srGAP2 Arginine Methylation Regulates Cell Migration and Cell Spreading through Promoting Dimerization

Abstract

The Slit-Robo GTPase-activating proteins (srGAPs) are critical for neuronal migration through inactivation of Rho GTPases Cdc42, Rac1, and RhoA. Here we report that srGAP2 physically interacts with protein arginine methyltransferase 5 (PRMT5). srGAP2 localizes to the cytoplasm and plasma membrane protrusion. srGAP2 knockdown reduces cell adhesion spreading and increases cell migration, but has no effect on cell proliferation. PRMT5 binds to the N terminus of srGAP2 (225-538 aa) and methylates its C-terminal arginine residue Arg-927. The methylation mutant srGAP2-R927A fails to rescue the cell spreading rate, is unable to localize to the plasma membrane leading edge, and perturbs srGAP2 homodimer formation mediated by the F-BAR domain. These results suggest that srGAP2 arginine methylation plays important roles in cell spreading and cell migration through influencing membrane protrusion.Cell migration is essential for a wide variety of physiological and pathological processes such as embryonic development, wound tissue repair, and tumor metastasis (1-2). Classically, the major driving force for migration in mammalian cells is thought to be provided by actin polymerization at the cell protrusion of the front edge and F-actin depolymerization to retract cell body at the rear (3). These coordinated processes are regulated by small Rho GTPases, especially Cdc42, Rac1, and RhoA (4 -5). Similar to other GTPases, Rho GTPases act as intracellular molecular switches, cycling between a GDPbound inactive form and a GTP-bound active form. The two interconvertible forms are controlled by two classes of proteins, guanine nucleotide exchange factors (GEFs), 2 which promote the exchange of GDP for GTP, and GTPase-activating proteins (GAPs), which increase the GTPase activity of Rho GTPases through converting GTP to GDP, thereby closing the switch (6 -7).srGAP is a family of RhoGAP proteins, identified in the SlitRobo signal pathway. The srGAP family consists of three members, srGAP1, -2, and -3. All of them possess three conserved domains, F-BAR, RhoGAP, and SH3 (8). The function of RhoGAP domain of srGAP family protein is extensively studied because of its role in the negative regulation of Rho GTPase activities important for cytoskeleton rearrangement (8 -9). In addition to the RhoGAP domain, studies have found that the F-BAR domain-containing proteins function as key regulators in membrane remodeling in eukaryotes. Each of these proteins shares a similar quaternary "banana-like" structure, which forms elongated dimers mediated by the anti-parallel association of ␣ helices in each monomer (10 -11). Moreover, positive charges within the "banana-like" structure are aligned to interact with negative charges of the membrane via electrostatic interactions (12). Therefore, the F-BAR domains act as membrane-associated scaffolds and deform cell membrane independently of its F-actin bundling activity (13-14).Based on structural characteristics and phylogenetic analysis, the BAR domain super family includes "classical" BAR domain, F-BAR...

miR-29 Promotes Murine Osteoclastogenesis by Regulating Osteoclast Commitment and Migration

“…srGAP2 is a Rho-GTPase-activating protein that participates in the same signaling pathway as CDC42 (35). Although srGAP2 function in osteoclasts has not been investigated, it has been shown to repress cell migration during neuronal development (35,57) (Fig.…”

“…8C). CDC42 is important for osteoclast function and migration, and srGAP2 participates in the same signaling pathway as CDC42 (35). Although expressed in osteoclasts, srGAP2 has not been studied in the osteoclast lineage.…”

PKA-induced dimerization of the RhoGAP DLC1 promotes its inhibition of tumorigenesis and metastasis

“…Apart from DLC1, other GAPs are regulated by dimerization; for example, the C-terminus of Slit-Robo GTPase-activating protein 2 (srGAP2) has been shown to be methylated by protein arginine methyltransferase 5, leading to the homodimerization of srGAP2 (ref. 32). The formation of srGAP2 homodimers promotes neuronal cell membrane protrusion and cell spreading.…”

ArabidopsisFloral Initiator SKB1 Confers High Salt Tolerance by Regulating Transcription and Pre-mRNA Splicing through Altering Histone H4R3 and Small Nuclear Ribonucleoprotein LSM4 Methylation

“…Response to Salt Stress SKB1, or PRMT5, methylates Arg residues in multiple histone and nonhistone proteins, including H4R3 Wang et al, 2007;Schmitz et al, 2008) and LSM4 in plants and H4R3, H3R8 (Pal et al, 2004), p53 (Jansson et al, 2008), p300 (Yang et al, 2009), 130-kD cis-Golgi matrix protein (GM130) (Zhou et al, 2010), slit-robo Rho GTPase activating protein 2 (srGAP2) (Guo and Bao, 2010), Ribosomal protein S10 (Rps10) (Ren et al, 2010) and SM (Friesen et al, 2001;Meister and Fischer, 2002) in animals. SKB1 localizes to both the cytoplasm and the nucleus.…”

“…SKB1, also named protein arginine methyltransferase 5 (PRMT5), is a type II Arg methyltransferase that catalyzes Arg symmetric dimethylation (Bedford and Richard, 2005;Pahlich et al, 2006;Bedford, 2007;Bedford and Clarke, 2009). In mammalian cells, PRMT5 methylates a wide spectrum of substrates, including histone and nonhistone proteins, to regulate gene transcription, RNA elongation, pre-mRNA splicing, protein interaction, and protein stability (Kwak et al, 2003;Pal et al, 2003;Liu et al, 2007;Chari et al, 2008;Guo and Bao, 2010;Ren et al, 2010;Zhou et al, 2010). We and others previously reported that Arabidopsis SKB1/PRMT5 promotes flowering by suppressing the expression of FLC through H4R3sme2 in the promoter region (Wang et al, 2007;Schmitz et al, 2008).…”