF specific RNAs were confirmed by an RT-PCR analysis in all organs examined. Levels of hDAF protein in the organs from the descendants of Dm1 line were higher than those in the corresponding human organs as determined by enzyme-linked immunosorbent assay. Immunohistochemical studies showed that the tissue distribution of hDAF in the descendants of both lines was similar to that of endogenous pMCP. The expression level of hDAF on the vascular endothelial cells in Dm1 line was twice that on the corresponding human cells. We tested whether proinflammatory cytokines upregulate an efficiency of pMCP promoter on hDAF expression in transgenic pigs. Although the expression of hDAF on the human endothelial cells increased with a combination of cytokines, tumor necrosis factor alpha and interferon-gamma, no cytokine-induced upregulation was seen in the cells of transgenic pigs. The endothelial cells from transgenic pigs exhibited high resistance to the human serum-mediated cytolysis.

Production of transgenic pigs expressing human DAF (CD55) regulated by the porcine MCP gene promoter

Production of transgenic pigs expressing human DAF (CD55) regulated by the porcine MCP gene promoter

Porcine MCP Gene Promoter Directs High Level Expression of Human DAF (CD55) in Transgenic Mice

Porcine membrane cofactor protein (pMCP), a complement regulatory protein, is widely expressed in various tissues. Particularly, it is highly expressed on vascular endothelium. The objective of this study was to investigate whether the pMCP gene promoter can induce efficient expression of a human complement regulatory protein, decay-accelerating factor (DAF; CD55) in transgenic mice. Two fragments of the 5’-flanking region of pMCP gene (0.9 kb and 5.4 kb) connected with human DAF minigene (0.9/hDAF and 5.4/hDAF) were used to produce transgenic mice. The expression of hDAF in heart, liver, kidney, lung, pancreas, brain and testis of the transgenic mice was examined by immunohistochemical analysis. The vascular endothelia and the nerves in all organs examined were intensely stained. The staining pattern in these tissues was similar in all transgenic mice examined regardless of the length of the promoters. The surface expression levels of hDAF on peripheral red blood cells and splenocytes from a mouse carrying 5.4/hDAF hemizygously was twice the level of expression on corresponding human cells. The red blood cells and splenocytes from the transgenic mice exhibited resistance to lysis by human serum in a manner dependent upon expressed hDAF level. The hearts from the transgenic mice functioned for a significantly longer time than those from normal mice under perfusion with human serum in the Langendorff perfusion system. These results demonstrated that the pMCP gene promoter is a good candidate of the regulatory element in the transgene to produce transgenic animals for xenotransplantation.

Membrane and secretory forms of mouse membrane cofactor protein (CD46) generated from a single gene through alternative splicing.

A cDNA encoding a new secretory form of mouse membrane cofactor protein (MCP, CD46) was identified additionally to the membrane form cDNA. The secretory MCP, predicted from the cDNA sequence, consisted of the conserved four short consensus repeats (SCRs) plus a four amino acid-stretch. Unlike human MCP which comprises many isoforms, mouse MCP cDNA predicted a single isoform of membrane MCP with cytoplasmic tail 1 (CYT1) and serine/ threonine-rich domain C (ST(C)). To clarify the genomic origin and monomorphic alteration of these cDNAs, we cloned and analyzed a mouse genomic DNA harboring the full coding sequence of MCP from a 129/SV mouse genomic library. The mouse Mcp was a single gene approximately 50 kilobases long. Eleven of the 14 coding exons of the human MCP gene and intron-exon boundary sequences were found to be conserved in the mouse gene. The STC homologue but not the STA or STB homologue in the mouse exons was functional: the latter being due to deletions and lack of consensus sequences for splicing. The sequence equivalent to cytoplasmic tail 2 (CYT2) has not been identified in the Mcp genome. Thus, the three exons (ST(A), ST(B), and probably CYT2) responsible for the polymorphism of human MCP by differential splicing were missing in the mouse Mcp gene. Unlike the case in humans, no Mcp-related genes or pseudogenes were observed in the mouse genome. The single mouse Mcp gene was mapped to the R-positive H5 band of mouse Chromosome 1 by FISH. Strikingly, one alternative exon with 73 base pairs (encoding the four new amino acids and a TGA stop codon) was discovered between the SCRIV and the STC exons; alternative splicing causes the generation of the secretory form of mouse MCP. These results on mouse MCP, together with the information concerning other mouse SCR proteins, infer that the regulator of complement activation (RCA) gene cluster is genetically diverged between humans and mice.

Vibriophage KVP40 and coliphage T4 genomes share a homologous 7-kbp region immediately upstream of the gene encoding the major capsid protein.

Vibriophage KVP40, a large tailed DNA phage morphologically similar to T-even coliphages, has a major capsid protein (Mcp) homologous to the equivalent protein, gp23(*), of coliphage T4. The sequence analysis was extended to a 7-kbp region immediately upstream of the mcp gene encoding the precursor of Mcp. The region as a whole was fairly homologous to the corresponding region of the T4 genome and contained 8 ORFs homologous to T4 genes 17, 18, 19, 20, 67, 68, 21, and 22 in the same order as in T4. These findings thus strongly suggest that these two phages are phylogenetically related.