Pumping the brakes on RAS - negative regulators and death effectors of RAS

Abstract

Mutations that activate the RAS oncoproteins are common in cancer. However, aberrant upregulation of RAS activity often occurs in the absence of activating mutations in the RAS genes due to defects in RAS regulators. It is now clear that loss of function of Ras GTPase-activating proteins (RasGAPs) is common in tumors, and germline mutations in certain RasGAP genes are responsible for some clinical syndromes. Although regulation of RAS is central to their activity, RasGAPs exhibit great diversity in their binding partners and therefore affect signaling by multiple mechanisms that are independent of RAS. The RASSF family of tumor suppressors are essential to RAS-induced apoptosis and senescence, and constitute a barrier to RAS-mediated transformation. Suppression of RASSF protein expression can also promote the development of excessive RAS signaling by uncoupling RAS from growth inhibitory pathways. Here, we will examine how these effectors of RAS contribute to tumor suppression, through both RAS-dependent and RAS-independent mechanisms.

Keywords: DAB2IP; GAP; NF1; RAS; RASA1; RASAL2; RASSF.

Fig. 1.

Canonical RAS signaling. RAS is able to activate multiple signaling cascades regulating growth, survival and metabolism. The first downstream effector of RAS identified was the serine/threonine kinase RAF. Activation of RAF initiates the RAF–MEK–ERK kinase cascade. At its terminus, phosphorylated ERK (ERK1/2, also known as MAPK3 and MAPK1, respectively) translocates to the nucleus and activates transcription factors that promote cell cycle progression and inhibit apoptosis. Additionally, RAS can activate PI3K and promote PI3K-AKT signaling. AKT, also known as protein kinase B (PKB), phosphorylates and inhibits Bad, thus derepressing the anti-apoptotic proteins Bcl-2 and Bcl-XL. AKT also activates IκB kinase (IKK), which in turn phosphorylates IκB, thus resulting in the derepression of the anti-apoptotic transcription factor NFκB. NFκB translocates to the nucleus where it activates genes promoting cell survival, migration and EMT.

Fig. 2.

Domain organization of RasGAPs. All RasGAPs contain a GAP domain or GAP-related domain (RasGAP). Apart from that, the RasGAP family is quite diverse in terms of domain composition and arrangement. The GAP domain of RASA1 is at its C-terminus. C2 and plekstrin homology (PH) domains facilitate membrane binding, while the N-terminal SH2 and SH3 domains promote protein-protein interactions. NF1 contains a lipid-binding Sec14-PH module that targets it to membranes. This protein also contains a cysteine/serine-rich domain (CSRD), which can be phosphorylated to modulate NF1 function, and the C-terminal domain (CTD). As members of the SynGAP subfamily, DAB2IP and RASAL2 have N-terminal PH and C2 domains and a centrally located GAP domain. For DAB2IP, the period-like domain (PER) and proline-rich region (PR), which are responsible for binding to TNFR and transducing TNF-mediated apoptosis, are C-terminal to the GAP domain. At the extreme C-terminus lies the leucine zipper (LZ) domain.

Fig. 3.

Overview of DAB2IP activity in cancer. DAB2IP is involved in a variety of cellular processes that overall confer a tumor-suppressive phenotype. As a RasGAP, DAB2IP can directly inhibit RAS activity by facilitating the hydrolysis of RAS-GTP. This suppresses RAF–MEK–ERK and PI3K–AKT signaling. DAB2IP can also disrupt VEGFR-mediated activation of PI3K by directly binding to activated (phosphorylated) VEGFR. DAB2IP is phosphorylated in a TNF-dependent manner. Phosphorylated DAB2IP can activate pro-apoptotic ASK–JNK signaling and simultaneously suppress TNF-mediated activation of NFκB. DAB2IP can also directly bind PI3K and AKT, inhibiting their activity, and these interactions require DAB2IP phosphorylation.

Fig. 4.

Overview of RAS regulation and signaling through effector proteins. RAS is a small GTPase that functions as molecular switch controlling cell growth and survival. Activation and deactivation are controlled by two families of proteins, GEFs and GAPs, respectively. When active, RAS in turn activates pro-proliferative signaling cascades, such as PI3K–AKT, RAF–MEK–ERK and RalGDS. Conversely, RAS can induce apoptosis and senescence by binding a family of effector proteins called RASSF, which function to scaffold active RAS to pro-death proteins, such as the MST kinase and BAX. At least one member of the RASSF family binds to a RasGAP, suggesting that RASSF proteins may directly facilitate the RAS-RasGAP interaction in a feedback mechanism (red box).