Cell biology. Smurfing at the leading edge.
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Animals↗
Biomedical subjects
Alan Hall
Publications and source records attributed to Alan Hall.
Mechanisms in LPA-induced tumor cell migration: critical role of phosphorylated ERK.
Lysophosphatidic acid (LPA) is a serum-borne phospholipid with hormone and growth factor-like properties. LPA has been shown to modulate tumor cell invasion and malignant cell growth. Here, we report that two human pancreatic carcinoma cell lines, PANC-1 and BxPC-3, express functionally active LPA receptors coupled to pertussis toxin-sensitive Gi/o-proteins. In contrast to other cell types, LPA does not act as a mitogen, but is an efficacious stimulator of cell migration of these tumor cells. LPA-induced chemotaxis is markedly dependent on activation of PTX-sensitive heterotrimeric G-proteins, on activation of the small GTPases Ras, Rac and RhoA, and on GTPase-dependent activation of ERK. LPA-induced ERK activation results in a transient translocation of the phosphorylated ERK to newly forming focal contact sites at the leading edge of the migrating cells. Inhibition of ERK activation and its subsequent translocation impaired LPA-induced chemotaxis and LPA-induced actin reorganization. Thus, pancreatic tumor cell migration in response to LPA is essentially controlled by activation of a Gi/o-ERK pathway and requires the LPA-induced activation of Ras, Rac1 and RhoA.
Actins↗
Cdc42 regulates GSK-3beta and adenomatous polyposis coli to control cell polarity.
Cell polarity is a fundamental property of all cells. In higher eukaryotes, the small GTPase Cdc42, acting through a Par6-atypical protein kinase C (aPKC) complex, is required to establish cellular asymmetry during epithelial morphogenesis, asymmetric cell division and directed cell migration. However, little is known about what lies downstream of this complex. Here we show, through the use of primary rat astrocytes in a cell migration assay, that Par6-PKCzeta interacts directly with and regulates glycogen synthase kinase-3beta (GSK-3beta) to promote polarization of the centrosome and to control the direction of cell protrusion. Cdc42-dependent phosphorylation of GSK-3beta occurs specifically at the leading edge of migrating cells, and induces the interaction of adenomatous polyposis coli (Apc) protein with the plus ends of microtubules. The association of Apc with microtubules is essential for cell polarization. We conclude that Cdc42 regulates cell polarity through the spatial regulation of GSK-3beta and Apc. This role for Apc may contribute to its tumour-suppressor activity.
Adenomatous Polyposis Coli Protein↗
Cell polarity: Par6, aPKC and cytoskeletal crosstalk.
Par6 and atypical protein kinase C are key players in the establishment of cell polarity. First discovered in Caenorhabditis elegans, the function of this protein complex is conserved in all multicellular organisms. Recent work is beginning to throw light on how it converts information generated by extracellular cues into intracellular asymmetry.
Animals↗
Rho GTPases in cell biology.
Rho GTPases are molecular switches that control a wide variety of signal transduction pathways in all eukaryotic cells. They are known principally for their pivotal role in regulating the actin cytoskeleton, but their ability to influence cell polarity, microtubule dynamics, membrane transport pathways and transcription factor activity is probably just as significant. Underlying this biological complexity is a simple biochemical idea, namely that by switching on a single GTPase, several distinct signalling pathways can be coordinately activated. With spatial and temporal activation of multiple switches factored in, it is not surprising to find Rho GTPases having such a prominent role in eukaryotic cell biology.
Animals↗
Activated Galphaq family members induce Rho GTPase activation and Rho-dependent actin filament assembly.
Rho GTPase is required for actin filament assembly and serum response element (SRE)-dependent gene transcription. Certain G protein-coupled receptors (GPCRs) induce Rho-dependent responses, but the intermediary signaling steps are poorly understood. The heterotrimeric Galpha12 family can induce Rho-dependent responses. In contrast, there are conflicting reports on the role of the Galphaq family in Rho signaling. We report that expression of activated Galphaq members, or activation of endogenous Galphaq via GPCR stimulation, induces SRE reporter activation via Rho, and increased GTP-Rho levels. Moreover, microinjection of activated Galphaq in fibroblasts induces actin stress fiber formation via Rho. Galphaq functionally cooperates with Lbc Rho guanine nucleotide exchange factor. Overall, these findings indicate that Galphaq family signals are sufficient to induce Rho-dependent cellular responses.
