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Murine monocyte chemoattractant protein (MCP)-5: a novel CC chemokine that is a structural and functional homologue of human MCP-1.

Sarafi MN, Garcia-Zepeda EA, MacLean JA, Charo IF, Luster AD - J. Exp. Med. (1997)

Bottom Line: Consistent with these results, MCP-5 induced a calcium flux in human embryonic kidney (HEK)-293 cells transfected with human and murine CCR2, a CC chemokine receptor expressed on monocytes.MCP-5 did not induce a calcium flux in HEK-293 cells transfected with CCR1, CCR3, or CCR5.These data indicate that MCP-5 is a novel and potent monocyte active chemokine that is involved in allergic inflammation and the host response to pathogens.

View Article: PubMed Central - PubMed

Affiliation: Infectious Disease Unit, Massachusetts General Hospital, Boston, USA.

ABSTRACT
The chemokines are a large family of cytokines that control the recruitment of leukocytes in immune and inflammatory responses. We describe the isolation of a novel murine CC chemokine that, based on its biological and structural features, we have named monocyte chemoattractant protein (MCP)-5. MCP-5 mapped to the CC chemokine cluster on mouse chromosome 11 and was most closely related to human MCP-1 in structure (66% amino acid identity). Purified recombinant MCP-5 protein was a potent chemoattractant for peripheral blood monocytes, was only weakly active on eosinophils at high doses, and was inactive on neutrophils. MCP-5 induced a calcium flux in peripheral blood mononuclear cells, but not in purified murine eosinophils or neutrophils. Consistent with these results, MCP-5 induced a calcium flux in human embryonic kidney (HEK)-293 cells transfected with human and murine CCR2, a CC chemokine receptor expressed on monocytes. MCP-5 did not induce a calcium flux in HEK-293 cells transfected with CCR1, CCR3, or CCR5. Constitutive expression of MCP-5 mRNA was detected predominantly in lymph nodes, and its expression was markedly induced in macrophages activated in vitro and in vivo. Moreover, MCP-5 expression was up-regulated in the lungs of mice following aerosolized antigen challenge of sensitized mice, and during the host response to infection with Nippostrongylus brasiliensis. These data indicate that MCP-5 is a novel and potent monocyte active chemokine that is involved in allergic inflammation and the host response to pathogens.

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Genomic organization, chromosomal mapping, nucleotide  sequence, and predicted amino acid sequence of the murine MCP-5 gene.  (A) Partial restriction map and genomic organization of the mouse MCP-5  gene. The mature mRNA is shown schematically below with the positions of the start codon (ATG) and stop codon (TGA). The scales are  shown below each drawing. (B) SSCP polymorphism. PCR primers in  intron 2 and exon 3, indicated by arrows in A, were used to amplify genomic DNA from C57BL/6J, Mus spretus, or the C57BL/6J × M. spretus  heterozygote, and analyzed using SSCP. The complete raw data for this  cross with references and notes are available on the World Wide Web at  http://www.jax.org/resources/documents/cmdata/BSS11data.html and the  Mouse Genome Database accession number is MGD-CREX-697. (C)  MCP-5 cDNA sequence. The filled triangles indicate the intron/exon  borders. The single underline indicate the ATTTA sequences that have  been reported to decrease mRNA stability. The double underlined sequence indicates the predicted polyadenylation signal. The sequence has  been deposited in GenBank/EMBL/DDBJ under the accession U66670.  The arrow indicates the predicted site for a signal peptidase cleavage.
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Figure 1: Genomic organization, chromosomal mapping, nucleotide sequence, and predicted amino acid sequence of the murine MCP-5 gene. (A) Partial restriction map and genomic organization of the mouse MCP-5 gene. The mature mRNA is shown schematically below with the positions of the start codon (ATG) and stop codon (TGA). The scales are shown below each drawing. (B) SSCP polymorphism. PCR primers in intron 2 and exon 3, indicated by arrows in A, were used to amplify genomic DNA from C57BL/6J, Mus spretus, or the C57BL/6J × M. spretus heterozygote, and analyzed using SSCP. The complete raw data for this cross with references and notes are available on the World Wide Web at http://www.jax.org/resources/documents/cmdata/BSS11data.html and the Mouse Genome Database accession number is MGD-CREX-697. (C) MCP-5 cDNA sequence. The filled triangles indicate the intron/exon borders. The single underline indicate the ATTTA sequences that have been reported to decrease mRNA stability. The double underlined sequence indicates the predicted polyadenylation signal. The sequence has been deposited in GenBank/EMBL/DDBJ under the accession U66670. The arrow indicates the predicted site for a signal peptidase cleavage.

Mentions: To isolate novel mouse CC chemokine genes, a murine genomic library was sequentially screened with a human MCP-4 cDNA probe under conditions of low stringency and a mouse JE cDNA probe under conditions of high stringency. 15 plaques were identified that hybridized more strongly with the human MCP-4 probe than with the murine JE probe. These plaques were purified and further analyzed by PCR using a set of degenerate primers. A PCR product was generated from five of these clones, and all contained the same novel sequence that had a high degree of homology with the MCP subfamily. Southern blot analysis of mouse genomic DNA established a restriction map (Fig. 1 A) and revealed that MCP-5 was a single copy gene (data not shown). One of the five overlapping MCP-5 genomic clones was partially sequenced to determine the intron/exon structure of the MCP-5 gene (Fig. 1 A), and to confirm the sequence of the PCR products. Like other CC chemokines, the MCP-5 gene contained three exons and two introns.


