Background: Neoplasia can be driven by mutations resulting in dysregulation of transcription. In the mesenchymal neoplasm, aggressive fibromatosis, subtractive hybridization identified sterile alpha motif domain 9 (SAMD9) as a substantially down regulated gene in neoplasia. SAMD9 was recently found to be mutated in normophosphatemic familial tumoral calcinosis. In this study, we studied the gene structure and function of SAMD9, and its paralogous gene, SAMD9L, and examined these in a variety of species.
We have investigated the possibility that the intestinal mucin rat Muc2 forms dimers during biosynthesis via intermolecular disulphide bridging of its C-terminal domains. Since the cysteine alignment of RMuc2 (and other secretory mucins) is similar to that of human von Willebrand factor, a similar C-tail to C-tail dimerization may occur in mucins. The C-terminal domain of RMuc2 (534 amino acids) was expressed in COS-1 cells, and the products monitored by SDS/PAGE and western blotting with three antibodies to different regions of the C-terminal domain. In cells, the expressed domain was glycosylated and formed disulphide-dependent dimers centred at approximately 150 kDa. The domain dimer, but not its precursor monomer, was secreted into the culture medium. The dimers in the media however, appeared to be 12Ϫ15-kDa heavier (i.e. had a slower mobility) than in cell lysates. Initial N-glycosylation, dimerization and secretion were inhibited by addition of tunicamycin to incubations, whereas benzyl-A-GalNAc did not interfere with these processes. However benzyl-A-GalNAc resulted in a decrease in the apparent size of secreted dimers, such that they now had the same mobility on gels as dimers normally seen in cell lysates (i.e. 150 kDa). A similar change in dimer size was observed after incubating untreated media samples with N-acetylneuraminidase. This suggests that benzyl-A-GalNAc caused inhibition of sialylation of cell dimers just before they were secreted. In summary, the C-terminal domain of RMuc2 can form disulphide-dependent dimers, and N-glycosylation is required for dimerization and subsequent secretion. A late sialylation event appears to precede the secretion of mucin domain dimers.Keywords : mucin ; Muc2; disulfide ; dimer; cystine knot.The internal epithelial surfaces of most organisms are cov-coprotein von Willebrand factor [6,8]. Other secretory mucins ered by a gel that is rich in secreted mucus glycoproteins (mu-show a similar cysteine distribution throughout their C-termini, cins). The gel lubricates the mucosal surface, forms a barrier to including frog integumentary mucin B.1 [9], bovine submaxilvarious chemicals, proteases, toxins and infectious agents, and is lary mucin [10], porcine submaxillary mucin [11] and human secreted in response to a wide variety of stimuli [1Ϫ4]. Several lung mucin MUC5AC [12,13]. The similarity includes a region domains of mucin molecules contribute to their function. The termed the 'cystine knot' motif, which is also present and plays protein core (or apomucin) usually consists centrally of a vari-an active role in the dimerization of transforming growth factorable number of imperfect tandem repeats which contribute to β2 [14], nerve-growth factor [15], Norrie disease protein [16, molecular rigidity because of extensive O-glycosylation of their 17], human chorionic gonadotropin [18] and glial cell line-deserine and threonine residues. The flanking end regions of most rived neurotrophic factor [19]. The involvement of the cystine secretory mucins are enriched in cystein...
Long range physical mapping within the p21 region of the X chromosome identified a CpG rich island approximately 180 kb centromeric to the chronic granulomatous disease (CGD) locus. The segments adjacent to the CpG island hybridized to discrete bands in DNAs of several species and when used to screen retinal cDNA libraries led to the identification of cDNAs that detected a mRNA of 2.1 kb in many tissues. Molecular characterization of corresponding genomic clones of this novel human gene confirmed the origin of the cDNA clones and indicated a genomic structure with five exons spanning a total of 9 kb. The complete cDNA sequence revealed that this gene contained a putative open reading frame of 116 amino acids with a 3' untranslated region of 1.74 kb. The amino acid sequence shows a high degree of similarity to the predicted product of the tctex-1 gene of the mouse t complex. As linkage studies and patients with deletions have implicated the Xp21 region as containing the retinitis pigmentosa defect (RP3), the gene was assessed as a candidate disease gene in RP3 families. A single base pair polymorphism was identified within the coding region but no disease associated changes were found by single strand conformational polymorphism and sequencing analysis of amplified exons of 20 RP patients. Analysis of a dinucleotide repeat polymorphism within this gene in families affected with RP3 suggested refinement of the RP3 region.
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