3T3 Cells↗
Guanine nucleotide exchange factors for Rho GTPases: turning on the switch.
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Animals↗
The Rho exchange factor Net1 is regulated by nuclear sequestration.
Net1 is a guanine nucleotide exchange factor specific for the small GTPase Rho. Oncogenic activation of Net1 occurs by truncation of the N-terminal part of the protein, which functions as a negative regulatory domain. Here, we have investigated the mechanism of Net1 regulation via its N terminus. We find that Net1 localizes to the nucleus, whereas oncogenic Net1 is found in the cytoplasm. Nuclear import of Net1 is mediated by two nuclear localization signals present in the N terminus of the protein, and forced cytoplasmic localization of Net1 is sufficient to activate Rho. In addition, the pleckstrin homology (PH) domain of Net1 acts as a nuclear export signal. Because an amino acid substitution in the PH domain that inhibits guanine nucleotide exchange factor activity does not inhibit nuclear export, we conclude that this PH domain has at least two functions. Together, our results suggest that Net1 can shuttle in and out of the nucleus, and that activation of Rho by Net1 is controlled by changes in its subcellular localization.
3T3 Cells↗
Rho GTPases in transformation and metastasis.
During the development and progression of human cancer, cells undergo numerous changes in morphology, proliferation, and transcriptional profile. Over the past couple of decades there have been intense efforts to understand the molecular mechanisms involved, and members of the Ras superfamily of small GTPases have emerged as important players. Mutated versions of the Ras genes were first identified in human cancers some 20 years ago, but more recently, the Rho branch of the family has been receiving increased attention. In addition to the experimental evidence implicating Rho GTPase signaling in promoting malignant transformation, genetic analysis of human cancers has now revealed a few examples of direct alterations in the genes encoding regulators of Rho GTPases. In this review, we discuss the evidence implicating Rho GTPases in transformation and metastasis, as well as the progress made toward identifying their biochemical mechanism of action.
Animals↗
Vav regulates activation of Rac but not Cdc42 during FcgammaR-mediated phagocytosis.
Phagocytosis is the process whereby cells direct the spatially localized, receptor-driven engulfment of particulate materials. It proceeds via remodeling of the actin cytoskeleton and shares many of the core cytoskeletal components involved in adhesion and migration. Small GTPases of the Rho family have been widely implicated in coordinating actin dynamics in response to extracellular signals and during diverse cellular processes, including phagocytosis, yet the mechanisms controlling their recruitment and activation are not known. We show herein that in response to ligation of Fc receptors for IgG (FcgammaR), the guanine nucleotide exchange factor Vav translocates to nascent phagosomes and catalyzes GTP loading on Rac, but not Cdc42. The Vav-induced Rac activation proceeds independently of Cdc42 function, suggesting distinct roles for each GTPase during engulfment. Moreover, inhibition of Vav exchange activity or of Cdc42 activity does not prevent Rac recruitment to sites of particle attachment. We conclude that Rac is recruited to Fcgamma membrane receptors in its inactive, GDP-bound state and that Vav regulates phagocytosis through subsequent catalysis of GDP/GTP exchange on Rac.
Animals↗
Rac regulates endothelial morphogenesis and capillary assembly.
Endothelial cells undergo branching morphogenesis to form capillary tubes. We have utilized an in vitro Matrigel overlay assay to analyze the role of the cytoskeleton and Rho GTPases during this process. The addition of matrix first induces changes in cell morphology characterized by the formation of dynamic cellular protrusions and the assembly of discrete aggregates or cords of aligned cells resembling primitive capillary-like structures, but without a recognizable lumen. This is followed by cell migration leading to the formation of a complex interconnecting network of capillary tubes with readily identifiable lumens. Inhibition of actin polymerization or actin-myosin contraction inhibits cell migration but has no effect on the initial changes in endothelial cell morphology. However, inhibition of microtubule dynamics prevents both the initial cell shape changes as well as cell migration. We find that the small GTPase Rac is essential for the matrix-induced changes in endothelial cell morphology, whereas p21-activated kinase, an effector of Rac, is required for cell motility. We conclude that Rac integrates signaling through both the actin and microtubule cytoskeletons to promote capillary tube assembly.
Actins↗