Differential effects of depletion of ARL1 and ARFRP1 on membrane trafficking between the trans-Golgi network and endosomes.
Nishimoto-Morita K
et al.
J Biol Chem 2009 Apr;284(16)10583-10592
Nishimoto-Morita K, Shin HW, Mitsuhashi H, Kitamura M, Zhang Q, Johannes L, Nakayama K.
J Biol Chem 2009 Apr;284(16)10583-10592
Abstract: ARFRP1 and ARL1, which are both ARF-like small GTPases, are mammalian orthologs of yeast Arl3p and Arl1p, respectively. In yeast, Arl3p targeted to trans-Golgi network (TGN) membranes activates Arl1p, and the activated Arl1p in turn recruits a GRIP domain-containing protein; this complex regulates retrograde transport to the TGN and anterograde transport from the TGN. In the present study, using RNA interference-mediated knockdown of ARFRP1 and ARL1, we have examined whether the orthologs of Arl3p-Arl1p-GRIP story serve similar functions in mammalian cells. However, we have unexpectedly found differential roles of ARL1 and ARFRP1. Specifically, ARL1 and ARFRP1 regulate retrograde transport of Shiga toxin to the TGN and anterograde transport of VSVG from the TGN, respectively. Furthermore, we have obtained evidence suggesting that a SNARE complex containing Vti1a, syntaxin 6, and syntaxin 16 is involved in Shiga toxin transport downstream of ARL1.
Characterization of a GTPase-activating protein that stimulates GTP hydrolysis by both ADP-ribosylation factor (ARF) and ARF-like proteins. Comparison to the ARD1 gap domain.
Ding M
et al.
J Biol Chem 1996 Sep;271(39)24005-24009
Ding M, Vitale N, Tsai SC, Adamik R, Moss J, Vaughan M.
J Biol Chem 1996 Sep;271(39)24005-24009
Abstract: ADP-ribosylation factors (ARFs) are approximately20-kDa guanine nucleotide-binding proteins that participate in vesicular transport in the Golgi and other intracellular compartments and stimulate cholera toxin ADP-ribosyltransferase activity. Both GTP binding and hydrolysis are necessary for its physiological functions, although purified mammalian ARF lacks detectable GTPase activity. An ARF GTPase-activating protein (GAP) was purified >15,000-fold from rat spleen cytosol using (NH4)2SO4 precipitation and chromatography on Ultrogel AcA 34, DEAE-Sephacel, heparin-Sepharose, hydroxylapatite, and Ultrogel AcA 44. In fractions ( approximately100-kDa proteins) from Ultrogel AcA 44, a major protein band of approximately50 kDa on SDS-polyacrylamide gel electrophoresis correlated with GAP activity, consistent with it being a homodimer, thus differing from an ARF GAP purified from rat liver (Makler, V., Cukierman, E., Rotman, M., Admon, A., and Cassel, D. (1995) J. Biol. Chem. 270, 5232-5237). Purified spleen GAP accelerated hydrolysis of GTP bound to recombinant ARF1, ARF3, ARF5, and ARF6; no effect of NH2-terminal myristoylation was observed. ARF GAP also activated GTP hydrolysis by ARL1, which is 56% identical in amino acid sequence to ARF1, but lacks ARF activity. ARD1 is a 64-kDa guanine nucleotide-binding protein that contains an 18-kDa ARF domain at its carboxyl terminus; the ARF domain lacks the amino-terminal alpha-helix found in native ARF and hence is similar to the amino-terminal truncated mutant Delta13ARF1. Both the ARF domain of ARD1 and Delta13ARF1 were poor substrates for ARF GAP. The non-ARF1 domain of ARD1 enhanced the GTPase activity of the ARF domain, but not that of the ARF proteins and Delta13ARF1, i.e. it lacks the relatively broad substrate specificity exhibited by ARF GAP.
