The LKB1-like Kinase Elm1 Controls Septin Hourglass Assembly and Stability by Regulating Filament Pairing

Abstract

Septins form rod-shaped hetero-oligomeric complexes that assemble into filaments and other higher-order structures, such as rings or hourglasses, at the cell division site in fungal and animal cells [1][2][3][4] to carry out a wide range of functions, including cytokinesis and cell morphogenesis. However, the architecture of septin higher-order assemblies and their control mechanisms, including regulation by conserved kinases [5,6], remain largely unknown. In the budding yeast Saccharomyces cerevisiae, the five mitotic septins (Cdc3, Cdc10, Cdc11, Cdc12, and Shs1) localize to the bud neck and form an hourglass before cytokinesis that acts as a scaffold for proteins involved in multiple processes as well as a membrane-diffusible barrier between the mother and developing bud [7][8][9]. The hourglass is remodeled into a double ring that sandwiches the actomyosin ring at the onset of cytokinesis [10][11][12][13]. How septins are assembled into a highly ordered hourglass structure at the division site [13] is largely unexplored. Here we show that the LKB1-like kinase Elm1, which has been implicated in septin organization [14], cell morphogenesis [15], and mitotic exit [16,17], specifically associates with the septin hourglass during the cell cycle and controls hourglass assembly and stability, especially for the daughter half, by regulating filament pairing and the functionality of its substrate, the septin-binding protein Bni5. This study illustrates how a protein kinase regulates septin architecture at the filament level and suggests that filament pairing is a highly regulated process during septin assembly and remodeling in vivo. ll

Biophysical properties governing septin assembly

“…Recent work investigating the architecture and arrangements of septin filaments in budding yeast via platinum replica EM have shed light on the intricate organizational transition of septin filaments through the cell cycle (Chen et al, 2020; Marquardt et al, 2020). After bud emergence, septin filaments aligned along the mother-bud axis are paired (Ong et al, 2014).…”

“…Unpaired circumferential filaments are critically dependent on the regulators Bud3, a reported RhoGEF, and Bud4, an anillin-like protein. Later in the cell cycle just prior to cytokinesis, septins dramatically rearrange into paired, circumferential filaments sandwiching the actomyosin ring in the middle (Marquardt et al, 2020; Ong et al, 2014). Future extract experiments will enable exploration into how these different regulators combinatorially modulate filament pairing.…”

The kinetic landscape and interplay of protein networks in cytokinesis

“…As YEF473A except TUB1::HPH-pHIS3-mRuby2-TUB1 (Marquardt, et al, 2020) t AflII-digested plasmid pRS305-CHS2-GFP, pRS305-CHS2(I750A)-GFP, pRS305-CHS2(S751A)-GFP, or pRS305-CHS2(N797Q)-GFP was integrated into the LEU2 locus of YEF9905 to generate YEF10361, YEF9921, YEF10092, or YEF10093, respectively. These strains were further selected on SC+5-FOA plate to remove the CHS2 cover plasmid to generate YEF10098, YEF10443, YEF10100, or YEF10101, respectively.…”

“…This study BiLab collection#E2341 pFA6a-His3MX6 (Longtine, et al, 1998) pFA6a-link-yomApple-CaURA3 (Lee, et al, 2013 pFA6a-link-GFPEnvy-SpHis5 (Slubowski, et al, 2015) Addgene Cat#60782 pFA6a-link-yoEGFP-NatMX6 (Marquardt, et al, 2020) BiLab collection#E2375 pFA6a-TRP1 (Longtine, et al, 1998) pG366…”

The Structural Biology of Septins and Their Filaments: An Update

“…The dynamics of septin filament assembly changes radically during the cell cycle, albeit in a highly regulated way, both in time and space (Marquardt et al, 2020). Several binding partners could be key players in this process in which they may act by regulating septin remodeling (Nakahira et al, 2010;Sandrock et al, 2011).…”