Bni5p, a Septin-Interacting Protein, Is Required for Normal Septin Function and Cytokinesis in Saccharomyces cerevisiae

In the budding yeast Saccharomyces cerevisiae, the Cdc3p, Cdc10p, Cdc11p, Cdc12p, and Sep7p/Shs1p septins assemble early in the cell cycle in a ring that marks the future cytokinetic site. The septins appear to be major structural components of a set of filaments at the mother-bud neck and function as a scaffold for recruiting proteins involved in cytokinesis and other processes. We isolated a novel gene, BNI5, as a dosage suppressor of the cdc12-6 growth defect. Overexpression of BNI5 also suppressed the growth defects of cdc10-1, cdc11-6, and sep7Δ strains. Loss of BNI5 resulted in a cytokinesis defect, as evidenced by the formation of connected cells with shared cytoplasms, and deletion of BNI5 in a cdc3-6, cdc10-1, cdc11-6, cdc12-6, or sep7Δ mutant strain resulted in enhanced defects in septin localization and cytokinesis. Bni5p localizes to the mother-bud neck in a septin-dependent manner shortly after bud emergence and disappears from the neck approximately 2 to 3 min before spindle disassembly. Two-hybrid, in vitro binding, and protein-localization studies suggest that Bni5p interacts with the N-terminal domain of Cdc11p, which also appears to be sufficient for the localization of Cdc11p, its interaction with other septins, and other critical aspects of its function. Our data suggest that the Bni5p-septin interaction is important for septin ring stability and function, which is in turn critical for normal cytokinesis.

在出芽酵母酿酒酵母（Saccharomyces cerevisiae）中，Cdc3p、Cdc10p、Cdc11p、Cdc12p及Sep7p/Shs1p等分隔素（septin）会在细胞周期早期组装成环状结构，标记未来的胞质分裂位点。这类分隔素是母芽颈处一组丝状体的主要结构组分，并可作为支架招募参与胞质分裂及其他生理过程的蛋白质。本研究通过剂量抑制筛选，分离得到了可挽救cdc12-6生长缺陷的新基因BNI5。过表达BNI5同样可抑制cdc10-1、cdc11-6及sep7Δ突变菌株的生长缺陷。BNI5缺失会引发胞质分裂缺陷，具体表现为形成胞质连通的粘连细胞；在cdc3-6、cdc10-1、cdc11-6、cdc12-6或sep7Δ突变菌株中敲除BNI5，则会加剧分隔素定位异常与胞质分裂缺陷。Bni5p会在出芽后不久以分隔素依赖的方式定位于母芽颈，并在纺锤体解体前约2~3分钟从颈区消失。双杂交、体外结合及蛋白质定位实验结果显示，Bni5p可与Cdc11p的N端结构域相互作用；而Cdc11p的N端结构域也足以介导其自身定位、与其他分隔素的相互作用，以及其功能的其他关键环节。本研究数据表明，Bni5p与分隔素的相互作用对分隔素环的稳定性与功能至关重要，而这又直接影响正常胞质分裂过程。