title: Structure/function analysis of tristetraprolin (TTP): p38 stress-activated protein kinase and lipopolysaccharide stimulation do not alter TTP function.
abstract: Tristetraprolin (TTP) is the only trans-acting factor shown to be capable of regulating AU-rich element-dependent mRNA turnover at the level of the intact animal; however, the mechanism by which TTP mediated RNA instability is unknown. Using an established model system, we performed structure/function analysis with TTP as well as examined the current hypothesis that TTP function is regulated by p38-MAPKAP kinase 2 (MK2) activation. Deletion of either the N- or C-terminal domains inhibited TTP function. Extensive mutagenesis, up to 16%, of serines and threonines, some of which were predicted to mediate proteasomal targeting, did not alter human TTP function. Mutation of the conserved MK2 phosphorylation sites enhanced human TTP function in both resting and p38-stress-activated protein kinase-MK2-activated cells. However, p38-stress-activated protein kinase-MK2 activation did not alter the activity of either wild-type or mutant TTP. TTP localized to the stress granules, with arsenite treatment reducing this localization. In contrast, arsenite treatment enhanced stress granule localization of the MK2 mutant, consistent with the involvement of additional pathways regulating this event. Finally, we determined that, in response to LPS stimulation, human TTP moves onto the polysomes, and this movement occurs in the absence of 14-3-3. Taken together, these data indicate that, although p38 activation alters TTP entry into the stress granule, it does not alter TTP function. Moreover, the interaction of TTP with 14-3-3, which may limit entry into the stress granule, is not involved in the downstream message stabilization events.
authors.affiliation: Department of Medicine, Dartmouth College, Lebanon, NH 03756, USA.
authors.forename: William F C
authors.initials: WF
authors.lastname: Rigby
authors.forename: Kristen
authors.initials: K
authors.lastname: Roy
authors.forename: Jane
authors.initials: J
authors.lastname: Collins
authors.forename: Sam
authors.initials: S
authors.lastname: Rigby
authors.forename: John E
authors.initials: JE
authors.lastname: Connolly
authors.forename: Donald B
authors.initials: DB
authors.lastname: Bloch
authors.forename: Seth A
authors.initials: SA
authors.lastname: Brooks
mesh_terms: D048948:14-3-3 Proteins / Q000737:chemistry / Q000378:metabolism / Q000502:physiology; D000595:Amino Acid Sequence; D000818:Animals; D018053:Arsenites / Q000494:pharmacology; D002460:Cell Line; D045744:Cell Line, Tumor; D003594:Cytoplasmic Granules / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D006801:Humans; D017874:Immediate-Early Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D047908:Intracellular Signaling Peptides and Proteins; D008070:Lipopolysaccharides / Q000737:chemistry*; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D011132:Polyribosomes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011237:Predictive Value of Tests; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D017434:Protein Structure, Tertiary / Q000235:genetics; D020871:RNA Stability / Q000502:physiology; D017384:Sequence Deletion; D012694:Serine / Q000235:genetics; D013312:Stress, Physiological / Q000139:chemically induced / Q000235:genetics / Q000378:metabolism; D013329:Structure-Activity Relationship; D013912:Threonine / Q000235:genetics; D014162:Transfection; D051816:Tristetraprolin; D048051:p38 Mitogen-Activated Protein Kinases / Q000737:chemistry* / Q000378:metabolism; D000099190:MAP-Kinase-Activated Kinase 2
chemical_list: D048948:14-3-3 Proteins; D018053:Arsenites; D004268:DNA-Binding Proteins; D017874:Immediate-Early Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008070:Lipopolysaccharides; D017346:Protein Serine-Threonine Kinases; D012694:Serine; D013912:Threonine; D051816:Tristetraprolin; D048051:p38 Mitogen-Activated Protein Kinases; D000099190:MAP-Kinase-Activated Kinase 2; C497552:ZFP36 protein, human; C497553:Zfp36 protein, mouse; C015001:arsenite