title: Cytohesin 2/ARF6 regulates preadipocyte migration through the activation of ERK1/2.
abstract: Preadipocyte migration is vital for the development of adipose tissue, which plays a crucial role in lipid metabolism. ADP-ribosylation factor 6 (ARF6) small GTPase, which regulates membrane trafficking, is activated by GTP-exchange factors (GEFs) such as cytohesin 2. Cytohesin 2 and ARF6 have previously been implicated in the regulation of 3T3-L1 preadipocyte migration. We investigated here the molecular mechanism underlying the cytohesin 2 and ARF6 mediated regulation of preadipocyte migration. Preadipocyte migration and the activation of ARF6 and ERK1/2 were studied by using a number of approaches, including pharmacological inhibitors, siRNA and the inhibitory peptides. The siRNA mediated down regulation of ARF6 and cytohesin 2 expression confirmed the requirement of both for migration of preadipocytes. Phosphatidylinositol 3-kinase (PI3K) and PI 4,5-bisphosphate (PIP2) have also found to be essential for the cytohesin 2/ARF6 induced preadipocyte migration. Pharmacological inhibition of the activation of ARF6, ERK1/2 or dynamin led to significant reduction in migration of 3T3-L1 preadipocytes. Furthermore, our study revealed the activation of ARF6 and ERK1/2 during migration of preadipocytes. In the migrating preadipocytes, ARF6 activation was inhibited with SecinH3 (cytohesin inhibitor) and LY294002 (PI3K inhibitor) whereas the ERK1/2 phosphorylation was inhibited with SecinH3, LY294002, PBP10 (a PIP2 sequester peptide) and PD98059 (MAPKK inhibitor). However, dynosore (dynamin inhibitor) had inhibited neither ARF6 activation nor ERK1/2 phosphorylation during preadipocyte migration. These results together suggest that cytohesin 2 activates ARF6 in a PI3K dependent manner and then the active ARF6 causes phosphorylation of ERK1/2 during preadipocyte migration.
authors.affiliation: Institute of Life Science 1, College of Medicine, Swansea University, Singleton Park, Swansea SA2 8PP, UK.
authors.forename: Jonathon C B
authors.initials: JC
authors.lastname: Davies
authors.affiliation: Institute of Life Science 1, College of Medicine, Swansea University, Singleton Park, Swansea SA2 8PP, UK.
authors.forename: Salman
authors.initials: S
authors.lastname: Tamaddon-Jahromi
authors.affiliation: Institute of Life Science 1, College of Medicine, Swansea University, Singleton Park, Swansea SA2 8PP, UK.
authors.forename: Riaz
authors.initials: R
authors.lastname: Jannoo
authors.affiliation: Institute of Life Science 1, College of Medicine, Swansea University, Singleton Park, Swansea SA2 8PP, UK. Electronic address: k.venkateswarlu@swansea.ac.uk.
authors.forename: Venkateswarlu
authors.initials: V
authors.lastname: Kanamarlapudi
mesh_terms: D041721:3T3-L1 Cells; D000089982:ADP-Ribosylation Factor 6; D020727:ADP-Ribosylation Factors / Q000502:physiology*; D017667:Adipocytes / Q000166:cytology*; D000818:Animals; D002465:Cell Movement / Q000502:physiology; D004789:Enzyme Activation; D020690:GTPase-Activating Proteins / Q000502:physiology*; D020935:MAP Kinase Signaling System; D051379:Mice; D015398:Signal Transduction
chemical_list: D000089982:ADP-Ribosylation Factor 6; D020690:GTPase-Activating Proteins; C102474:cytohesin-2; D020727:ADP-Ribosylation Factors; C000715727:Arf6 protein, mouse
keywords: ARF6; BFA; Cytohesin 2; Dynamin; Dynasore; ERK; LY294002; PBP10; PD98059; Preadipocytes migration; QS11; SecinH3