Alteration of O-GlcNAcylation affects serine phosphorylation and regulates gene expression and activity of pyruvate kinase M2 in colorectal cancer cells

被引:36
作者
Chaiyawat, Parunya [1 ]
Chokchaichamnankit, Daranee [2 ]
Lirdprapamongkol, Kriengsak [1 ,2 ]
Srisomsap, Chantragan [1 ,2 ]
Svasti, Jisnuson [1 ,2 ]
Champattanachai, Voraratt [1 ,2 ]
机构
[1] Chulabhom Grad Inst, Appl Biol Sci Program, Bangkok, Thailand
[2] Chulabhorn Res Inst, Biochem Lab, Bangkok 10210, Thailand
关键词
colorectal cancer; hexosamine biosynthesis pathway; O-GlcNAcylation; phosphorylation; pyruvate kinase M2; PROTEOMIC ANALYSIS; PROTEINS; NUCLEAR; GLYCOSYLATION; SOLUBILITY; METABOLISM; DYNAMICS; GLYCINE; BREAST; GROWTH;
D O I
10.3892/or.2015.4178
中图分类号
R73 [肿瘤学];
学科分类号
100214 ;
摘要
O-GlcNAcylation is a dynamic post-translational modification that has extensive crosstalk with phosphorylation either at the same or adjacent sites of various proteins. We have previously reported that O-GlcNAcylation level was increased in primary breast and colorectal cancer, but the interplay of the two modifications remains unclear. Therefore, we explored crosstalk of the modifications by RNA interference against O-GlcNAc transferase (OGT) in colorectal cancer cells. Two-dimensional immunoblotting and mass spectrometric analysis showed that the levels of O-GlcNAc and senile phosphorylation of many proteins including serine hydroxymethyltransferase, cytokeratin-8, pyruvate kinase M2 (PKM2), heterogeneous nuclear ribonucleoprotein L, and lamin-B1, were reduced in siOGT cells compared to siScramble cells. In HT29 cells, immunoprecipitated PKM2 revealed decreased O-GlcNAc and serine phosphorylation levels after siOGT knockdown, but increased levels after treatment with Thiamet-G, an inhibitor of O-GlcNAcase (OGA). In addition, when global O-GlcNAcylation was enhanced by treating cells with Thiamet-G, PKM2 expression level was upregulated, but PKM2-specific activity was decreased. On the other hand, in OGT knockdown cells, PKM2 expression level was downregulated, but PKM2-specific activity was increased. Moreover, the metastatic colorectal cancer cells, SW620, had more O-GlcNAc-PKM2 and showed lower PKM2-specific activity compared to the non-metastatic colorectal cancer 5W480 cells. These results suggested roles of O-GlcNAcylation in modulating serine phosphorylation, as well as in regulating PKM2 activity and expression. Interfering levels of O-GIcNAcylation of PKM2 may be a novel target in controlling cancer metabolism and tumorigenesis of colorectal cancer.
引用
收藏
页码:1933 / 1942
页数:10
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