SUMOylation-dependent Functions And Mechanisms Of SNIP1 In The TGF? Signaling Pathway

Transforming growth factor ?(TGF?)is a multifunctional factor and plays an important role in promoting tumor metastasis.In canonical TGF? signaling,the ligand binds to and activates the receptors,which phosphorylate Smad2/3.Phosphorylated Smad2/3 forms a complex with Smad4 and the Smad complex undergoes nuclear translocation.Inside the nucleus,the Smad complex binds to DNA,recruits transcription co-activators p300/CBP,and induces or represses the expression of target genes.Smad Nuclear Interacting Protein 1(SNIP 1)was found as a Smad-interacting protein and could inhibit TGF? signaling by disrupting the interaction between Smad4 and p300.Although SNIP1 is crucial in the regulation of TGF? signaling pathway,its regulation via post-translational modifications has not been reported.SUMOylation is a type of posttranslational modification by small-ubiquitin-like proteins called SUMO.The function of SUMOylation varies as it has been linked to a variety of cellular activities,including genomic integrity,subcellular localization and transcriptional regulation.Preliminary studies in our lab showed that SNIP1 could be SUMOylated at three lysine residues,Lys5,Lys30 and Lys108.Thus,we sought to determine whether SUMO modification affected the transcriptional activity of SNIP1 in TGF? signaling pathway.We found that SUMO modification disabled the inhibitory effect of SNIP1 in the TGF? pathway.SBE-luc and 3TP-luc reporter gene studies showed that the SUMO modification suppressed the inhibitory activity of SNIP1 in the TGFP signaling pathway.The same result was obtained when the mRNA and protein levels of TGFP target genes such as PAI-1 and MMP-2 were analyzed.Mechanistically,we revealed that while SNIP1 decreased stability of the Smad complex and disrupted the interactions between Smad and p300,SUMO-modified form of SNIP1 lost these activities.Functionally,SUMO modification disabled the inhibitory effect of SNIP1 on TGF?-induced migratory and invasive responses in A549 cells.We also found that SNIP1 and its SUMOylation affected the activation of RhoA.Opposing to the inhibitory effect of SNIP1,SUMOylated SNIP1 increased the level of GTP-bound RhoA and the morphology of F-actin.Thus,our novel studies demonstrated that SUMOylation suppressed the activity of SNIP1 in the TGFP signaling pathway and enabled suitable conditions for TGF?-induced tumor cell migration and invasion.