Abstract
Septins comprise a conserved family of GTPases important in cytokinesis. These proteins polymerize into filaments from rod-shaped heteromeric septin complexes. Septins interact with one another at two interfaces (NC and G) that alternate within the complex. Here, we show that small mutations at the N terminus greatly enhance the formation of SEPT2 homopolymers. Taking advantage of this mutation to examine polymer formation using SEPT2 alone, we show that both NC and G interfaces are required for filament formation. However, co-expression of wild type SEPT2 with SEPT2 containing mutations at either NC or G interfaces revealed that only the NC mutant suppressed filament formation. NC mutants are able to interact with one another at putative G interfaces, whereas G mutants fail to interact at NC interfaces. In addition, all promiscuous septin pairwise interactions occur at the G interface. These findings suggest that G interface interactions must occur before NC interactions during polymer formation.
Highlights
Septins assemble into oligomers by interactions at “NC” and “G” interfaces by unknown mechanisms
Immunoprecipitation revealed that the GFP-NC mutant bound to FLAG-SEPT2 ⌬15, similar to wild type, whereas there was no appreciable binding with the G mutant (Fig. 4B). These results show that the NC mutant interferes with filament formation by interacting with SEPT2 ⌬15, presumably by its functional G interface, whereas the G mutant cannot bind to wild type SEPT2 ⌬15 by its functional NC interface
N Terminus of SEPT2 Inhibits Homotypic Polymer Formation— Crystallography of septins revealed that in the heterotypic 7-62-2-6-7 complex, SEPT2 interacts with itself at the NC interface when SEPT6 and SEPT7 are present, yet SEPT2 alone formed a dimer through interaction at the G interface [33]
Summary
Septins assemble into oligomers by interactions at “NC” and “G” interfaces by unknown mechanisms. Crystal structure of a bacterially expressed preselected mammalian septin complex consisting of SEPT2, SEPT6, and SEPT7 follow a similar apolar organizational arrangement with 7-6-2-2-6-7 [33]. We recently confirmed this order in HeLa cells and showed that, as in yeast, septins form octamers with SEPT9 at the ends and organized in the sequence 9-7-6-2-2-6-7-9 [34]. We have found a mutation that promotes the self-assembly of SEPT2 into homotypic filaments, thereby simplifying the analysis of how septin assembly occurs In this unique scenario, we have probed the relative affinities of the NC and G interfaces. Septin heterotypic complexes appear to begin with an initial G interaction followed by an NC interaction
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
