Molecular cloning and characterization of chemokine-like factor 1 (CKLF1), a novel human cytokine with unique structure and potential chemotactic activity.
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Cytokines are small proteins that have an essential role in the immune and inflammatory responses. The repertoire of cytokines is becoming diverse and expanding. Here we report the identification and characterization of a novel cytokine designated as chemokine-like factor 1 (CKLF1). The full-length cDNA of CKLF1 is 530 bp long and a single open reading frame encoding 99 amino acid residues. CKLF1 bears no significant similarity to any other known cytokine in its amino acid sequence. Expression of CKLF1 can be partly inhibited by interleukin 10 in PHA-stimulated U937 cells. Recombinant CKLF1 is a potent chemoattractant for neutrophils, monocytes and lymphocytes; moreover, it can stimulate the proliferation of murine skeletal muscle cells. These results suggest that CKLF1 might have important roles in inflammation and in the regeneration of skeletal muscle. (+info)
Proteolipid protein 2 mRNA is expressed in the rabbit embryo during gastrulation.
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Differential display technology applied to rabbit blastocysts identified an mRNA that encodes a motif similar to that of the proteolipid protein PLP2/A4 of man, mouse and sheep. The open reading frame (456bp) has 88% amino acid identity to human PLP2/A4. The gene is maximally expressed at the beginning of gastrulation: in situ hybridizations exhibited a sickle-shaped area of labelling at the posterior pole of day 7 post-coitum embryos, which appeared at day 6.5 and decreased in size up to day 8. Weaker labelling was found in the extraembryonic mesoderm, in the anterior part of the primitive streak and in the trophoblast. Time and site of gene expression coincide with emerging morphogenetic activities at the posterior pole of the embryo at the beginning of gastrulation. (+info)
Overexpression of chemokine-like factor 2 promotes the proliferation and survival of C2C12 skeletal muscle cells.
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Chemokine-like factor 1 (CKLF1) is a novel cytokine first cloned from U937 cells. It contains different splicing forms and has chemotactic effects on a wide spectrum of cells both in vitro and in vivo; it can also stimulate the regeneration of skeletal muscle cells in vivo, but the mechanism remains unclear. To probe the myogenesis function of CKLF2, which is the largest isoform of CKLFs, C2C12 murine myoblasts were stably transfected with human CKLF2 eukaryotic expression vector. Compared with control vector transfected C2C12 cells, CKLF2 overexpression causes accelerated myoblast proliferation as determined by cell counting and [(3)H]TdR incorporation assays. In addition, CKLF2 overexpression also promotes cell differentiation, which was determined by higher expression levels of myogenin, creatine kinase, myosin and the accelerated myoblast fusion. Further analysis also indicates that CKLF2 could activate the transcription activity of the bHLH/MyoD and MEF2 families. Finally, DNA synthesis and myotube formation could also be promoted by growing C2C12 cells in conditioned media from CKLF2-transfected cells. These findings strongly suggest a role for human CKLF2 in regulation of skeletal muscle myogenesis. (+info)
Uncleaved BAP31 in association with A4 protein at the endoplasmic reticulum is an inhibitor of Fas-initiated release of cytochrome c from mitochondria.
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BAP31 is a polytopic integral protein of the endoplasmic reticulum membrane and, like BID, is a preferred substrate of caspase-8. Upon Fas/CD95 stimulation, BAP31 is cleaved within its cytosolic domain, generating proapoptotic p20 BAP31. In human KB epithelial cells expressing the caspase-resistant mutant crBAP31, Fas stimulation resulted in cleavage of BID and insertion of BAX into mitochondrial membrane, but subsequent oligomerization of BAX and BAK, egress of cytochrome c to the cytosol, and apoptosis were impaired. Bap31-null mouse cells expressing crBAP31 cannot generate the endogenous p20 BAP31 cleavage product, yet crBAP31 conferred resistance to cellular condensation and cytochrome c release in response to activation of ectopic FKBPcasp8 by FK1012z. Full-length BAP31, therefore, is a direct inhibitor of these caspase-8-initiated events, acting independently of its ability to sequester p20, with which it interacts. Employing a novel split ubiquitin yeast two-hybrid screen for BAP31-interacting membrane proteins, the putative ion channel protein of the endoplasmic reticulum, A4, was detected and identified as a constitutive binding partner of BAP31 in human cells. Ectopic A4 that was introduced into A4-deficient cells cooperated with crBAP31 to resist Fas-induced egress of cytochrome c from mitochondria and cytoplasmic apoptosis. (+info)
Chemokine-like factor 1, a novel cytokine, contributes to airway damage, remodeling and pulmonary fibrosis.
