Abstract
Integrins are heterodimeric glycoproteins that bind cells to extracellular matrix. Upon integrin clustering, multimolecular integrin adhesion complexes (IACs) are formed, creating links to the cell cytoskeleton. We have previously observed decreased cell migration and increased sensitivity to microtubule (MT) poisons, paclitaxel and vincristine, in the melanoma cell line MDA-MB-435S upon transfection with integrin αV-specific siRNA, suggesting a link between adhesion and drug sensitivity. To elucidate the underlying mechanism, we determined αV-dependent changes in IAC composition. Using mass spectrometry (MS)-based proteomics, we analyzed the components of isolated IACs of MDA-MB-435S cells and two MDA-MB-435S-derived integrin αV-specific shRNA-expressing cell clones with decreased expression of integrin αV. MS analysis showed that cells preferentially use integrin αVβ5 for the formation of IACs. The differential analysis between MDA-MB-435S cells and clones with decreased expression of integrin αV identified key components of integrin αVβ5 adhesion complexes as talins 1 and 2, α-actinins 1 and 4, filamins A and B, plectin and vinculin. The data also revealed decreased levels of several components of the cortical microtubule stabilization complex, which recruits MTs to adhesion sites (notably liprins α and β, ELKS, LL5β, MACF1, KANK1, and KANK2), following αV knockdown. KANK2 knockdown in MDA-MB-435S cells mimicked the effect of integrin αV knockdown and resulted in increased sensitivity to MT poisons and decreased migration. Taken together, we conclude that KANK2 is a key molecule linking integrin αVβ5 IACs to MTs, and enabling the actin-MT crosstalk that is important for both sensitivity to MT poisons and cell migration.
Highlights
Integrins are cell-surface adhesion molecules that connect cells to other cells and components of the extracellular matrix (ECM)
We have recently shown that knockdown of integrin subunit αV, via transient transfection of integrin αV-specific siRNA, increased sensitivity of MDA-MB-435S cells to PTX and VCR and decreased migration
ΑV integrins, which have been implicated in tumor growth and angiogenesis, as well as in sensitivity to chemo- and radiotherapy, have long been recognized as therapeutic targets
Summary
Integrins are cell-surface adhesion molecules that connect cells to other cells and components of the ECM. Other proteins are recruited to their cytoplasmic tails to form multimolecular IACs which establish the linkage between integrins and the cell cytoskeleton (Winograd-Katz et al, 2014). Integrins together with their associated IAC components have been termed the adhesome (Zaidel-Bar et al, 2007; Kuo et al, 2011; Schiller et al, 2011; Byron et al, 2015; Jones et al, 2015). Talin coordinates the microtubule (MT) cytoskeleton at adhesion sites through the interaction with KN motif and ankyrin repeat domain-containing (KANK) proteins (Bouchet et al, 2016; Sun et al, 2016), which stimulates FA turnover (Stehbens and Wittmann, 2012)
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