Human core 1 beta3-galactosyltransferase (T-synthase) generates the core 1 O-glycan Galbeta1 3GalNAc-alpha1-Ser/Thr or T-antigen, which is a precursor for many extended O-glycans in animal glycoproteins. Lack of T-synthase activity leads to expression of the Tn antigen GalNAc-alpha 1 -Ser/Thr, a common tumor associated antigen. This application arises from our discovery that T-synthase activity in cells requires the co-expression of a unique protein that we termed Cosmc (COre 1 beta3-Gal-T Specific Molecular Chaperone). Cosmc may be a unique chaperone primarily involved in folding and maturation of T-synthase. Cosmc is X-linked (Xq23) and its Cdna predicts a 318 amino acid transmembrane protein (approximately 36.4 kDa polypeptide size) with type-II membrane topology. Both human lymphoblastoid Jurkat and LSC colon carcinoma cell lines contain a normal gene and mRNA encoding T-synthase, but lack T-synthase activity. However, both cell lines contain a mutated, non-functional Cosmc;expression of wild-type Cosmc cDNA in both cell lines restores T-synthase activity and T-antigen expression. In cells lacking Cosmc, newly synthesized T-synthase is targeted to proteasomes for degradation. Cosmc binds ATP, consistent with a chaperone function, and Cosmc is primarily localized in the ER and Tsynthase is primarily in the Golgi. We hypothesize that Cosmc acts as a specific molecular chaperone in assisting the folding, stability, and/or targeting of T-synthase. To test this hypothesis we propose three specific aims.
Aim 1 - We will define the subcellular localization of Cosmc and T-synthase and explore the mechanisms of Cosmc localization and its potential chaperone/escort function.
Aim 2 - We will define whether Cosmc assists other glycosyltransferases and define its potential interaction with T-synthase and/or other proteins (e.g. chaperones) and ATP.
Aim 3 - To define other possible biological functions of Cosmc and directly test our hypothesis, we will generate mice containing conventional and targeted, endothelial cell-specific deletions of Cosmc. An understanding of the structure and function of Cosmc should provide important new insights into the molecular basis of several human diseases, including IgA nephropathy, Tn-Syndrome, Henoch-Schonlein purpura, and malignant transformation associated with Tn antigen expression and lack of T-synthase activity.
|Hanes, Melinda S; Moremen, Kelley W; Cummings, Richard D (2017) Biochemical characterization of functional domains of the chaperone Cosmc. PLoS One 12:e0180242|
|Aryal, Rajindra P; Ju, Tongzhong; Cummings, Richard D (2014) Identification of a novel protein binding motif within the T-synthase for the molecular chaperone Cosmc. J Biol Chem 289:11630-41|
|Aryal, Rajindra P; Ju, Tongzhong; Cummings, Richard D (2012) Tight complex formation between Cosmc chaperone and its specific client non-native T-synthase leads to enzyme activity and client-driven dissociation. J Biol Chem 287:15317-29|
|Wang, Yingchun; Jobe, Shawn M; Ding, Xiaokun et al. (2012) Platelet biogenesis and functions require correct protein O-glycosylation. Proc Natl Acad Sci U S A 109:16143-8|
|Ju, Tongzhong; Xia, Baoyun; Aryal, Rajindra P et al. (2011) A novel fluorescent assay for T-synthase activity. Glycobiology 21:352-62|
|Sun, Qian; Ju, Tongzhong; Cummings, Richard D (2011) The transmembrane domain of the molecular chaperone Cosmc directs its localization to the endoplasmic reticulum. J Biol Chem 286:11529-42|
|Ju, Tongzhong; Otto, Vivianne I; Cummings, Richard D (2011) The Tn antigen-structural simplicity and biological complexity. Angew Chem Int Ed Engl 50:1770-91|
|Wang, Yingchun; Ju, Tongzhong; Ding, Xiaokun et al. (2010) Cosmc is an essential chaperone for correct protein O-glycosylation. Proc Natl Acad Sci U S A 107:9228-33|
|Ju, Tongzhong; Cummings, Richard D (2010) Functional assays for the molecular chaperone cosmc. Methods Enzymol 479:107-22|
|Aryal, Rajindra P; Ju, Tongzhong; Cummings, Richard D (2010) The endoplasmic reticulum chaperone Cosmc directly promotes in vitro folding of T-synthase. J Biol Chem 285:2456-62|
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