The Anillin-Related Region of Bud4 Is the Major Functional Determinant for Bud4's Function in Septin Organization during Bud Growth and Axial Bud Site Selection in Budding Yeast

Abstract

bThe anillin-related protein Bud4 of Saccharomyces cerevisiae is required for axial bud site selection by linking the axial landmark to the septins, which localize at the mother bud neck. Recent studies indicate that Bud4 plays a role in septin organization during cytokinesis. Here we show that Bud4 is also involved in septin organization during bud growth prior to cytokinesis, as bud4⌬ shs1⌬ cells displayed an elongated bud morphology and defective septin organization at 18°C. Bud4 overexpression also affected septin organization during bud growth in shs1⌬ cells at 30°C. Bud4 was previously thought to associate with the septins via its central region, while the C-terminal anillin-related region was not involved in septin association. Surprisingly, we found that the central region of Bud4 alone targets to the bud neck throughout the cell cycle, unlike full-length Bud4, which localizes to the bud neck only during G 2 /M phase. We identified the anillin-related region to be a second targeting domain that cooperates with the central region for proper septin association. In addition, the anillin-related region could largely mediate Bud4's function in septin organization during bud growth and bud site selection. We show that this region interacts with the C terminus of Bud3 and the two segments depend on each other for association with the septins. Moreover, like the bud4⌬ mutant, the bud3⌬ mutant genetically interacts with shs1⌬ and cdc12-6 mutants in septin organization, suggesting that Bud4 and Bud3 may cooperate in septin organization during bud growth. These observations provide new insights into the interaction of Bud4 with the septins and Bud3.

Coordinate action of distinct sequence elements localizes checkpoint kinase Hsl1 to the septin collar at the bud neck inSaccharomyces cerevisiae

“…(2010) and confirmed here, both Gin4 and Kcc4 have functional KA1 domains at their extreme C-termini, and it has been shown that Gin4 contributes to regulating the lipid distribution in the inner leaflet of the PM in a way that affects the efficiency of cytokinesis ( Roelants et al ., 2015 ). Similarly, the septin-binding proteins Bud4 ( Kang et al ., 2013 ; Wu et al ., 2015 ) and Boi1 (and its paralogue Boi2; Hallett et al ., 2002 ), which are necessary for proper septin organization for bud emergence, have phosphoinositide-binding pleckstrin homology domains at their extreme C-termini ( Yu et al ., 2004 ), and PtdIns4,5P 2 is another lipid that appears to be enriched at the bud neck ( Garrenton et al ., 2010 ). Moreover, septin filament assembly is robustly stimulated in vitro by the presence of PtdIns4,5P 2 in membranes ( Bertin et al ., 2010 ; Bridges et al ., 2014 ); in the cell, the septin filaments in the collar are tightly apposed to the PM at the bud neck, and this association is required for their function ( Bezanilla et al ., 2015 ; Finnigan et al ., 2015b ).…”

Septins and Generation of Asymmetries in Fungal Cells

“…At cytokinesis, septins split into a double ring, so the mother and the daughter cells each inherit one half of the original septin ring. These septin rings then act as a scaffold for landmark proteins such as Bud4, a fungal anillin-like protein (65, 66, 99, 125). The association between Bud4 and septins is not as straightforward as septins simply tethering Bud4, but instead there is clearly a conserved reciprocal relationship.…”

“…The association between Bud4 and septins is not as straightforward as septins simply tethering Bud4, but instead there is clearly a conserved reciprocal relationship. In S. cerevisiae , Bud4 is required for normal septin organization and the splitting of the septin ring; the very process Bud4 itself relies on for segregation to each cell after division (32, 65, 66, 99, 123, 125). The synergistic links between Bud4 and septins are conserved across a variety of fungi including S. pombe , where the Bud4 homolog, Mid2 acts to stabilize the septin rings during cytokinesis similar to that in budding yeast (6).…”

Anillin‐related protein Mid1 regulates timely formation of the contractile ring in the fission yeast Schizosaccharomyces japonicus

“…; Wu et al . ). In the filamentous fungus Neurospora crassa , the Bud4 homologue is required for the process of septum formation by interacting with the Rho family small GTPases (Justa‐Schuch et al .…”

“…BUD4 is one of them. This gene product controls bud site selection by interacting with the bud neck filaments consisted of septin in the budding yeast Saccharomyces cerevisiae (Sanders & Herskowitz 1996;Gladfelter et al 2005;Wu et al 2015). In the filamentous fungus Neurospora crassa, the Bud4 homologue is required for the process of septum formation by interacting with the Rho family small GTPases (Justa-Schuch et al 2010;Delgado-Alvarez et al 2014).…”