title: Mitotic arrest by tumor suppressor RASSF1A is regulated via CHK1 phosphorylation.
abstract: The tumor suppressor RAS-association domain family 1 isoform A (RASSF1A) is known to play an important role in cell-cycle regulation. However, the molecular details about RASSF1A protein regulation are unclear. In this report, checkpoint kinase 1 (CHK1) is identified as a novel RASSF1A kinase that phosphorylates RASSF1A in vitro and under cellular conditions. Using tandem mass spectrometry and biochemical analysis, it was determined that CHK1 phosphorylates RASSF1A on Serine 184, which has been shown to be mutated in a subset of human primary nasopharyngeal carcinomas. Furthermore, Serine 184 phosphorylation of RASSF1A was significantly diminished by a CHK1-specific kinase inhibitor. Similarly, a kinase-dead CHK1 mutant was unable to phosphorylate Serine 184 whereas constitutively active-CHK1 enhanced phosphorylation. Molecular substitution of Serine 184 with aspartic acid, mimicking phosphorylation, abolished the ability of RASSF1A to interact with microtubules and induce M-phase arrest. Combined, these data indicate that phosphorylation of RASSF1A by CHK1 is important for mitotic regulation and provide valuable new insight into the regulatory mechanisms of RASSF1A function.
  
  
  CONCLUSIONS
  This study reveals that CHK1-mediated phosphorylation of RASSF1A, at Serine 184, plays an important role in cell-cycle regulation and highlights that mutation of this CHK1 phosphorylation site in nasopharyngeal carcinoma has disease relevance.
authors.affiliation: Department of Pharmacology, State University of New York, Upstate Medical University, 750 East Adams St., Syracuse, NY 13210. huangy@upstate.edu.
authors.forename: Lingyan
authors.initials: L
authors.lastname: Jiang
authors.forename: Rong
authors.initials: R
authors.lastname: Rong
authors.forename: M Saeed
authors.initials: MS
authors.lastname: Sheikh
authors.forename: Ying
authors.initials: Y
authors.lastname: Huang
mesh_terms: D002277:Carcinoma; D045744:Cell Line, Tumor; D000071877:Checkpoint Kinase 1; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D059566:M Phase Cell Cycle Checkpoints / Q000235:genetics*; D061986:MCF-7 Cells; D009154:Mutation / Q000235:genetics; D000077274:Nasopharyngeal Carcinoma; D009303:Nasopharyngeal Neoplasms / Q000235:genetics*; D010766:Phosphorylation / Q000235:genetics; D011494:Protein Kinases / Q000378:metabolism*; D025521:Tumor Suppressor Proteins / Q000235:genetics*
chemical_list: C411762:RASSF1 protein, human; D025521:Tumor Suppressor Proteins; D011494:Protein Kinases; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1