Disease relevance of Clgn

• The defective zona pellucida-adhesion phenotype of sperm from calmegin-deficient mice is reminiscent of certain cases of unexplained infertility in human males [1].
• The putative chaperone Calmegin is required for sperm fertility in mouse and the relevance of the gene to certain cases of human male infertility has been suggested [2].
• The cDNA microarray analysis of the human fibrosarcoma cells treated with trichostatin A (TSA) showed an increased level of calmegin mRNA [3].
• Analysis of short oligo(A) fragments in synchronized L5178y mouse lymphoma cells after labeling with [3H]Ado revealed that the percentage of A2-6 sequences on the total radioactivity amounted in S-phase cells to 1.6%, while the value obtained for the stationary L-cell system was 8.0% [4].

High impact information on Clgn

• Here we show that calmegin binds to nascent polypeptides during spermatogenesis, and have analysed its physiological function by targeted disruption of its gene [1].
• These results suggest that spermatogenic cell endoplasmic reticulum has a unique calcium binding protein, calnexin-t, which appears to be a calnexin variant [5].
• Molecular cloning and sequencing of calnexin-t. An abundant male germ cell-specific calcium-binding protein of the endoplasmic reticulum [5].
• Molecular cloning of a novel Ca(2+)-binding protein (calmegin) specifically expressed during male meiotic germ cell development [6].
• Furthermore, we demonstrated the 45Ca2+ binding ability of this antigen by a 45Ca2+ overlay assay, and the name calmegin is proposed for this antigen [6].

Biological context of Clgn

• Acid phosphatase treatment of the 101-kDa form generated a 93-kDa band that in turn could be recovered to the 101-kDa form by incubation with HeLa cell S100 fraction, indicating that the 101-kDa form is a phosphorylated type of calnexin-t. The sites of phosphorylation were shown to be restricted to the cytoplasmic domain [7].
• Our results suggest that the structure of the ER luminal domain of calnexin-t is likely to differ in middle pachytene versus haploid germ cell phases [7].
• Using the fluorescence in situ hybridization (FISH) technique, human Calmegin gene was mapped to chromosome 4q28.3-q31.1 [2].
• This suggests that calmegin is not required for the morphogenesis of male germ cells [8].
• A switch in the phosphorylation state of the dimeric form of the Meg1 protein correlates with progression through meiosis in the mouse [9].

Anatomical context of Clgn

• This diminished amount of ADAM3 in the Triton X-114 detergent-enriched phase may explain the inability of Clgn(-/-) and Ace(-/-) sperm to bind to the zona pellucida [10].
• Since CLGN is a molecular chaperone for membrane transport of target proteins and ACE is a membrane protein, we looked for ACE on the sperm membranes from Clgn(-/-) mice [10].
• Loss of the endoplasmic reticulum resident chaperone calmegin leads to the production of sterile sperm that do not bind to the egg zona pellucida (M. Ikawa et al., 1997, Nature 387, 607-611) [11].
• In the present study, we demonstrate that calmegin -/- sperm were defective in migrating into the oviducts and in binding to the egg plasma membrane [11].
• Despite the impaired adhesive function, calmegin -/- sperm could fertilize eggs when zonae pellucidae were partially dissected, and eggs fertilized in this manner could develop normally to term [11].

Associations of Clgn with chemical compounds

• Male mice deficient for the calmegin (Clgn) or the angiotensin-converting enzyme (Ace) gene show impaired sperm migration into the oviduct and loss of sperm-zona pellucida binding ability in vitro [10].
• The induction of calmegin mRNA by TSA was added by the treatment with 5-aza-2'-deoxycytidine (5'Aza-dC), implying that epigenetic alterations are involved in the transcriptional repression of the gene [3].

Physical interactions of Clgn

• Gel-shift and yeast one-hybrid experiments showed that Tcfl5 interacts with a non-canonical CACGCG site that is present in the Clgn promoter [12].

Other interactions of Clgn

• Calnexin-t (calmegin) is a male germ cell-specific variant of calnexin, a membrane bound-molecular chaperone in the endoplasmic reticulum (ER) [7].
• This screening resulted in the identification of mouse Tcfl5 as a candidate Clgn promoter-binding protein [12].
• Because fertilin beta is absent from calmegin-deficient mice, these results also suggest that the role of fertilin beta in sperm-EPM interaction needs to be reevaluated [13].

Analytical, diagnostic and therapeutic context of Clgn

• Using immunoprecipitation techniques, calmegin was found to bind to sperm membrane proteins, fertilin alpha and beta, during spermatogenesis [11].
• Light microscopic immunocytochemistry showed that calmegin appeared in early pachytene spermatocytes, with the highest expression in round and elongating spermatids, and disappeared in the maturation phase of spematids at step 15 [8].
• To further characterize calmegin, we analyzed the precise stage of expression and the intracellular localization of this protein in germ cells during mouse spermatogenesis by an immunoperoxidase technique using the anti-calmegin monoclonal antibody TRA369 [8].
• Western blot analysis of immunoprecipitated Meg1 protein revealed multiple bands (in the range of M(r) 12,000-18,000), some of which where recognized by anti-phosphotyrosine antibodies, suggesting that in vivo, Meg1 appears in multiple phosphorylated forms [9].

References