MEKK1-dependent phosphorylation of calponin-3 tunes cell contractility

Abstract

MEKK1 (also known as MAP3K1), which plays a major role in MAPK signaling, has been implicated in mechanical processes in cells, such as migration. Here, we identify the actin-binding protein calponin-3 as a new MEKK1 substrate in the signaling that regulates actomyosinbased cellular contractility. MEKK1 colocalizes with calponin-3 at the actin cytoskeleton and phosphorylates it, leading to an increase in the cell-generated traction stress. MEKK1-mediated calponin-3 phosphorylation is attenuated by the inhibition of myosin II activity, the disruption of actin cytoskeletal integrity and adhesion to soft extracellular substrates, whereas it is enhanced upon cell stretching. Our results reveal the importance of the MEKK1-calponin-3 signaling pathway to cell contractility.

A cryptic tubulin-binding domain links MEKK1 to microtubule remodelling

“…How MEKK1 is regulated and the specific substrates at the leading edge both remain pertinent questions. Logical candidates for interaction partners include: Rho GTPases, previously shown to regulate MEKK1 and actin dynamics (Dogterom and Koenderink, 2019;Gallagher et al, 2004); and Calponin-3, a MEKK1 kinase substrate within the actin cytoskeleton known to impact cellular contractility (Hirata et al, 2016).…”

“…The ability to phosphorylate or ubiquitinate substrates under different circumstances allows MEKK1 to function as a central signalling hub-it can promote signalling via the ERK, JNK, p38, NF-κB, or other pathways depending on cellular context (Lee et al, 1997(Lee et al, , 1998Xia et al, 1998;Yujiri et al, 1998). Direct targets for MEKK1 kinase activity include: MAP2Ks, leading to activation of JNK, ERK, and p38 pathways; Inhibitor of κ kinases (IκKs), leading to NF-κB transcription factor family activation (Deak and Templeton, 1997;Lee et al, 1998;Xu et al, 1996); or migrationlinked proteins such as calponin-3 (Hirata et al, 2016). MEKK1 can ubiquitinate itself in a phosphorylation-dependent manner (Witowsky and Johnson, 2003), leading to attenuation of its own kinase activity, and the diminished activity of downstream ERK and JNK pathways.…”

Calponin-3 deficiency augments contractile activity, plasticity, fibrogenic response and Yap/Taz transcriptional activation in lens epithelial cells and explants

“…CNN3 has been shown to be involved in trophoblast cell fusion, wound healing and neural tube morphogenesis largely through its role in actin cytoskeletal organization 15,22,23 . A definitive role has also been recently demonstrated for CNN3 in regulating contractile activity and force generation in different types of cells 21,24 .Our recent cDNA microarray and RNA-seq based analysis of gene expression profiles of neonatal and adult mouse lenses revealed relatively high level expression of acidic calponin (CNN3) with little to no expression of basic (CNN1) and neutral (CNN2) calponin isoforms 25 . Intrigued by this observation, we initiated studies to understand the significance of what appears to be tissue-specific expression profile of CNN-3 in the lens, and to evaluate a possible role for CNN3 in lens function.…”

“…CNN3 has been shown to be involved in trophoblast cell fusion, wound healing and neural tube morphogenesis largely through its role in actin cytoskeletal organization 15,22,23 . A definitive role has also been recently demonstrated for CNN3 in regulating contractile activity and force generation in different types of cells 21,24 .…”

“…Interestingly, while CNN1 and CNN2 null mice grow and breed normally 13 , CNN3 null mice display embryonic and neonatal lethality with severe defects in brain development 15 , indicating distinct roles for different calponin isoforms despite their shared biological roles in actin cytoskeletal networks, signaling and contraction 13 . Calponins are known to be serine/threonine phosphorylated by various kinases including protein kinase C, Ca 2+ /calmodulin kinase II, Rho kinase (CNN3) and MEKK1 (CNN3), with phosphorylation being reported to decrease actin interaction and suppression of the inhibitory effect of calponin on myosin Mg 2+ ATPase activity 13,[18][19][20][21][22] . CNN3 has been shown to be involved in trophoblast cell fusion, wound healing and neural tube morphogenesis largely through its role in actin cytoskeletal organization 15,22,23 .…”