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.
Highlights
The ARF/ARL3 family of small GTPases play crucial roles in membrane trafficking and in other cellular processes by interacting with various effector proteins (1, 2)
Dominant-negative Mutants of ARL1 Might Disorganize the trans-Golgi network (TGN)—We previously showed that exogenous expression of an HA-tagged dominant-negative ARFRP1 mutant, ARFRP1(T31N)-HA, in HeLa cells causes redistribution of ARL1 and golgin-245 from the TGN (Fig. 1, A and AЈ, and supplemental Fig. S1) and blocks both exit of vesicular stomatitis virus G protein (VSVG) from the TGN and retrograde transport of Shiga toxin B fragment (StxB) from endosomes to the TGN
We have revealed two important points regarding the roles of ARL1 and ARFRP1 in mammalian cells
Summary
The ARF/ARL3 family of small GTPases play crucial roles in membrane trafficking and in other cellular processes by interacting with various effector proteins (1, 2). Dominant-negative Mutants of ARL1 Might Disorganize the TGN—We previously showed that exogenous expression of an HA-tagged dominant-negative ARFRP1 mutant, ARFRP1(T31N)-HA, in HeLa cells causes redistribution of ARL1 and golgin-245 from the TGN (Fig. 1, A and AЈ, and supplemental Fig. S1) and blocks both exit of VSVG from the TGN (see supplemental Fig. S2) and retrograde transport of StxB from endosomes to the TGN (see supplemental Fig. S3).
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