The CD9/CD81 tetraspanin complex and tetraspanin CD151 regulate α3β1 integrin-dependent tumor cell behaviors by overlapping but distinct mechanisms.

Elisabeth Gustafson-Wagner,Christopher S Stipp,Maddy Parsons

doi:10.1371/journal.pone.0061834

Elisabeth Gustafson-Wagner, Christopher S Stipp + Show 1 more

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https://doi.org/10.1371/journal.pone.0061834

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Journal: PloS one	Publication Date: Apr 17, 2013
Citations: 45	License type: CC BY 4.0

Affiliation: University of Iowa

Abstract

Integrin α3β1 potently promotes cell motility on its ligands, laminin-332 and laminin-511, and this may help to explain why α3β1 has repeatedly been linked to breast carcinoma progression and metastasis. The pro-migratory functions of α3β1 depend strongly on lateral interactions with cell surface tetraspanin proteins. Tetraspanin CD151 interacts directly with the α3 integrin subunit and links α3β1 integrin to other tetraspanins, including CD9 and CD81. Loss of CD151 disrupts α3β1 association with other tetraspanins and impairs α3β1-dependent motility. However, the extent to which tetraspanins other than CD151 are required for specific α3β1 functions is unclear. To begin to clarify which aspects of α3β1 function require which tetraspanins, we created breast carcinoma cells depleted of both CD9 and CD81 by RNA interference. Silencing both of these closely related tetraspanins was required to uncover their contributions to α3β1 function. We then directly compared our CD9/CD81-silenced cells to CD151-silenced cells. Both CD9/CD81-silenced cells and CD151-silenced cells showed delayed α3β1-dependent cell spreading on laminin-332. Surprisingly, however, once fully spread, CD9/CD81-silenced cells, but not CD151-silenced cells, displayed impaired α3β1-dependent directed motility and altered front-rear cell morphology. Also unexpectedly, the CD9/CD81 complex, but not CD151, was required to promote α3β1 association with PKCα in breast carcinoma cells, and a PKC inhibitor mimicked aspects of the CD9/CD81-silenced cell motility defect. Our data reveal overlapping, but surprisingly distinct contributions of specific tetraspanins to α3β1 integrin function. Importantly, some of CD9/CD81's α3β1 regulatory functions may not require CD9/CD81 to be physically linked to α3β1 by CD151.

Highlights

Integrins, the major family of cellular receptors for extracellular matrix proteins, comprise 18 a and 8 b subunits, which assemble into 24 known ab heterodimers with different ligand binding specificities [1]
The first major finding of our study is that the CD9/CD81 complex is required for certain aspects of a3b1 integrin function in tumor cells
One recent study showed that a novel, PKCa inhibitory peptide can strongly inhibit spontaneous metastasis in the 4T1 murine mammary carcinoma cell model [73]

Summary

Introduction

The major family of cellular receptors for extracellular matrix proteins, comprise 18 a and 8 b subunits, which assemble into 24 known ab heterodimers with different ligand binding specificities [1]. The laminin-binding integrins, a3b1, a6b1, a6b4, and a7b1, constitute a distinct subfamily These integrins play essential roles in the morphogenesis and maintenance of skin, kidney, and lung epithelia (a3 and a6 integrins) and muscle (a7 integrin) by binding to laminin isoforms in the basement membranes underlying these tissues [2,3,4]. In addition to ligand preference, the laminin-binding integrins share other biochemical similarities, including palmitoylation of the a3, a6, and b4 integrin subunit cytoplasmic tails [5], and physical interactions with tetraspanin proteins in the cell membrane [6,7,8]. Localization of the laminin-binding integrins to TEMs may provide access to a distinct array of cytoplasmic signaling proteins, including PI 4-kinase, ERM proteins, and classical PKC isoforms [6,9,10,11,12]

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