Control of cellular responses to mechanical cues through YAP/TAZ regulation

Abstract

To perceive their three-dimensional environment, cells and tissues must be able to sense and interpret various physical forces like shear, tensile, and compression stress. These forces can be generated both internally and externally in response to physical properties, like substrate stiffness, cell contractility, and forces generated by adjacent cells. Mechanical cues have important roles in cell fate decisions regarding proliferation, survival, and differentiation as well as the processes of tissue regeneration and wound repair. Aberrant remodeling of the extracellular space and/or defects in properly responding to mechanical cues likely contributes to various disease states, such as fibrosis, muscle diseases, and cancer. Mechanotransduction involves the sensing and translation of mechanical forces into biochemical signals, like activation of specific genes and signaling cascades that enable cells to adapt to their physical environment. The signaling pathways involved in mechanical signaling are highly complex, but numerous studies have highlighted a central role for the Hippo pathway and other signaling networks in regulating the YAP and TAZ (YAP/TAZ) proteins to mediate the effects of mechanical stimuli on cellular behavior. How mechanical cues control YAP/TAZ has been poorly understood. However, rapid progress in the last few years is beginning to reveal a surprisingly diverse set of pathways for controlling YAP/TAZ. In this review, we will focus on how mechanical perturbations are sensed through changes in the actin cytoskeleton and mechanosensors at focal adhesions, adherens junctions, and the nuclear envelope to regulate YAP/TAZ.

Keywords: Hippo pathway; LATS (Warts, Wts); TAZ; actin; adherens junction; cell signaling; focal adhesions; mechanotransduction; yes-associated protein (YAP).

Figure 1.

Overview of YAP regulation by mechanical stimuli. Changes in the mechanical environment of the cell can control YAP activity through multiple Hippo-dependent and -independent mechanisms. The Hippo pathway acts to inhibit YAP nuclear localization. Several upstream Hippo pathway kinases like MST1/2, MAP4K-family, and TAO phosphorylate LATS1/2 in the presence of its activator, MOB1. The active, phosphorylated form of LATS1/2 phosphorylates YAP, retaining it in the cytoplasm, thereby resulting in YAP inhibition. Tension sensed at focal adhesions, adherens junctions, and the nuclear envelope, as well as changes in F-actin levels in the cytoplasm and the nucleus, control YAP activity through both Hippo-dependent and -independent pathways. F-actin levels can be influenced by the mechanical environment through tension sensing at the adherens junctions, focal adhesions, and nucleus. In turn, F-actin can affect tension experienced by the cell at each of these structures. In sum, increases in F-actin and tension inhibit Hippo signaling and promote YAP activity.

Figure 2.

Changes in F-actin levels in response to mechanical stimuli regulate YAP activity. F-actin modulates YAP activity through both Hippo-dependent and -independent mechanisms. A, in general, disassembly or loss of F-actin filaments results in LATS1/2 activation and a concomitant inhibition of YAP activity by various upstream pathways. PKA phosphorylates and activates LATS1/2, thereby promoting YAP phosphorylation and inactivation. Other upstream kinases, like MAP4K-family and TAO, activate LATS1/2; however, whether they are directly regulated by F-actin levels is not known. Additionally, AMOT can directly sense F-actin levels to influence YAP activity. When actin levels are low, AMOT is free to bind SAV-MST1/2, LATS1/2, and YAP to promote LATS1/2 activation and inhibition of YAP by sequestering it in the cytoplasm. AMOT also binds NF2 to stimulate its ability to activate LATS1/2. AMOT can also bind directly to YAP, independent of Hippo signaling, to retain it in the cytoplasm. When nuclear F-actin levels are low, the Arid1A-SWI/SNF chromatin remodeling complex inhibits any nuclear YAP by blocking its interaction with the transcription factor TEAD. B, conversely, F-actin assembly mediated by G-protein–coupled receptor signaling, shear stress, substrate stiffness, or other factors enables actin binding to AMOT. Thus, AMOT is no longer free to bind YAP, LATS1/2, and SAV-MST1/2, which removes inhibitory signals to allow nuclear translocation of YAP, rendering it active. Nuclear F-actin assembly triggered by mechanical forces also promotes YAP activity. Arid1A-SWI/SNF associates with nuclear F-actin, allowing YAP to bind TEAD.