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BACKGROUND: Chemokine-like factor 1 (CKLF1) was recently identified as a novel cytokine. The full-length CKLF1 cDNA contains 530 bp encoding 99 amino acid residues with a CC motif similar to that of other CC family chemokines. Recombinant CKLF1 exhibits chemotactic activity on leucocytes and stimulates proliferation of murine skeletal muscle cells. We questioned whether CKLF1 could be involved in the pathogenesis of inflammation and proliferation in the lung. Therefore we used efficient in vivo gene delivery method to investigate the biological effect of CKLF1 in the murine lung. METHODS: CKLF1-expressing plasmid, pCDI-CKLF1, was constructed and injected into the skeletal muscles followed by electroporation. Lung tissues were obtained at the end of week 1, 2, 3 and 4 respectively after injection. The pathological changes in the lungs were observed by light microscope. RESULTS: A single intramuscular injection of CKLF1 plasmid DNA into BALB/c mice caused dramatic pathological changes in the lungs of treated mice. These changes included peribronchial leukocyte infiltration, epithelial shedding, collagen deposition, proliferation of bronchial smooth muscle cells and fibrosis of the lung. CONCLUSIONS: The sustained morphological abnormalities of the bronchial and bronchiolar wall, the acute pneumonitis and interstitial pulmonary fibrosis induced by CKLF1 were similar to phenomena observed in chronic persistent asthma, acute respiratory distress syndrome and severe acute respiratory syndrome. These data suggest that CKLF1 may play an important role in the pathogenesis of these important diseases and the study also implies that gene electro-transfer in vivo could serve as a valuable approach for evaluating the function of a novel gene in animals. (+info)
Regulation of EGF receptor signaling by the MARVEL domain-containing protein CKLFSF8.
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It is known that chemokine-like factor superfamily 8 (CKLFSF8), a member of the CKLF superfamily, has four putative transmembrane regions and a MARVEL domain. Its structure is similar to TM4SF11 (plasmolipin) and widely distributed in normal tissue. However, its function is not yet known. We show here that CKLFSF8 is associated with the epidermal growth factor receptor (EGFR) and that ectopic expression of CKLFSF8 in several cell lines suppresses EGF-induced cell proliferation, whereas knockdown of CKLFSF8 by siRNA promotes cell proliferation. In cells overexpressing CKLFSF8, the initial activation of EGFR was not affected, but subsequent desensitization of EGF-induced signaling occurred rapidly. This attenuation was correlated with an increased rate of receptor endocytosis. In contrast, knockdown of CKLFSF8 by siCKLFSF8 delayed EGFR endocytosis. These results identify CKLFSF8 as a novel regulator of EGF-induced signaling and indicate that the association of EGFR with four transmembrane proteins is critical for EGFR desensitization. (+info)
Characterization and expression profile of CMTM3/CKLFSF3.
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CMTM/CKLFSF is a novel family of proteins linking chemokines and TM4SF. In humans, these proteins are encoded by nine genes, CKLF and CMTM1-8/CKLFSF1-8. Here we report the characteristics and expression profile of CMTM3/CKLFSF3. Human CMTM3/CKLFSF3 has a high sequence identity among various species and similar characteristics as its mouse and rat homologues. Established by results both of RT-PCR and Quantitative Real-time PCR, the gene is highly transcribed in testis, leukocytes and spleen. For further verification, we generated a polyclonal antibody against human CMTM3/CKLFSF3 and found that the protein is highly expressed in the testis and some cells of PBMCs. Therefore, CMTM3/CKLFSF3 is an evolutionarily conserved gene that may have important roles in the male reproductive system and immune system. Further studies are necessary to validate its functions in the two systems. (+info)
Expression and localization of CKLFSF2 in human spermatogenesis.
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AIM: To investigate the expression and subcellular localization of chemokine-like factor superfamily 2 (CKLFSF2) in human testis and its potential role in spermatogenesis. METHODS: A specific polyclonal antibody against CKLFSF2 was raised. The expression and cellular localization of CKLFSF2 in the seminiferous tubules was checked by immunohistochemistry method. Also, in situ hybridization was applied to localize the mRNA distribution. The EGFP-CKLFSF2 fusion protein was expressed in COS-7 cells to localize its subcellular location in vitro. In addition, the abnormal expression of CKLFSF2 in testes of patients with male infertility was assayed by reverse transcription polymerase chain reaction (RT-PCR) and immunohistochemistry methods. RESULTS: Having a close correlation with spermatogenesis defects, CKLFSF2 was specifically expressed in meiotic and post-meiotic germ cells, which were localized to the endoplasmic reticulum (ER) near the Golgi apparatus. CONCLUSION: CKLFSF2 could play important roles in the process of meiosis and spermiogenesis, and might be involved in the vesicular transport or membrane apposition events in the endoplasmic reticulum. (+info)