Small GTPases最新文献

Angiogenesis is the formation of new blood vessels from pre-existing ones. Angiogenesis requires endothelial cells to change shape and polarity, as well as acquire the ability to directionally migrate ‒ processes that are classically regulated by the Rho family of GTPases. RhoJ (previously TCL) is an endothelium enriched Rho GTPase with a 78% amino acid similarity to the ubiquitously expressed Cdc42. In our recent publication, we demonstrate that α5β1 integrin co-traffics with RhoJ. RhoJ specifically represses the internalization of the active α5β1 conformer, leading to a reduced ability of endothelial cells to form fibronectin fibrils. Surprisingly, this function of RhoJ is in opposition to the role of Cdc42, a known driver of fibrillogenesis. Intriguingly, we discovered that the competition for limiting amounts of the shared effector, PAK3, could explain the ability of these two Rho GTPases to regulate fibrillogenesis in opposing directions. Consequently, RhoJ null mice show excessive fibronectin deposition around retinal vessels, possibly due to the unopposed action of Cdc42. Our work suggests that the functional antagonism between RhoJ and Cdc42 could restrict fibronectin remodelling to sites of active angiogenesis to form a provisional matrix for vessel growth. One correlate of our findings is that RhoJ dependent repression of fibronectin remodelling could be atheroprotective in quiescent vessels.

Malignant melanoma is characterized by mutations in a number of driver genes, most notably BRAF and NRAS. Recent genomic analyses revealed that 4-9% of sun-exposed melanomas bear activating mutations in RAC1, which encodes a small GTPase that is known to play key roles in cell proliferation, survival, and migration. The RAC1 protein activates several effector pathways, including Group A p21-activated kinases (PAKs), phosphoinositol-3-kinases (PI3Ks), in particular the beta isoform, and the serum-response factor/myocardin-related transcription factor (SRF/MRTF). Having previously shown that inhibition of Group A PAKs impedes oncogenic signalling from RAC1^P29S, we here extend this analysis to examine the roles of PI3Ks and SRF/MRTF in melanocytes and/or in a zebrafish model. We demonstrate that a selective Group A PAK inhibitor (Frax-1036), a pan-PI3K (BKM120), and two PI3Kβ inhibitors (TGX221, GSK2636771) impede the growth of melanoma cells driven by mutant RAC1 but not by mutant BRAF, while other PI3K selective inhibitors, including PI3Kα, δ and γ, are less effective. Using these compounds as well as an SRF/MRTF inhibitor (CCG-203,971), we observed similar results in vivo, using embryonic zebrafish development as a readout. These results suggest that targeting Group A PAKs, PI3Kβ, and/or SRF/MRTF represent a promising approach to suppress RAC1 signalling in malignant melanoma.

Cytokinesis in fission yeast involves actomyosin ring constriction concurrent to septum synthesis followed by septum digestion resulting in cell separation. A recent report indicates that endocytosis is required for septum synthesis and cell separation. The conserved GTPase Cdc42 is required for membrane trafficking and promotes endocytosis. Cdc42 is activated by Guanine nucleotide exchange factors (GEFs). Cdc42 GEFs have been shown to promote timely initiation of septum synthesis and proper septum morphology. Here we show that Cdc42 promotes the recruitment of the major primary septum synthesizing enzyme Bgs1 and consequent ring constriction. Cdc42 is also required for proper localization of the septum digesting glucanases at the division site. Thus, Cdc42 is required to promote multiple steps during cytokinesis.

Fatty acid synthase (FASN) is commonly overexpressed in prostate cancer and associated with tumour progression. FASN is responsible for de novo synthesis of the fatty acid palmitate; the building block for protein palmitoylation. A functional role for FASN in regulating cell proliferation is widely accepted. We recently reported that FASN activity can also mediate prostate cancer HGF-mediated cell motility. Moreover, we found that modulation of FASN expression specifically impacts on the palmitoylation of RhoU. Findings we will describe here. We now report that loss of FASN expression also impairs HGF-mediated cell dissociation responses. Taken together our results provide compelling evidence that FASN activity directly promotes cell migration and supports FASN as a potential therapeutic target in metastatic prostate cancer.

The Ran pathway has a well-described function in nucleocytoplasmic transport, where active Ran dissociates importin/karyopherin-bound cargo containing a nuclear localization signal (NLS) in the nucleus. As cells enter mitosis, the nuclear envelope breaks down and a gradient of active Ran forms where levels are highest near chromatin. This gradient plays a crucial role in regulating mitotic spindle assembly, where active Ran binds to and releases importins from NLS-containing spindle assembly factors. An emerging theme is that the Ran gradient also regulates the actomyosin cortex for processes including polar body extrusion during meiosis, and cytokinesis. For these events, active Ran could play an inhibitory role, where importin-binding may help promote or stabilize a conformation or interaction that favours the recruitment and function of cortical regulators. For either spindle assembly or cortical polarity, the gradient of active Ran determines the extent of importin-binding, the effects of which could vary for different proteins.

