Structural and Mechanistic Insights into the Regulation of the Fundamental Rho Regulator RhoGDI by Lysine Acetylation

Rho proteins are small GTP/GDP-binding proteins primarily involved in cytoskeleton regulation. Their GTP/GDP cycle is often tightly connected to a membrane/cytosol cycle regulated by the Rho guanine nucleotide dissociation inhibitor (RhoGDI). RhoGDI has been regarded as a housekeeping regulator essential to control homeostasis of Rho proteins. Recent proteomic screens showed that RhoGDI is extensively lysine-acetylated. Here, we present the first comprehensive structural and mechanistic study to show how RhoGDI function is regulated by lysine acetylation. We discover that lysine acetylation impairs Rho protein binding and increases guanine nucleotide exchange factor-catalyzed nucleotide exchange on RhoA, these two functions being prerequisites to constitute a bona fide GDI displacement factor. RhoGDI acetylation interferes with Rho signaling, resulting in alteration of cellular filamentous actin. Finally, we discover that RhoGDI is endogenously acetylated in mammalian cells, and we identify CBP, p300, and pCAF as RhoGDI-acetyltransferases and Sirt2 and HDAC6 as specific deacetylases, showing the biological significance of this post-translational modification

Nucleotide-dependent farnesyl switch orchestrates polymerization and membrane binding of human guanylate-binding protein 1

Dynamin-like proteins (DLPs) mediate various membrane fusion and fission processes within the cell, which often require the polymerization of DLPs. An IFN-inducible family of DLPs, the guanylate-binding proteins (GBPs), is involved in antimicrobial and antiviral responses within the cell. Human guanylate-binding protein 1 (hGBP1), the founding member of GBPs, is also engaged in the regulation of cell adhesion and migration. Here, we show how the GTPase cycle of farnesylated hGBP1 (hGBP1F) regulates its self-assembly and membrane interaction. Using vesicles of various sizes as a lipid bilayer model, we show GTP-dependent membrane binding of hGBP1F. In addition, we demonstrate nucleotide-dependent tethering ability of hGBP1F. Furthermore, we report nucleotide-dependent polymerization of hGBP1F, which competes with membrane binding of the protein. Our results show that hGBP1F acts as a nucleotide-controlled molecular switch by modulating the accessibility of its farnesyl moiety, which does not require any supportive proteins