Novel intestinal splice variants of RNA-binding protein CUGBP2: isoform-specific effects on mitotic catastrophe.
CUG triplet repeat-binding protein 2 (CUGBP2) is a RNA-binding protein that regulates mRNA translation and modulates apoptosis. Here, we report the identification of two splice variants (termed variants 2 and 3) in cultured human intestinal epithelial cells and in mouse gastrointestinal tract. The variants are generated from alternative upstream promoters resulting in the inclusion of additional NH(2)-terminal residues. Although variant 2 is the predominant isoform in normal intestine, its expression is reduced, whereas variant 1 is overexpressed following gamma-irradiation. All three variants bind cyclooxygenase-2 (COX-2) mRNA. However, only variant 1 inhibits the translation of the endogenous COX-2 mRNA and a chimeric luciferase mRNA containing the COX-2 3'untranslated region. Furthermore, whereas variant 1 is predominantly nuclear, variants 2 and 3 are predominantly cytoplasmic. These data imply that the additional amino acids affect CUGBP2 function. Previous studies have demonstrated that variant 1 induces intestinal epithelial cells to undergo apoptosis. However, in contrast to variant 1, the two novel variants do not affect proliferation or apoptosis of HCT116 cells. In addition, only variant 1 induced G(2)/M cell cycle arrest, which was overcome by prostaglandin E(2). Moreover, variant 1 increased cellular levels of phosphorylated p53 and Bax and decreased Bcl2. Caspase-3 and -9 were also activated, suggesting the initiation of the intrinsic apoptotic pathway. Furthermore, increased phosphorylation of checkpoint kinase (Chk)1 and Chk2 kinases and increased nuclear localization of Cdc2 and cyclin B1 suggested that cells were in mitotic transition. Taken together, these data demonstrate that cells expressing CUGBP2 variant 1 undergo apoptosis during mitosis, suggesting mitotic catastrophe.
Ramalingam
Satish
S
Dept. of Medicine, Univ. of Oklahoma Health Sciences Ctr., 920 Stanton L. Young Blvd., WP1360, Oklahoma City, OK 73126, USA.
Natarajan
Gopalan
G
Schafer
Chris
C
Subramaniam
Dharmalingam
D
May
Randal
R
Ramachandran
Ilangovan
I
Queimado
Lurdes
L
Houchen
Courtney W
CW
Anant
Shrikant
S
D017398:Alternative Splicing / Q000528:radiation effects; D000818:Animals; D017209:Apoptosis / Q000235:genetics; D000067878:CELF Proteins; D002453:Cell Cycle / Q000235:genetics; D018797:Cell Cycle Proteins / Q000378:metabolism; D003110:Colonic Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D051546:Cyclooxygenase 2 / Q000235:genetics / Q000378:metabolism; D015232:Dinoprostone / Q000172:deficiency / Q000378:metabolism; D004847:Epithelial Cells / Q000378:metabolism* / Q000473:pathology / Q000528:radiation effects; D015972:Gene Expression Regulation, Neoplastic; D045325:HCT116 Cells; D019073:HT29 Cells; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism*; D007422:Intestines / Q000473:pathology / Q000528:radiation effects; D051379:Mice; D008810:Mice, Inbred C57BL; D008938:Mitosis / Q000235:genetics; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D014176:Protein Biosynthesis; D020033:Protein Isoforms / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
D000067878:CELF Proteins; C117363:CELF2 protein, human; D018797:Cell Cycle Proteins; D009419:Nerve Tissue Proteins; D020033:Protein Isoforms; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human; D015232:Dinoprostone