TIA-1 or TIAR is required for DT40 cell viability.
TIA-1 and TIAR are a pair of related RNA-binding proteins which have been implicated in apoptosis. We show that chicken DT40 cells with both tia-1 alleles and one tiar allele disrupted (tia-1(-/-)tiar(-/+) cells) are viable. However, their growth and survival in medium containing low serum levels is significantly reduced compared with DT40 cells. The remaining intact tiar allele in tia-1(-/-)tiar(-/+) cells can only be disrupted if TIA-1 expression is first restored to the cells by transfection of a TIA-1 expression vector. We conclude that DT40 cells require either TIA-1 or TIAR for viability. TIA-1 overexpression in tia-1(-/-)tiar(-/+) cells leads to a radical drop in TIAR levels, by inducing efficient splicing of two tiar alternative exons carrying in-frame stop codons. In wild-type DT40 cells, tiar transcripts including these exons can also be detected. These transcripts increase significantly in abundance in cycloheximide-treated cells, suggesting that splicing of the exons exposes mRNAs to nonsense-mediated mRNA decay. TIA-1 or TIAR depletion leads to a marked drop in splicing of the exons. The human tiar gene contains a corresponding pair of TIA-1-inducible alternative exons, and we show that there is very high sequence conservation between chickens and humans of the exon pair and parts of the flanking introns. The TIA-1/TIAR responsiveness of these alternative tiar exons is likely to be of physiological importance for controlling TIAR levels.
Le Guiner
Caroline
C
INSERM U463, Institut de Biologie-CHR, 9 Quai Moncousu, 44093 Nantes Cedex 1, France.
Gesnel
Marie-Claude
MC
Breathnach
Richard
R
D000483:Alleles; D000818:Animals; D017209:Apoptosis; D001483:Base Sequence; D002455:Cell Division; D002460:Cell Line; D002470:Cell Survival; D002645:Chickens; D003062:Codon; D005091:Exons; D006801:Humans; D007438:Introns; D008565:Membrane Proteins / Q000235:genetics / Q000502:physiology*; D008969:Molecular Sequence Data; D039102:Poly(A)-Binding Proteins; D011506:Proteins; D012333:RNA, Messenger / Q000032:analysis; D016601:RNA-Binding Proteins / Q000235:genetics / Q000502:physiology*; D000074019:T-Cell Intracellular Antigen-1
D003062:Codon; D008565:Membrane Proteins; D039102:Poly(A)-Binding Proteins; D011506:Proteins; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D000074019:T-Cell Intracellular Antigen-1; C071558:TIA1 protein, human; C076751:TIAL1 protein, human