A Rab10:RalA G protein cascade regulates insulin-stimulated glucose uptake in adipocytes.
Bottom Line: Once activated, Rab10 can increase the GTP binding of RalA by recruiting the Ral guanyl nucleotide exchange factor, Rlf/Rgl2.Overexpression of membrane-tethered Rlf compensates for the loss of Rab10 in Glut4 translocation, suggesting that Rab10 recruits Rlf to membrane compartments for RalA activation and that RalA is downstream of Rab10.Together these studies identify a new G protein cascade in the regulation of insulin-stimulated Glut4 trafficking and glucose uptake.
Affiliation: Life Sciences Institute, University of Michigan Medical School, Ann Arbor, MI 48109.Show MeSH
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Mentions: Rab10 and RalA are established components of the insulin regulatory network and are required for Glut4 translocation and glucose uptake in adipocytes (Chen et al., 2007; Sano et al., 2007, 2008). Because Rlf appears to be an intermediate in this activation cascade, we hypothesized that the GEF may also be essential. We depleted Rlf using siRNA in 3T3-L1 adipocytes before assaying 2-deoxyglucose uptake. The expression of Rlf was reduced by ∼70% after incubation with siRNA oligos (Figure 5B). Knockdown of Rlf caused a 15–20% loss in insulin-stimulated glucose uptake, comparable to the effect achieved by knockdown of RalA (Chen et al., 2007) and Rab10 (Figure 5A). Because RalA and Rab10 are required for translocation of Glut4 storage vesicles to the plasma membrane upon insulin stimulation (Chen et al., 2007; Sano et al., 2007), we also tested the effect of Rlf knockdown on Glut4 translocation. Knockdown of Rlf in 3T3-L1 adipocytes stably expressing Myc-Glut4–enhanced green fluorescent protein (eGFP; Bogan et al., 2001; Lodhi et al., 2007) caused a significant reduction (∼25%) in surface Glut4 levels similar to the effect of knockdown of RalA and Rab10 (Figure 5, C–E). Together these data suggest that Rlf is a new component required for Glut4 translocation and is essential for maximal glucose uptake in adipocytes.
Affiliation: Life Sciences Institute, University of Michigan Medical School, Ann Arbor, MI 48109.