Regulation of glioma cell migration by serine-phosphorylated P311.
P311, an 8-kDa polypeptide, was previously shown to be highly expressed in invasive glioma cells. Here, we report the functional characteristics of P311 with regard to influencing glioma cell migration. P311 is constitutively serine-phosphorylated; decreased phosphorylation is observed in migration-activated glioma cells. The primary amino acid sequence of P311 indicates a putative serine phosphorylation site (S59) near the PEST domain. Site-directed mutagenesis of S59A retarded P311 degradation and induced glioma cell motility. In contrast, S59D mutation resulted in the rapid degradation of P311 and reduced glioma cell migration. Coimmunoprecipitation coupled with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis identified Filamin A as a binding partner of P311, and immunofluorescence studies showed that both proteins colocalized at the cell periphery. Moreover, P311-induced cell migration was abrogated by inhibition of beta1 integrin function using TACbeta1A, a dominant-negative inhibitor of beta1 integrin signaling, suggesting that P311 acts downstream of beta1 signaling. Finally, overexpression of P311 or P311 S59A mutant protein activates Rac1 GTPase; small interfering RNA-mediated depletion of Rac1 suppresses P311-induced motility. Collectively, these results suggest a role for levels of P311 in regulating glioma motility and invasion through the reorganization of actin cytoskeleton at the cell periphery.
McDonough
Wendy S
WS
The Translational Genomics Research Institute, Phoenix, AZ 85004, USA.
Tran
Nhan L
NL
Berens
Michael E
ME
D000595:Amino Acid Sequence; D045744:Cell Line, Tumor; D002465:Cell Movement; D003285:Contractile Proteins / Q000378:metabolism; D064448:Filamins; D005910:Glioma / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D008840:Microfilament Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D009361:Neoplasm Invasiveness; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000378:metabolism*; D015513:Oncogene Proteins / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000235:genetics; D017434:Protein Structure, Tertiary; D034741:RNA, Small Interfering / Q000378:metabolism; D012694:Serine / Q000378:metabolism*; D020830:rac1 GTP-Binding Protein / Q000378:metabolism
D003285:Contractile Proteins; D064448:Filamins; D008840:Microfilament Proteins; C428027:NREP protein, human; D009419:Nerve Tissue Proteins; D015513:Oncogene Proteins; D034741:RNA, Small Interfering; D012694:Serine; D011493:Protein Kinase C; D020830:rac1 GTP-Binding Protein