title: The Rho guanine nucleotide exchange factor Lsc homo-oligomerizes and is negatively regulated through domains in its carboxyl terminus that are absent in novel splenic isoforms.
abstract: Rho GTPases control fundamental cellular processes, including cytoskeletal reorganization and transcription. Rho guanine nucleotide exchange factors (GEFs) compose a large (>65) and diverse family of related proteins that activate Rho GTPases. Lsc/p115-RhoGEF is a Rho-specific GEF required for normal B and T lymphocyte function. Despite its essential role in lymphocytes, Lsc/p115-RhoGEF signaling in vivo is not well understood. To define Lsc/p115-RhoGEF signaling pathways in vivo, we set out to identify proteins that interact with regulatory regions of Lsc. The 146-amino acid C terminus of Lsc contains a predicted coiled-coil domain, and we demonstrated that deletion of this C terminus confers a gain of function in vivo. Surprisingly, a yeast two-hybrid screen for proteins that interact with this regulatory C terminus isolated a larger C-terminal fragment of Lsc itself. Co-immunoprecipitation experiments in mammalian cells demonstrated that Lsc specifically homo-oligomerizes and that the coiled-coil domain in the C terminus is required for homo-oligomerization. Mutagenesis experiments revealed that homo-oligomerization and negative regulation are distinct functions of the C terminus. Two novel isoforms of Lsc found in the spleen lack portions of this C terminus, including the coiled-coil domain. Importantly, the C termini of both isoforms confer a gain of function and eliminate homo-oligomerization. These results define two important features of Lsc signaling. First, Lsc homo-oligomerizes and is negatively regulated through domains in its C terminus; and second, functionally distinct isoforms of Lsc lacking these domains are present in the spleen.
authors.affiliation: Department of Medicine, Weill Medical College of Cornell University, New York, New York 10021, USA.
authors.forename: Thomas M
authors.initials: TM
authors.lastname: Eisenhaure
authors.forename: Sanjeev A
authors.initials: SA
authors.lastname: Francis
authors.forename: L David
authors.initials: LD
authors.lastname: Willison
authors.forename: Shaun R
authors.initials: SR
authors.lastname: Coughlin
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Lerner
mesh_terms: D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D020662:Guanine Nucleotide Exchange Factors; D007536:Isomerism; D008322:Mammals; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D016296:Mutagenesis; D017434:Protein Structure, Tertiary; D011518:Proto-Oncogene Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D064067:Rho Guanine Nucleotide Exchange Factors; D013154:Spleen / Q000378:metabolism*; D020798:Two-Hybrid System Techniques; D020741:rho GTP-Binding Proteins / Q000378:metabolism
chemical_list: C577231:Arhgef1 protein, mouse; D020662:Guanine Nucleotide Exchange Factors; D011518:Proto-Oncogene Proteins; D064067:Rho Guanine Nucleotide Exchange Factors; D020741:rho GTP-Binding Proteins