MST Kinases Monitor Actin Cytoskeletal Integrity and Signal via c-Jun N-Terminal Kinase Stress-Activated Kinase To Regulate p21^Waf1/Cip1 Stability

Abstract

As well as providing a structural framework, the actin cytoskeleton plays integral roles in cell death, survival, and proliferation. The disruption of the actin cytoskeleton results in the activation of the c-Jun N-terminal kinase (JNK) stress-activated protein kinase (SAPK) pathway; however, the sensor of actin integrity that couples to the JNK pathway has not been characterized in mammalian cells. We now report that the mammalian Ste20-like (MST) kinases mediate the activation of the JNK pathway in response to the disruption of the actin cytoskeleton. One consequence of actin disruption is the JNK-mediated stabilization of p21 Waf1/Cip1 (p21) via the phosphorylation of Thr57. The expression of MST1 or MST2 was sufficient to stabilize p21 in a JNK-and Thr57-dependent manner, while the stabilization of p21 by actin disruption required MST activity. These data indicate that, in addition to being components of the Salvador-Warts-Hippo tumor suppressor network and binding partners of c-Raf and the RASSF1A tumor suppressor, MST kinases serve to monitor cytoskeletal integrity and couple via the JNK SAPK pathway to the regulation of a key cell cycle regulatory protein.The actin cytoskeleton is a dynamic structure that determines cell morphology and motility. In addition, the cytoskeleton also influences other biological functions, such as proliferation, survival, and death, although the mechanistic details linking the cytoskeleton to these processes have not been fully elucidated. Considerable effort has focused on characterizing the signal transduction pathways that control cytoskeletal organization (33). The actin cytoskeleton itself also may regulate cell signaling; for example, mechanical stretching, shear stress, and cytoskeletal disruption each have been shown to activate stress-activated protein kinase (SAPK) pathways (34). Although in Saccharomyces cerevisiae an actin integrity-responsive pathway has been identified in which actin cytoskeleton disassembly results in the activation of the Ssk2p kinase that lies upstream of the Hog1 SAPK pathway (7, 56), an analogous pathway in mammalian cells has not been delineated.SAPK pathways are specific examples of mitogen-activated protein kinase (MAPK) cascades (43). At the bottom of archetypal MAPK pathways are signal-propagating kinases such as ERK1 and ERK2; in the case of SAPK signaling, the similarly positioned kinases are JNK and p38 family members.MAPK are phosphorylated and regulated by MAPK kinases (MAP2K); for c-Jun N-terminal kinase (JNK), the MAP2K are MKK4 and MKK7, while for p38 they are MKK3 and MKK6. Moving stepwise further upstream are MAP3K and MAP4K, although in some pathways there may be no need for a MAP4K, the Ras activation of the MAP3K Raf in the ERK MAPK pathway being one example.Although much recent interest has focused on their antiproliferative and proapoptotic functions as a component of the Salvador-Warts-Hippo tumor suppressor network (31) and as binding partners of the c-Raf MAP3K (42) and RASSF1A tumor suppressor (39), the mammalian Ste2...

Actin-Capping Protein and the Hippo pathway regulate F-actin and tissue growth in Drosophila

“…S4 in the supplementary material), other members of the pathway might be mislocalized in the presence of excess Factin. Interestingly, in mouse fibroblasts, the Hpo orthologs MST1 and MST2 colocalize with F-actin structures and are activated upon F-actin depolymerization (Densham et al, 2009), suggesting that by tethering Hpo pathway components, F-actin dynamics modulates their activity. Finally, the F-actin network might act as a mechanical transducer.…”

“…Interestingly, the Merlin-actin cytoskeleton association is required for growth suppression and inhibition of epidermal growth factor (EGFR) signaling (Curto and McClatchey, 2008). Moreover, F-actin depolymerization promotes activation of the Hpo orthologs MST1 and MST2 in mouse fibroblasts (Densham et al, 2009). These observations suggest a role for Factin dynamics in modulating Hpo pathway activity.…”

The Hippo pathway regulates stem cell proliferation, self-renewal, and differentiation

“…In mammalian cells, Mst1/2 are also activated by binding to Ras association domain family (RASSF) proteins (Oh et al, 2006;Guo et al, 2007) possibly due to alteration of Mst1/2 subcellular localization (Khokhlatchev et al, 2002;Praskova et al, 2004). Recently, Mst1/2 were reported to partially co-localize with actin cytoskeleton, disruption of which leads to mild activation of the kinase (Densham et al, 2009). The activation of Mst1/2 leads to phosphorylation and activation of their direct substrates, Lats1/2 (Chan et al, 2005).…”

Mutual regulation between Hippo signaling and actin cytoskeleton

“…Several studies indicated that the Hippo pathway regulates actin cytoskeleton in Drosophila (Fang and Adler, 2010;Fernández et al, 2011;Lucas et al, 2013). Others demonstrated the interaction of some of the core Hippo pathway components with F-actin regulators and with β-actin itself (Hirota et al, 2000;Yang et al, 2004;Densham et al, 2009;Rauskolb et al, 2011;Visser-Grieve et al, 2011). Although the biological signifi cance of this reverse regulation is not fully understood, it may play an important role in establishing a feedback loop.…”

“…In NIH3T3 cells, immunohistochemistry identifi ed the co-localization between actin cytoskeleton and Mst2. Moreover, inactivation of Rho GTPase by C3 or disruption of F-actin by LatB or CytoD induced the kinase activity of Mst2 (Densham et al, 2009). Understanding if this regulation of Mst2 by actin cytoskeleton activates Hippo pathway needs more examinations.…”