TI - SOCS3 inhibiting migration of A549 cells correlates with PYK2 signaling in vitro . AB - Background Suppressor of cytokine signaling 3 ( SOCS3 ) is considered to inhibit cytokine responses and play a negative role in migration of various cells . Proline-rich tyrosine kinase 2 ( PYK2 ) is a non receptor kinase and has been found crucial to cell motility . However , little is known about whether SOCS3 could regulate PYK2 pro-migratory function in lung cancer . Methods The methylation status of SOCS3 was investigated in HBE and A549 cell lines by methylation-specific PCR . A549 cells were either treated with a demethylation agent 5-aza-2'-deoxycytidine or transfected with three SOCS3 mutants with various functional domains deleted . Besides , cells were pretreated with a proteasome inhibitor beta-lactacystin where indicated . The effects of SOCS3 up-regulation on PYK2 expression , PYK2 and ERK1/2 PHOSphorylations were assessed by western blot using indicated antibodies . RT-PCR was used to estimate PYK2 mRNA levels . Transwell experiments were performed to evaluate cell migration . Results SOCS3 expression was found impaired in A549 cells and higher PYK2 activity was correlated with enhanced cell migration . We identified that SOCS3 was aberrantly methylated in the exon 2 , and 5-aza-2'-deoxycytidine restored SOCS3 expression . Reactivation of SOCS3 attenuated PYK2 expression and PHOSphorylation , cell migration was inhibited as well . Transfection studies indicated that exogenous SOCS3 interacted with PYK2 , and both the Src homology 2 ( SH2 ) and the kinase inhibitory region ( KIR ) domains of SOCS3 contributed to PYK2 binding . Furthermore , SOCS3 was found to inhibit PYK2-associated ERK1/2 activity in A549 cells . SOCS3 possibly promoted degradation of PYK2 in a SOCS-box -dependent manner and interfered with PYK2-related signaling events , such as cell migration . Conclusion These data indicate that SOCS3 negatively regulates cell motility and decreased SOCS3 induced by methylation may confer a migration advantage to A549 cells . These results also suggest a negative role of SOCS3 in PYK2 signaling , and a previously unidentified regulatory mechanism for PYK2 function .