Pro‐migratory and TGF‐β‐activating functions of αvβ6 integrin in pancreatic cancer are differentially regulated via an Eps8‐dependent GTPase switch

Abstract

The integrin αvβ6 is up‐regulated in numerous carcinomas, where expression commonly correlates with poor prognosis. αvβ6 promotes tumour invasion, partly through regulation of proteases and cell migration, and is also the principal mechanism by which epithelial cells activate TGF‐β1; this latter function complicates therapeutic targeting of αvβ6, since TGF‐β1 has both tumour‐promoting and ‐suppressive effects. It is unclear how these different αvβ6 functions are linked; both require actin cytoskeletal reorganization, and it is suggested that tractive forces generated during cell migration activate TGF‐β1 by exerting mechanical tension on the ECM‐bound latent complex. We examined the functional relationship between cell invasion and TGF‐β1 activation in pancreatic ductal adenocarcinoma (PDAC) cells, and confirmed that both processes are αvβ6‐dependent. Surprisingly, we found that cellular functions could be biased towards either motility or TGF‐β1 activation depending on the presence or absence of epidermal growth factor receptor pathway substrate 8 (Eps8), a regulator of actin remodelling, endocytosis, and GTPase activation. Similar to αvβ6, we found that Eps8 was up‐regulated in >70% of PDACs. In complex with Abi1/Sos1, Eps8 regulated αvβ6‐dependent cell migration through activation of Rac1. Down‐regulation of Eps8, Sos1 or Rac1 suppressed cell movement, while simultaneously increasing αvβ6‐dependent TGF‐β1 activation. This latter effect was modulated through increased cell tension, regulated by Rho activation. Thus, the Eps8/Abi1/Sos1 tricomplex acts as a key molecular switch altering the balance between Rac1 and Rho activation; its presence or absence in PDAC cells modulates αvβ6‐dependent functions, resulting in a pro‐migratory (Rac1‐dependent) or a pro‐TGF‐β1 activation (Rho‐dependent) functional phenotype, respectively. © 2017 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Targeting the complexity of Src signalling in the tumour microenvironment of pancreatic cancer: from mechanism to therapy

“…Furthermore, integrins also play a role in angiogenesis, by providing a docking site for several cell types, including endothelial cells, endothelial stem cells and inflammatory cells, at the site of angiogenesis [65]. Upregulation of amb6-integrins occurs in a variety of tumours, including PDAC, where it has been shown to activate TGF-b, stimulating tumour cell epithelial-to-mesenchymal transition (EMT) and stromal myofibroblast differentiation [66], which has in turn been shown to either promote [67] or restrict tumour growth and progression [68]. The association between amb6-integrins and increased migration, invasion and cell survival is partly due to the regulation of proteases (MMPs), and urokinasetype plasminogen activator (uPA) [63, 66,[69][70][71].…”

“…Upregulation of amb6-integrins occurs in a variety of tumours, including PDAC, where it has been shown to activate TGF-b, stimulating tumour cell epithelial-to-mesenchymal transition (EMT) and stromal myofibroblast differentiation [66], which has in turn been shown to either promote [67] or restrict tumour growth and progression [68]. The association between amb6-integrins and increased migration, invasion and cell survival is partly due to the regulation of proteases (MMPs), and urokinasetype plasminogen activator (uPA) [63, 66,[69][70][71]. In PDAC specifically, overexpression of integrin amb3/ amb6 has been previously shown to associate with poor survival of patients as well as lymph node metastasis [59,72], and recent findings indicate that the stromal localisation and levels of active a5b1-integrin and FAK can identify two readily distinguishable desmoplastic phenotypes in pancreatic cancer.…”

Integrin αVβ6-EGFR crosstalk regulates bidirectional force transmission and controls breast cancer invasion

“…Relative to other fibronectin-binding integrins, αVβ6 exhibits distinct biophysical properties that promote force-generation, rigidity sensing and increase matrix stiffening 15 . Moreover, TGFβ is mechanically activated by αVβ6; requiring transmission of intracellular actomyosin-dependent forces to be applied on latent TGFβ via αVβ6 [32][33][34] .Biochemical integrin internalisation assays confirmed that EGF stimulation triggered endocytosis of αVβ6 ( Figure 6A). Therefore, we investigated whether αVβ6-EGFR crosstalk could regulate the ability of αVβ6 to transmit cytoskeletal mechanical forces to the extracellular environment.…”

“…Therefore, we investigated whether αVβ6-EGFR crosstalk could regulate the ability of αVβ6 to transmit cytoskeletal mechanical forces to the extracellular environment. We performed traction force microscopy, using polyacrylamide hydrogels coated with the αVβ6-selective ligand LAP, to measure αVβ6-dependent mechanical force application 32 . These assays revealed that stimulation with EGF induced a marked suppression in the ability of MDA-MB-468 cells to apply αVβ6-dependent mechanical force on LAP ( Figure 6B).…”

“…Polyacrylamide hydrogels embedded with FluoSpheres® carboxylate-modified 0.2 μm fluorescent (505/515) microspheres were synthesised as described previously 32 . Hydrogels were treated with sulfo-SANPAH under UV light (365nm), then coated with LAP (0.5μg/ml) overnight at 4°C.…”