Figure 3.

Regulation of YAP activity by adherens junctions signaling. A, under conditions of high mechanical tension (low density), cadherin-mediated junctions function to trigger YAP activity. The α-catenin protein binds both β-catenin and actin stress fibers and is thus subject to pulling forces from neighboring cells and tension generated by the actin-myosin cytoskeleton. Tension causes α-catenin to undergo a conformational change, which increases its binding to LIMD1 and vinculin. LIMD1 promotes LATS1/2 recruitment to junctions and inhibition. Vinculin bound to α-catenin binds to F-actin and recruits TRIP6. TRIP6 competes with LATS1/2 activator MOB1 and binds LATS1/2 at adherens junctions, thereby inhibiting it. Together, LIMD1 and TRIP6 inhibition of LATS1/2 allow YAP to translocate to the nucleus. B, at high cell density, reduced tension possibly caused by loss of stress fibers causes α-catenin to revert to a closed conformation, thereby impeding the junctional recruitment of LIMD1, vinculin, TRIP6, and LATS1/2, rendering LATS1/2 free to bind MOB1 and become active (phosphorylated) and inhibit YAP. Additionally, high cell densities promote circumferential actin belt contraction in some cell types, which releases Merlin bound to E-cadherin junctions. Merlin then enters the nucleus to drive the nuclear export of YAP. Once in the cytoplasm, YAP is phosphorylated by kinases, including LATS1/2. α-Catenin inhibits YAP activity by binding to phosphorylated YAP, in a complex with 14-3-3, and protects it from dephosphorylation by PP2A phosphatase.

Figure 4.

YAP regulation by focal adhesion signaling. Integrin engagement with extracellular matrix on stiff substrates triggers activation of focal adhesion signaling driven by FAK and SRC tyrosine kinases. FAK and SRC kinases modulate Hippo pathway and YAP activity in various ways. Both kinases function to activate YAP to promote cell proliferation on stiff substrates. FAK and SRC can directly phosphorylate YAP to promote its activity. In addition, FAK and SRK directly phosphorylate MOB1 and LATS1/2, respectively, to inhibit their activity. They also inhibit LATS1/2 less directly through several signaling pathways. Integrin signaling through a FAK-SRC-PI3K-PDK1 pathway inhibits LATS1/2 activity, thereby enhancing YAP nuclear enrichment. FAK-dependent activation of a RAC-PAK signaling cascade also ensures inhibition of LATS1/2 activity by phosphorylating and inhibiting NF2. Another mechanism by which focal adhesions regulate Hippo signaling is by modulating PIP₂ levels. On stiff substrates, active FAK induces PLC, which reduces PIP₂. Reduction of PIP₂ shuts down the signaling cascade that eventually leads to the activation of RAP2. RAP2 acts both to inhibit Rho, a known inhibitor of LATS1/2, and to activate LATS1/2 by activating MAP4K kinases. Thus, increasing substrate stiffness impedes RAP2 activation by inhibiting upstream PIP₂, thereby promoting nuclear translocation and activation of YAP.

Figure 5.

Model showing force dependent nuclear localization of YAP. A, cells grown on soft substrates are round, and the nucleus is poorly coupled to the substrate. As a result, the nucleus maintains its round shape, and YAP is retained in the cytoplasm because of the permeability barrier of the nuclear pores. B, increasing substrate stiffness causes enhanced coupling of the nucleus to the cell substrate due to increased connections between focal adhesions and LINC complexes in the nuclear membrane via actin stress fibers. This results in increased contractile forces on the nucleus, causing it to flatten as the cell assumes a more spread-out morphology. Nuclear flattening causes the nuclear pores to be more permeable to YAP, resulting in increased levels of nuclear YAP.