TY - JOUR
T1 - Nuclear protein TIA-1 regulates COL2A1 alternative splicing and interacts with precursor mRNA and genomic DNA
AU - McAlinden, Audrey
AU - Liang, Li
AU - Mukudai, Yoshiki
AU - Imamura, Toshihiro
AU - Sandell, Linda J.
PY - 2007/8/17
Y1 - 2007/8/17
N2 - The RNA-binding protein TIA-1 (T-cell-restricted intracellular antigen-1) functions in regulating post-transcriptional mechanisms, including precursor mRNA (pre-mRNA) alternative splicing and mRNA translation. Utilizing a mini-gene consisting of part of the type II procollagen gene (COL2A1), we show that TIA-1 interacts with a conserved AU-rich cis element in COL2A1 intron 2 and modulates alternative splicing of exon 2. This unique, highly conserved cis element containing stemloop secondary structure was previously identified in our laboratory as an essential motif that controls the developmentally regulated exon 2 splicing switch during chondrogenesis (McAlinden, A., Havlioglu, N., Liang, L., Davies, S. R., and Sandell, L. J. (2005) J. Biol. Chem. 280, 32700-32711). In vivo binding of endogenous TIA-1 to the AU-rich cis element in COL2A1 pre-mRNA was confirmed by the ribonucleoprotein immunoprecipitation assay. Importantly, we also show that TIA-1 interacts with the equivalent DNA sequence with a preference for single-stranded rather than double-stranded DNA. Chromatin immunoprecipitation assays (including an additional RNase step) confirmed this interaction in vivo. Competition assays showed that TIA-1 apparently binds with higher affinity to DNA than to RNA. Finally, we show that this strong DNA-TIA-1 interaction can be disrupted by an RNA polymerase during active transcription. This suggests a potentially novel, dual role for TIA-1 in shuttling between DNA and RNA ligands to coregulate COL2A1 expression at the level of transcription and pre-mRNA alternative splicing.
AB - The RNA-binding protein TIA-1 (T-cell-restricted intracellular antigen-1) functions in regulating post-transcriptional mechanisms, including precursor mRNA (pre-mRNA) alternative splicing and mRNA translation. Utilizing a mini-gene consisting of part of the type II procollagen gene (COL2A1), we show that TIA-1 interacts with a conserved AU-rich cis element in COL2A1 intron 2 and modulates alternative splicing of exon 2. This unique, highly conserved cis element containing stemloop secondary structure was previously identified in our laboratory as an essential motif that controls the developmentally regulated exon 2 splicing switch during chondrogenesis (McAlinden, A., Havlioglu, N., Liang, L., Davies, S. R., and Sandell, L. J. (2005) J. Biol. Chem. 280, 32700-32711). In vivo binding of endogenous TIA-1 to the AU-rich cis element in COL2A1 pre-mRNA was confirmed by the ribonucleoprotein immunoprecipitation assay. Importantly, we also show that TIA-1 interacts with the equivalent DNA sequence with a preference for single-stranded rather than double-stranded DNA. Chromatin immunoprecipitation assays (including an additional RNase step) confirmed this interaction in vivo. Competition assays showed that TIA-1 apparently binds with higher affinity to DNA than to RNA. Finally, we show that this strong DNA-TIA-1 interaction can be disrupted by an RNA polymerase during active transcription. This suggests a potentially novel, dual role for TIA-1 in shuttling between DNA and RNA ligands to coregulate COL2A1 expression at the level of transcription and pre-mRNA alternative splicing.
UR - http://www.scopus.com/inward/record.url?scp=34548129306&partnerID=8YFLogxK
U2 - 10.1074/jbc.M702717200
DO - 10.1074/jbc.M702717200
M3 - Article
C2 - 17580305
AN - SCOPUS:34548129306
SN - 0021-9258
VL - 282
SP - 24444
EP - 24454
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 33
ER -