Murine monocyte chemoattractant protein (MCP)-5: a novel CC chemokine that is a structural and functional homologue of human MCP-1.

Sarafi MN, Garcia-Zepeda EA, MacLean JA, Charo IF, Luster AD - J. Exp. Med. (1997)

Genomic organization, chromosomal mapping, nucleotide  sequence, and predicted amino acid sequence of the murine MCP-5 gene.  (A) Partial restriction map and genomic organization of the mouse MCP-5  gene. The mature mRNA is shown schematically below with the positions of the start codon (ATG) and stop codon (TGA). The scales are  shown below each drawing. (B) SSCP polymorphism. PCR primers in  intron 2 and exon 3, indicated by arrows in A, were used to amplify genomic DNA from C57BL/6J, Mus spretus, or the C57BL/6J × M. spretus  heterozygote, and analyzed using SSCP. The complete raw data for this  cross with references and notes are available on the World Wide Web at  http://www.jax.org/resources/documents/cmdata/BSS11data.html and the  Mouse Genome Database accession number is MGD-CREX-697. (C)  MCP-5 cDNA sequence. The filled triangles indicate the intron/exon  borders. The single underline indicate the ATTTA sequences that have  been reported to decrease mRNA stability. The double underlined sequence indicates the predicted polyadenylation signal. The sequence has  been deposited in GenBank/EMBL/DDBJ under the accession U66670.  The arrow indicates the predicted site for a signal peptidase cleavage.
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Related In: Results  -  Collection

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Figure 1: Genomic organization, chromosomal mapping, nucleotide sequence, and predicted amino acid sequence of the murine MCP-5 gene. (A) Partial restriction map and genomic organization of the mouse MCP-5 gene. The mature mRNA is shown schematically below with the positions of the start codon (ATG) and stop codon (TGA). The scales are shown below each drawing. (B) SSCP polymorphism. PCR primers in intron 2 and exon 3, indicated by arrows in A, were used to amplify genomic DNA from C57BL/6J, Mus spretus, or the C57BL/6J × M. spretus heterozygote, and analyzed using SSCP. The complete raw data for this cross with references and notes are available on the World Wide Web at http://www.jax.org/resources/documents/cmdata/BSS11data.html and the Mouse Genome Database accession number is MGD-CREX-697. (C) MCP-5 cDNA sequence. The filled triangles indicate the intron/exon borders. The single underline indicate the ATTTA sequences that have been reported to decrease mRNA stability. The double underlined sequence indicates the predicted polyadenylation signal. The sequence has been deposited in GenBank/EMBL/DDBJ under the accession U66670. The arrow indicates the predicted site for a signal peptidase cleavage.
Mentions: To isolate novel mouse CC chemokine genes, a murine genomic library was sequentially screened with a human MCP-4 cDNA probe under conditions of low stringency and a mouse JE cDNA probe under conditions of high stringency. 15 plaques were identified that hybridized more strongly with the human MCP-4 probe than with the murine JE probe. These plaques were purified and further analyzed by PCR using a set of degenerate primers. A PCR product was generated from five of these clones, and all contained the same novel sequence that had a high degree of homology with the MCP subfamily. Southern blot analysis of mouse genomic DNA established a restriction map (Fig. 1 A) and revealed that MCP-5 was a single copy gene (data not shown). One of the five overlapping MCP-5 genomic clones was partially sequenced to determine the intron/exon structure of the MCP-5 gene (Fig. 1 A), and to confirm the sequence of the PCR products. Like other CC chemokines, the MCP-5 gene contained three exons and two introns.

Bottom Line: Consistent with these results, MCP-5 induced a calcium flux in human embryonic kidney (HEK)-293 cells transfected with human and murine CCR2, a CC chemokine receptor expressed on monocytes.MCP-5 did not induce a calcium flux in HEK-293 cells transfected with CCR1, CCR3, or CCR5.These data indicate that MCP-5 is a novel and potent monocyte active chemokine that is involved in allergic inflammation and the host response to pathogens.

View Article: PubMed Central - PubMed

Affiliation: Infectious Disease Unit, Massachusetts General Hospital, Boston, USA.

ABSTRACT
The chemokines are a large family of cytokines that control the recruitment of leukocytes in immune and inflammatory responses. We describe the isolation of a novel murine CC chemokine that, based on its biological and structural features, we have named monocyte chemoattractant protein (MCP)-5. MCP-5 mapped to the CC chemokine cluster on mouse chromosome 11 and was most closely related to human MCP-1 in structure (66% amino acid identity). Purified recombinant MCP-5 protein was a potent chemoattractant for peripheral blood monocytes, was only weakly active on eosinophils at high doses, and was inactive on neutrophils. MCP-5 induced a calcium flux in peripheral blood mononuclear cells, but not in purified murine eosinophils or neutrophils. Consistent with these results, MCP-5 induced a calcium flux in human embryonic kidney (HEK)-293 cells transfected with human and murine CCR2, a CC chemokine receptor expressed on monocytes. MCP-5 did not induce a calcium flux in HEK-293 cells transfected with CCR1, CCR3, or CCR5. Constitutive expression of MCP-5 mRNA was detected predominantly in lymph nodes, and its expression was markedly induced in macrophages activated in vitro and in vivo. Moreover, MCP-5 expression was up-regulated in the lungs of mice following aerosolized antigen challenge of sensitized mice, and during the host response to infection with Nippostrongylus brasiliensis. These data indicate that MCP-5 is a novel and potent monocyte active chemokine that is involved in allergic inflammation and the host response to pathogens.

Show MeSH
Related in: MedlinePlus