The primary cilium and the immunological synapse are both specialized functional plasma membrane domains that share several similarities. Signalling output of membrane domains is regulated, spatially and temporally, by segregating and focusing lipids and proteins. ARL3, a small GTPase, plays a major role in concentrating lipid-modified proteins in both the immunological synapse and the primary cilia. Here in this review we will introduce the role of ARL3 in health and disease and its role in polarizing signalling at the primary cilia and immunological synapses.

Cancer is the second leading cause of death globally, and colorectal cancer (CRC) is among the five most common cancers. The small GTPase KRAS is an oncogene that is mutated in ~30% of all CRCs. Pharmacological treatments of CRC are currently unsatisfactory, but much hope rests on network-centric approaches to drug development and cancer treatment. These approaches, however, require a better understanding of how networks downstream of Ras oncoproteins are connected in a particular tissue context - here colon and CRC. Previously we have shown that competition for binding to a 'hub' protein, such as Ras, can induce a rewiring of signal transduction networks. In this study, we analysed 56 established and predicted effectors that contain a structural domain with the potential ability to bind to Ras oncoproteins and their link to pathways coordinating intestinal homoeostasis and barrier function. Using protein concentrations in colon tissue and Ras-effector binding affinities, a computational network model was generated that predicted how effectors differentially and competitively bind to Ras in colon context. The model also predicted both qualitative and quantitative changes in Ras-effector complex formations with increased levels of active Ras - to simulate its upregulation in cancer - simply as an emergent property of competition for the same binding interface on the surface of Ras. We also considered how the number of Ras-effector complexes at the membrane can be increased by additional domains present in some effectors that are recruited to the membrane in response to specific conditions (inputs/stimuli/growth factors) in colon context and CRC.

LRRK2 is a multi-domain Ser/Thr kinase that is associated with inherited and sporadic cases of Parkinson's disease. Many mutations linked to disease are associated within a central ROC-COR regulatory region and the subsequent kinase domain, leading to enhanced catalytic activity. The N-terminus of human LRRK2 consists of armadillo repeat motifs (ARMs) followed by ankyrin repeats (ANKs). Recently, Rab GTPases have emerged as key players in LRRK2 function, both as substrates of the kinase, and as regulators of the catalytic activity. Rabs recruit effector proteins via their GTP-dependent switch 1 and 2 regions to distinct sub-cellular compartments to regulate membrane trafficking. LRRK2 phosphorylates Rab8, Rab10 and Rab12 in switch 2, and this activity is regulated via interactions with Rab29. Furthermore, the related Rab32-subfamily GTPases, Rab32 and Rab38, have also been shown to interact with LRRK2. Here, we have mapped the interactions of the Rab32-subfamily to the ARM domain of LRRK2. The complexes are dependent on the GTP state of the Rabs in vitro, implying that LRRK2 may be an effector of the Rab32-subfamily of small GTPases. X-ray crystal structures of the Rab32-family GTPases and subsequent mutational studies reveal that a positively charged residue in switch 1 is critical for binding of Rab32/38 to LRRK2. Homology modelling and mutational analyses of the ARM domain point to a patch of negatively charged residues that contribute to complex formation. These structural and biochemical studies provide a framework for understanding the molecular basis for Rab regulation of LRRK2 and its role in Parkinson's disease.

Glioblastoma is an aggressive and incurable form of brain cancer. Both mutation analysis in human glioblastoma and mouse modelling studies have shown that aberrant activation of the PI 3-kinase pathway is a central driver of glioblastoma malignancy. The small GTPase Rac is activated downstream of this pathway, mediating a subset of the effects of aberrant PI 3-kinase pathway activation. Here I discuss the current state of our knowledge on Rac activation mechanisms in glioblastoma. Current knowledge on roles for specific PI 3-kinase pathway responsive Rac guanine nucleotide exchange factors in glioblastoma is reviewed. Rac is best known for its role in promoting cell motility and invasion, but there is also evidence for roles in multiple other cellular processes with cancer relevance, including proliferation, differentiation, apoptosis, DNA damage responses, metabolism, angiogenesis and immunosuppression. I review what is known about the role of Rac in these processes in glioblastoma. Finally, I assess possible strategies to inhibit this pathway in glioblastoma through either direct inhibition of Rac or inhibition of upstream activators or downstream mediators of Rac signalling.

Cell migration frequently involves the formation of lamellipodial protrusions, the initiation of which requires Rac GTPases signalling to heteropentameric WAVE regulatory complex (WRC). While Rac-related RhoG and Cdc42 can potently stimulate lamellipodium formation, so far presumed to occur by upstream signalling to Rac activation, we show here that the latter can be bypassed by RhoG and Cdc42 given that WRC has been artificially activated. This evidence arises from generation of B16-F1 cells simultaneously lacking both Rac GTPases and WRC, followed by reconstitution of lamellipodia formation with specific Rho-GTPase and differentially active WRC variant combinations. We conclude that formation of canonical lamellipodia requires WRC activation through Rac, but can possibly be tuned, in addition, by WRC interactions with RhoG and Cdc42.