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A Complete Survey of RhoGDI Targets Reveals Novel Interactions with Atypical Small GTPases.

Accepted version
Peer-reviewed

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Authors

Ahmad Mokhtar, Ana Masara Binti 
Ahmed, Samrein BM 
Darling, Nicola J 
Harris, Matthew 

Abstract

There are three RhoGDIs in mammalian cells, which were initially defined as negative regulators of Rho family small GTPases. However, it is now accepted that RhoGDIs not only maintain small GTPases in their inactive GDP-bound form but also act as chaperones for small GTPases, targeting them to specific intracellular membranes and protecting them from degradation. Studies to date with RhoGDIs have usually focused on the interactions between the "typical" or "classical" small GTPases, such as the Rho, Rac, and Cdc42 subfamily members, and either the widely expressed RhoGDI-1 or the hematopoietic-specific RhoGDI-2. Less is known about the third member of the family, RhoGDI-3 and its interacting partners. RhoGDI-3 has a unique N-terminal extension and is found to localize in both the cytoplasm and the Golgi. RhoGDI-3 has been shown to target RhoB and RhoG to endomembranes. In order to facilitate a more thorough understanding of RhoGDI function, we undertook a systematic study to determine all possible Rho family small GTPases that interact with the RhoGDIs. RhoGDI-1 and RhoGDI-2 were found to have relatively restricted activity, mainly binding members of the Rho and Rac subfamilies. RhoGDI-3 displayed wider specificity, interacting with the members of Rho, Rac, and Cdc42 subfamilies but also forming complexes with "atypical" small Rho GTPases such as Wrch2/RhoV, Rnd2, Miro2, and RhoH. Levels of RhoA, RhoB, RhoC, Rac1, RhoH, and Wrch2/RhoV bound to GTP were found to decrease following coexpression with RhoGDI-3, confirming its role as a negative regulator of these small Rho GTPases.

Description

Keywords

Amino Acid Sequence, Cell Membrane, GTP-Binding Proteins, Guanine Nucleotide Dissociation Inhibitors, HEK293 Cells, Humans, Monomeric GTP-Binding Proteins, Protein Binding, rho GTP-Binding Proteins, rho Guanine Nucleotide Dissociation Inhibitor alpha, rho Guanine Nucleotide Dissociation Inhibitor beta, rho Guanine Nucleotide Dissociation Inhibitor gamma, rho-Specific Guanine Nucleotide Dissociation Inhibitors

Journal Title

Biochemistry

Conference Name

Journal ISSN

0006-2960
1520-4995

Volume Title

60

Publisher

American Chemical Society (ACS)

Rights

All rights reserved
Sponsorship
Medical Research Council (MR/J007803/1)
Isaac Newton Trust (1307(b))
BBSRC (1643678)
Medical Research Council (G0700057)
Biotechnology and Biological Sciences Research Council (1643678)
This research was supported by a studentship to AMbAM from the Ministry of Higher Education, Malaysia, a BBSRC Doctoral Training Partnership studentship (BB/JO1454/1) to MH, an Isaac Newton Trust Research Grant Award (13.07(b)) to DO and HRM and an MRC project grant (G0700057) to DO and HRM