HuR Regulates Alternative Splicing of the TRA2β Gene in Human Colon Cancer Cells under Oxidative Stress

Hu antigen R (HuR) regulates stress responses through stabilizing and/or facilitating the translation of target mRNAs. The humanTRA2 β gene encodes splicing factor transformer 2β (Tra2β) and generates 5 mRNA isoforms (TRA2β1 to -5 ) through alternative splicing. Exposure of HCT116 colon cancer cells to sodium arsenite stimulated checkpoint kinase 2 (Chk2)- and mitogen-activated protein kinase p38 (p38MAPK )-mediated phosphorylation of HuR at positions S88 and T118. This induced an association between HuR and the 39-nucleotide (nt) proximal region ofTRA2 β exon 2, generating aTRA2β4 mRNA that includes exon 2, which has multiple premature stop codons. HuR knockdown or Chk2/p38MAPK double knockdown inhibited the arsenite-stimulated production ofTRA2β4 and increased Tra2β protein, facilitating Tra2β-dependent inclusion of exons in target pre-mRNAs. The effects of HuR knockdown or Chk2/p38MAPK double knockdown were also confirmed using aTRA2 β minigene spanning exons 1 to 4, and the effects disappeared when the 39-nt region was deleted from the minigene. In endogenous HuR knockdown cells, the overexpression of a HuR mutant that could not be phosphorylated (with changes of serine to alanine at position 88 [S88A], S100A, and T118A) blocked the associatedTRA2β4 interaction andTRA2β4 generation, while the overexpression of a phosphomimetic HuR (with mutations S88D, S100D, and T118D) restored theTRA2β4 -related activities. Our findings revealed the potential role of nuclear HuR in the regulation of alternative splicing programs under oxidative stress.