SHP-2 acts via ROCK to regulate the cardiac actin cytoskeleton

Abstract

SUMMARYNoonan syndrome is one of the most common causes of human congenital heart disease and is frequently associated with missense mutations in the protein phosphatase SHP-2. Interestingly, patients with acute myelogenous leukemia (AML), acute lymphoblastic leukemia (ALL), juvenile myelomonocytic leukemia (JMML) and LEOPARD syndrome frequently carry a second, somatically introduced subset of missense mutations in SHP-2. To determine the cellular and molecular mechanisms by which SHP-2 regulates heart development and, thus, understand how Noonan-associated mutations affect cardiogenesis, we introduced SHP-2 encoding the most prevalent Noonan syndrome and JMML mutations into Xenopus embryos. Resulting embryos show a direct relationship between a Noonan SHP-2 mutation and its ability to cause cardiac defects in Xenopus; embryos expressing Noonan SHP-2 mutations exhibit morphologically abnormal hearts, whereas those expressing an SHP-2 JMML-associated mutation do not. Our studies indicate that the cardiac defects associated with the introduction of the Noonan-associated SHP-2 mutations are coupled with a delay or arrest of the cardiac cell cycle in M-phase and a failure of cardiomyocyte progenitors to incorporate into the developing heart. We show that these defects are a result of an underlying malformation in the formation and polarity of cardiac actin fibers and F-actin deposition. We show that these defects can be rescued in culture and in embryos through the inhibition of the Rho-associated, coiled-coil-containing protein kinase 1 (ROCK), thus demonstrating a direct relationship between SHP-2 N308D and ROCK activation in the developing heart.

ROCKs cause SHP-wrecks and broken hearts

“…An additional remarkable finding from this study was that regardless of the fact that the N308D SHP-2 variant was overexpressed throughout the entire animal, the only apparent defect was improper heart formation 13 . We surmise that the nature of the specificity of the cardiac defects induced by N308D could relate to the fact that ROCK is particularly abundant in developing hearts 23 and that precise regulation of this pathway is crucial for the appropriate regionalized co-ordination of myocyte growth and migration required for cardiac morphogenesis.…”

“…Given the findings that ROCK inhibition can fully rescue cardiac malformations induced by SHP-2 variants, 13 , 25 we propose that novel treatment strategies with ROCK inhibitors might be efficacious for NS patients. In this regard, it is promising that the ROCK inhibitor, Fasudil, has been approved for use in adult patients to treat vascular defects and hypertension 26 , 27 .…”

“…In contrast, aberrant fibroblast growth factor receptor-mediated ERK signaling has been implicated in NS-associated SHP-2 cardiac phenotypes in vivo 6 , 7 . However, the role of SHP-2 is not confined to these growth conduits as more recent data indicates that SHP-2 can influence actin remodeling and cytoskeletal shape by modulating the RhoA-ROCK pathway 12 , 13 . Indeed, recent mechanistic studies indicate that both Rho-selective GAPs (p190 RhoGAP A and B), and the essential Rho effector, rho-associated kinase (ROCK) are bona fide SHP-2 substrates 9 - 11 …”

The role of the protein tyrosine phosphatase SHP2 in cardiac development and disease

“…Interestingly, NS SHP2 mutations, through hyperactivation of ROCK, the downstream effector of RhoA, lead to defective formation and polarity of cardiac actin fibers and F-actin deposition in Xenopus embryos (Table 2). This, in turn, results in smaller hearts that fail to undergo complete looping and chamber formation and have delayed morphogenetic movements [58], suggesting that SHP2’s actions on actin polymerization in the myocyte itself might be important for cardiac looping (Figure 4, lower panels).…”

Expanding the genetic toolkit in Xenopus: Approaches and opportunities for human disease modeling

“…Xenopus animals and oocytes are used extensively to understand normal organ function and disease in humans (Labonne and Zorn, 2015), including cardiac congenital heart disorders and heterotaxy (Boskovski et al, 2013; Duncan and Khokha, 2016; Fakhro et al, 2011; Kaltenbrun et al, 2011; Langdon et al, 2012; 2007), gastrointestinal and pancreatic diseases (Kofent and Spagnoli, 2016; Pearl et al, 2009; 2011; Salanga and Horb, 2015; Womble et al, 2016), endocrine functions and disorders (Buchholz, 2015), kidney disease (Lienkamp, 2016), lung development (Rankin et al, 2011; 2015; Wallmeier et al, 2014), cancer (Chernet and Levin, 2013; Cross and Powers, 2009; Hardwick and Philpott, 2015; Haynes-Gilmore et al, 2014; Van Nieuwenhuysen et al, 2015; Wylie et al, 2015), ciliopathies (Kim et al, 2010; Klos Dehring et al, 2013; Ma et al, 2014), orofacial defects (Dickinson, 2016), and neurodevelopmental disorders (Erdogan et al, 2016; Pratt and Khakhalin, 2013). Looking forward, Xenopus is poised to take advantage of the new developments in genomics and genome engineering to better understand the molecular mechanisms underlying human disease (Harland and Grainger, 2011; Labonne and Zorn, 2015).…”