Abstract
Focal adhesion assembly and disassembly are essential for cell migration and cancer invasion, but the detailed molecular mechanisms regulating these processes remain to be elucidated. Phosphatidylinositol phosphate kinase type Iγ (PIPKIγ) binds talin and is required for focal adhesion formation in EGF-stimulated cells, but its role in regulating focal adhesion dynamics and cancer invasion is poorly understood. We show here that overexpression of PIPKIγ promoted focal adhesion formation, whereas cells expressing either PIPKIγK188,200R or PIPKIγD316K, two kinase-dead mutants, had much fewer focal adhesions than those expressing WT PIPKIγ in CHO-K1 cells and HCT116 colon cancer cells. Furthermore, overexpression of PIPKIγ, but not PIPKIγK188,200R, resulted in an increase in both focal adhesion assembly and disassembly rates. Depletion of PIPKIγ by using shRNA strongly inhibited formation of focal adhesions in HCT116 cells. Overexpression of PIPKIγK188,200R or depletion of PIPKIγ reduced the strength of HCT116 cell adhesion to fibronection and inhibited the invasive capacities of HCT116 cells. PIPKIγ depletion reduced PIP2 levels to ∼40% of control and PIP3 to undetectable levels, and inhibited vinculin localizing to focal adhesions. Taken together, PIPKIγ positively regulates focal adhesion dynamics and cancer invasion, most probably through PIP2-mediated vinculin activation.
Highlights
Focal adhesions (FAs, called cell-matrix adhesions) are specific types of large macromolecular assemblies at the ventral surface of cells, functioning as both mechanical machineries and regulatory signaling hubs [1,2]
Phosphatidylinositol phosphate kinase type Ic (PIPKIc) promotes focal adhesion formation To examine a possible role for PIPKIc in focal adhesion formation, CHO-K1 cells were transfected with EGFP-PIPKIc or EGFP vector, respectively
EGFP-tagged mutant PIPKIcK188,200R and WT PIPKIc were stably expressed in CHO-K1 cells, and their effects on focal adhesion formation were examined after paxillin staining
Summary
Focal adhesions (FAs, called cell-matrix adhesions) are specific types of large macromolecular assemblies at the ventral surface of cells, functioning as both mechanical machineries and regulatory signaling hubs [1,2]. Temporal and spatial regulation of focal adhesion assembly/disassembly is required for cell migration [3]. Focal adhesion assembly and disassembly are implicated in cancer invasion, a prerequisite for metastasis. DRR (DownRegulated in Renal cell carcinoma) associates with actin and microtubules and stimulates glioma invasion by promoting focal adhesion disassembly [6]. The actin cross-linking protein filamin A suppresses focal adhesion disassembly and breast cancer cell invasion [7]. Rho/Rock signaling promotes tumor cell migration and invasion by regulating focal adhesion dynamics through caveolin-1 phosphorylation [8]. FAK promotes focal adhesion disassembly and cancer invasion [9,10,11]. Focal adhesion dynamics and signaling pathways that regulate this process could be an attractive target for cancer therapy
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