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A Rab10:RalA G protein cascade regulates insulin-stimulated glucose uptake in adipocytes.

Karunanithi S, Xiong T, Uhm M, Leto D, Sun J, Chen XW, Saltiel AR - Mol. Biol. Cell (2014)

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.

View Article: PubMed Central - PubMed

Affiliation: Life Sciences Institute, University of Michigan Medical School, Ann Arbor, MI 48109.

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Related in: MedlinePlus

Rab10 and RalA reside in insulin-sensitive GSVs. (A) Subcellular fractionation of 3T3-L1 adipocytes. Indicated fractions were run on SDS–PAGE and immunoblotted with antibodies as specified. HDM, high-density microsomes; LDM, low-density microsomes; N/M, nuclear/mitochondrial; PM, plasma membrane. L.E and D.E. denote light and dark exposures of the RalA blot, respectively. (B) LDM fractions from several independent experiments as in A were pooled to obtain an enriched fraction, which was then run on 14% iodixanol gradient. Fractions 1–11 represent lighter to denser peaks in the gradient. Insulin-sensitive pool of GSVs (fractions 8–10) is indicated by a box (peak 1). Peak 2 represents the second-less-responsive recycling endosomes, containing Glut4 (fractions 2–4). (C) Proposed model for the role of Rab10 in Glut4 translocation. Rab10 recruits Rlf to insulin-sensitive GSVs to promote GEF activity on RalA. Activated RalA allows Glut4 translocation through its interactions with the myosin motor and the exocyst.
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Figure 7: Rab10 and RalA reside in insulin-sensitive GSVs. (A) Subcellular fractionation of 3T3-L1 adipocytes. Indicated fractions were run on SDS–PAGE and immunoblotted with antibodies as specified. HDM, high-density microsomes; LDM, low-density microsomes; N/M, nuclear/mitochondrial; PM, plasma membrane. L.E and D.E. denote light and dark exposures of the RalA blot, respectively. (B) LDM fractions from several independent experiments as in A were pooled to obtain an enriched fraction, which was then run on 14% iodixanol gradient. Fractions 1–11 represent lighter to denser peaks in the gradient. Insulin-sensitive pool of GSVs (fractions 8–10) is indicated by a box (peak 1). Peak 2 represents the second-less-responsive recycling endosomes, containing Glut4 (fractions 2–4). (C) Proposed model for the role of Rab10 in Glut4 translocation. Rab10 recruits Rlf to insulin-sensitive GSVs to promote GEF activity on RalA. Activated RalA allows Glut4 translocation through its interactions with the myosin motor and the exocyst.

Mentions: We performed subcellular fractionation of 3T3-L1 adipocytes to assess the insulin-dependent changes in the cellular localization of RalA and Rab10. As expected, Glut4, IRAP, and RalA were enriched in the plasma membrane fractions upon insulin stimulation, with a concurrent loss of these proteins in the low-density microsome (LDM) fractions. Of interest, Rab10 localization mirrored that of RalA (Figure 7A).


A Rab10:RalA G protein cascade regulates insulin-stimulated glucose uptake in adipocytes.

Karunanithi S, Xiong T, Uhm M, Leto D, Sun J, Chen XW, Saltiel AR - Mol. Biol. Cell (2014)

Rab10 and RalA reside in insulin-sensitive GSVs. (A) Subcellular fractionation of 3T3-L1 adipocytes. Indicated fractions were run on SDS–PAGE and immunoblotted with antibodies as specified. HDM, high-density microsomes; LDM, low-density microsomes; N/M, nuclear/mitochondrial; PM, plasma membrane. L.E and D.E. denote light and dark exposures of the RalA blot, respectively. (B) LDM fractions from several independent experiments as in A were pooled to obtain an enriched fraction, which was then run on 14% iodixanol gradient. Fractions 1–11 represent lighter to denser peaks in the gradient. Insulin-sensitive pool of GSVs (fractions 8–10) is indicated by a box (peak 1). Peak 2 represents the second-less-responsive recycling endosomes, containing Glut4 (fractions 2–4). (C) Proposed model for the role of Rab10 in Glut4 translocation. Rab10 recruits Rlf to insulin-sensitive GSVs to promote GEF activity on RalA. Activated RalA allows Glut4 translocation through its interactions with the myosin motor and the exocyst.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

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Figure 7: Rab10 and RalA reside in insulin-sensitive GSVs. (A) Subcellular fractionation of 3T3-L1 adipocytes. Indicated fractions were run on SDS–PAGE and immunoblotted with antibodies as specified. HDM, high-density microsomes; LDM, low-density microsomes; N/M, nuclear/mitochondrial; PM, plasma membrane. L.E and D.E. denote light and dark exposures of the RalA blot, respectively. (B) LDM fractions from several independent experiments as in A were pooled to obtain an enriched fraction, which was then run on 14% iodixanol gradient. Fractions 1–11 represent lighter to denser peaks in the gradient. Insulin-sensitive pool of GSVs (fractions 8–10) is indicated by a box (peak 1). Peak 2 represents the second-less-responsive recycling endosomes, containing Glut4 (fractions 2–4). (C) Proposed model for the role of Rab10 in Glut4 translocation. Rab10 recruits Rlf to insulin-sensitive GSVs to promote GEF activity on RalA. Activated RalA allows Glut4 translocation through its interactions with the myosin motor and the exocyst.
Mentions: We performed subcellular fractionation of 3T3-L1 adipocytes to assess the insulin-dependent changes in the cellular localization of RalA and Rab10. As expected, Glut4, IRAP, and RalA were enriched in the plasma membrane fractions upon insulin stimulation, with a concurrent loss of these proteins in the low-density microsome (LDM) fractions. Of interest, Rab10 localization mirrored that of RalA (Figure 7A).

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.

View Article: PubMed Central - PubMed

Affiliation: Life Sciences Institute, University of Michigan Medical School, Ann Arbor, MI 48109.

Show MeSH
Related in: MedlinePlus