Borg/septin interactions and the assembly of mammalian septin heterodimers, trimers, and filaments.
Septins constitute a family of guanine nucleotide-binding proteins that were first discovered in the yeast Saccharomyces cerevisiae but are also present in many other eukaryotes. In yeast they congregate at the bud neck and are required for cell division. Their function in metazoan cells is uncertain, but they have been implicated in exocytosis and cytokinesis. Septins have been purified from cells as hetero-oligomeric filaments, but their mechanism of assembly is unknown. Further studies have been limited by the difficulty in expressing functional septin proteins in bacteria. We now show that stable, soluble septin heterodimers can be produced by co-expression from bicistronic vectors in bacteria and that the co-expression of three septins results in their assembly into filaments. Pre-assembled dimers and trimers bind guanine nucleotide and show a slow GTPase activity. The assembly of a heterodimer from monomers in vitro is accompanied by GTP hydrolysis. Borg3, a downstream effector of the Cdc42 GTPase, binds specifically to a septin heterodimer composed of Sept6 and Sept7 and to the Sept2/6/7 trimer, but not to septin monomers or to other heterodimers. Septins associate through their C-terminal coiled-coil domains, and Borg3 appears to recognize the interface between these domains in Sept6 and Sept7.
Sheffield
Peter J
PJ
Department of Microbiology and Center for Cell Signaling, University of Virginia School of Medicine, Charlottesville, Virginia 22908, USA.
Oliver
Carey J
CJ
Kremer
Brandon E
BE
Sheng
Sitong
S
Shao
Zhifeng
Z
Macara
Ian G
IG
D000818:Animals; D001665:Binding Sites; D001798:Blood Proteins / Q000737:chemistry* / Q000378:metabolism; D003001:Cloning, Molecular; D003598:Cytoskeletal Proteins; D019281:Dimerization; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D020549:GTP Phosphohydrolase Activators; D020558:GTP Phosphohydrolases / Q000378:metabolism; D020555:GTP-Binding Protein Regulators; D019204:GTP-Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism; D006868:Hydrolysis; D007700:Kinetics; D046911:Macromolecular Substances; D008322:Mammals; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D016601:RNA-Binding Proteins; D012441:Saccharomyces cerevisiae / Q000502:physiology; D020764:cdc42 GTP-Binding Protein / Q000378:metabolism; D020741:rho GTP-Binding Proteins
D001798:Blood Proteins; C422984:CDC42EP2 protein, human; C422985:CDC42EP4 protein, human; D003598:Cytoskeletal Proteins; D020549:GTP Phosphohydrolase Activators; D020555:GTP-Binding Protein Regulators; D046911:Macromolecular Substances; D021122:Protein Subunits; D016601:RNA-Binding Proteins; D006160:Guanosine Triphosphate; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins; D020764:cdc42 GTP-Binding Protein; D020741:rho GTP-Binding Proteins