Phosphorylation of tristetraprolin, a potential zinc finger transcription factor, by mitogen stimulation in intact cells and by mitogen-activated protein kinase in vitro.
Tristetraprolin (TTP) is a potential transcription factor that contains three PPPPG repeats and two putative CCCH zinc fingers. TTP is encoded by the early response gene Zfp-36, which is highly expressed in response to growth factors and in several hematopoietic cell lines. In the present studies, we investigated the possibility that TTP is phosphorylated in intact cells. In NIH/3T3 cells that were made to overexpress TTP constitutively, we found that the protein was phosphorylated on serine residues, and that this phosphorylation was rapidly (within 10 min) stimulated by several mitogens. In cell-free assays, recombinant mouse TTP was a substrate for the mitogen-activated protein (MAP) kinase. By a combination of protease digestion experiments and site-directed mutagenesis strategies, we found that serine 220 was phosphorylated by p42 MAP kinase in vitro. Expression of mutant TTP in fibroblasts confirmed that serine 220 was one of the major, mitogen-stimulated phosphorylation sites on the protein in intact cells. These results suggest that TTP may be phosphorylated by MAP kinases in vivo and that this phosphorylation may regulate its function.
Taylor
G A
GA
Howard Hughes Medical Institute Laboratories, Duke University Medical Center, Durham, North Carolina 27710, USA.
Thompson
M J
MJ
Lai
W S
WS
Blackshear
P J
PJ
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D017931:DNA Primers; D004268:DNA-Binding Proteins; D006801:Humans; D017874:Immediate-Early Proteins; D051379:Mice; D019950:Mitogen-Activated Protein Kinase 1; D008934:Mitogens / Q000494:pharmacology*; D008969:Molecular Sequence Data; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D012694:Serine / Q000378:metabolism; D013379:Substrate Specificity; D014157:Transcription Factors / Q000378:metabolism*; D051816:Tristetraprolin; D016335:Zinc Fingers
D017931:DNA Primers; D004268:DNA-Binding Proteins; D017874:Immediate-Early Proteins; D008934:Mitogens; D011506:Proteins; D014157:Transcription Factors; D051816:Tristetraprolin; C497552:ZFP36 protein, human; C497553:Zfp36 protein, mouse; D012694:Serine; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D019950:Mitogen-Activated Protein Kinase 1