Emerging role for AS160/TBC1D4 and TBC1D1 in the regulation of GLUT4 traffic

Abstract

Vesicular traffic of the glucose transporter GLUT4 occurs in response to insulin, muscle contraction, and metabolic stimuli that lead to changes in the energy status of the cell. These stimuli are associated with linked kinase cascades that lead to changes in glucose uptake that meet the energy challenges imposed on the highly regulated cell types in insulin-responsive tissues. The need to mechanistically link these kinase-associated stimuli to identifiable intermediates in vesicular traffic has long been known but has been difficult to fulfill. The Rab-GTPase-activating proteins AS160 and TBC1D1 have now emerged as strong candidates to fill this void. Here we review the initial discovery of these proteins as phosphorylated substrates for Akt and the more recent emerging data that indicate that these proteins are substrates for additional kinases that are downstream of contraction and energy status signaling. The mechanism of coupling these phosphorylated proteins to vesicle traffic appears to be dependent on linking to small GTPase of the Rab family. We examine the current state of a hypothesis that suggests that phosphorylation of the Rab-GTPase-activating proteins leads to increased GTP loading of Rab proteins on GLUT4 vesicles and subsequently to increased interaction with Rab effectors that control GLUT4 vesicle translocation.

Fig. 1.

Currently proposed signaling pathways for insulin- and contraction-stimulated glucose transporter 4 (GLUT4) translocation in muscle. Insulin stimulates Akt through 2 distinct upstream mediators, phosphoinositide-dependent protein kinase-1 (PDK1) and mammalian target of rapamycin complex-2 (mTORC2). Activated Akt phosphorylates AS160 and TBC1D1 mainly at Thr⁶⁴² and Thr⁵⁹⁶, respectively. This enhances 14-3-3 binding to these proteins, which is proposed to inhibit Rab-GTPase-activating protein (GAP) activity toward particular Rab isoform(s). Inhibition of GAP promotes conversion of less active GDP-loaded Rab to more active GTP-loaded Rab. The more active GTP-loaded Rab then allows GLUT4 storage vesicles to move to and fuse with the plasma membrane. Contraction through both energy depletion (i.e., an elevated AMP/ATP ratio) and elevated intracellular [Ca²⁺] leads to activation of AMP-activated protein kinase (AMPK) via LKB1 and Ca²⁺/calmodulin-dependent protein kinase kinase (CaMKK), respectively (and possibly other Ca²⁺-regulated protein kinases). These regulators lead to AS160 and TBC1D1 phosphorylation at multiple phosphorylation sites. This is thought to regulate function of these proteins and GLUT4 trafficking by largely uncharacterized mechanism(s). IRS, insulin receptor substrate; PI3K, phosphatidylinositol 3-kinase.

Fig. 2.

Schematic domain structures of human AS160/TBC1D4 and TBC1D1. The amino acid sequences used are National Center for Biotechnology Information (NCBI; gi: 114688046) for human AS160/TBC1D4 and NCBI (gi: 50658061) for human TBC1D1. The 2 phosphotyrosine-binding (PTB) domains and the GAP domains are those predicted by analysis with the Pfam program (http://www.sanger.ac.uk/). Sano et al. (51) originally found that insulin stimulates phosphorylation of AS160 at 5 sites via Akt, and mutation of AS160 in which 4 of phosphorylation sites (Ser³¹⁸, Ser⁵⁸⁸, Thr⁶⁴², Ser⁷⁵¹) had been mutated to alanine (AS160-4P) abolished insulin-stimulated GLUT4 translocation in 3T3-L1 adipocytes. Akt phosphorylates Thr⁵⁹⁶, and AMPK phosphorylates Ser²³⁷ of TBC1D1 in vitro and in intact cells. CBD, calmodulin-binding domain.