Copyright © Johanna Ivaska. All rights reserved.
Keywords: AMPK, integrin activity, matrix remodelling, Ras-association domain, RNAi screens, SHANK, SHARPIN, talin, tensin
Introduction: Integrin heterodimers can exist in a bent/closed conformation with low affinity for extracellular ligands (‘inactive’) or in an extended/open conformation with a high affinity for ligands (‘active’). As a consequence of this conformational switch, integrins are able to transmit signals bidirectionally across the cell membrane. Engagement of extracellular ligand by integrins elicits signalling responses within the cell (‘outside-in’ signalling), whereas binding of intracellular proteins such as talin and kindlins to the β-integrin tail (NPXY motifs) promote the ligand-binding receptor conformation (‘inside-out’ signalling).
Our research: Our RNAi screens have thus far revealed several candidate proteins implicated in the regulation of integrin activity and importantly have led to the identification of SHARPIN, SHANKs and AMPK as negative regulators of integrin function (see Publications: Rantala et al., 2011; Lilja et al., 2017; Georgiadou et al., 2017 and below for a summary of these works).
SHARPIN deficiency causes aberrant collagen fibre organization in the mouse mammary gland stroma. Second harmonic generation (SHG) and unfiltered multiphoton (MP) imaging of collagen fibres surrounding terminal end buds in 6-7 weeks old wild-type (wt) and Sharpin-/- mouse mammary glands are shown. Adapted from Peuhu et al. 2017)
AMPK (5' AMP-activated protein kinase) is an important energy sensor and has been extensively studied as a central regulator of cellular metabolism; however, in recent years it has become apparent that AMPK directly phosphorylates non-metabolic proteins and regulates other cellular functions, such as transcription, cell polarity, mitosis, migration and adhesion. We found that inhibition of AMPK triggers a significant increase in integrin activity in fibroblasts, accompanied by enhanced formation of fibrillar adhesions (centrally located α5β1 integrin-ECM contacts rich in tensin) and fibronectin fibrillogenesis. In line with these data, we discovered an upregulation in tensin expression, the major component of fibrillar adhesions, following AMPK silencing or knockdown. As tensins bind to overlapping talin-binding sites within the β-integrin tail, we propose that tensins maintain integrins in an active state following talin-dependent activation. Importantly the role of AMPK in this process, as a regulator of tensin expression, provides a link between energy sensing and integrin signalling that may have wide implications in fibrosis and cancer (see Publications; Georgiadou et al., 2017).
SHARPIN is a widely expressed multifunctional protein implicated in cancer, inflammation and linear ubiquitination. We discovered that siRNA-mediated silencing of SHARPIN expression results in increased integrin activity, implicating SHARPIN as a negative regulator of integrin activation and signalling. We identified a conserved SHARPIN binding site within the α-integrin cytoplasmic domain and found that SHARPIN-integrin interaction inhibits the recruitment of integrin activators to the β-tail (see Publications; Rantala et al., 2011).
More recently, we demonstrated a role for SHARPIN in epithelial homeostasis in the developing mouse mammary gland. We observed increased intergin activity and altered stromal matrix deposition in vivo in the absence of SHARPIN, leading to diminished mammary ductal invasion and growth (see Publications; Peuhu et al., 2017).