Kovar David R, Kuhn Jeffrey R, Tichy Andrea L, Pollard Thomas D
Department of Molecular, Cellular, and Developmental Biology, Yale University, New Haven, CT 06520, USA.
J Cell Biol. 2003 Jun 9;161(5):875-87. doi: 10.1083/jcb.200211078.
Cytokinesis in most eukaryotes requires the assembly and contraction of a ring of actin filaments and myosin II. The fission yeast Schizosaccharomyces pombe requires the formin Cdc12p and profilin (Cdc3p) early in the assembly of the contractile ring. The proline-rich formin homology (FH) 1 domain binds profilin, and the FH2 domain binds actin. Expression of a construct consisting of the Cdc12 FH1 and FH2 domains complements a conditional mutant of Cdc12 at the restrictive temperature, but arrests cells at the permissive temperature. Cells overexpressing Cdc12(FH1FH2)p stop growing with excessive actin cables but no contractile rings. Like capping protein, purified Cdc12(FH1FH2)p caps the barbed end of actin filaments, preventing subunit addition and dissociation, inhibits end to end annealing of filaments, and nucleates filaments that grow exclusively from their pointed ends. The maximum yield is one filament pointed end per six formin polypeptides. Profilins that bind both actin and poly-l-proline inhibit nucleation by Cdc12(FH1FH2)p, but polymerization of monomeric actin is faster, because the filaments grow from their barbed ends at the same rate as uncapped filaments. On the other hand, Cdc12(FH1FH2)p blocks annealing even in the presence of profilin. Thus, formins are profilin-gated barbed end capping proteins with the ability to initiate actin filaments from actin monomers bound to profilin. These properties explain why contractile ring assembly requires both formin and profilin and why viability depends on the ability of profilin to bind both actin and poly-l-proline.
在大多数真核生物中，胞质分裂需要肌动蛋白丝和肌球蛋白II环的组装与收缩。裂殖酵母粟酒裂殖酵母在收缩环组装的早期需要formin蛋白Cdc12p和脯氨酰肌动蛋白结合蛋白（Cdc3p）。富含脯氨酸的formin同源（FH）1结构域结合脯氨酰肌动蛋白结合蛋白，而FH2结构域结合肌动蛋白。由Cdc12的FH1和FH2结构域组成的构建体的表达在限制温度下可补充Cdc12的条件突变体，但在允许温度下会使细胞停滞。过表达Cdc12（FH1FH2）p的细胞会因过多的肌动蛋白电缆而停止生长，但没有收缩环。与封端蛋白一样，纯化的Cdc12（FH1FH2）p封端肌动蛋白丝的刺端，阻止亚基的添加和解离，抑制丝的端对端退火，并使仅从其尖端生长的丝成核。最大产量是每六个formin多肽产生一个丝的尖端。同时结合肌动蛋白和聚-L-脯氨酸的脯氨酰肌动蛋白结合蛋白抑制Cdc12（FH1FH2）p的成核作用，但单体肌动蛋白的聚合更快，因为丝从其刺端以与未封端丝相同的速率生长。另一方面，即使在存在脯氨酰肌动蛋白结合蛋白的情况下，Cdc12（FH1FH2）p也会阻止退火。因此，formin蛋白是脯氨酰肌动蛋白结合蛋白门控的刺端封端蛋白，具有从与脯氨酰肌动蛋白结合蛋白结合的肌动蛋白单体引发肌动蛋白丝的能力。这些特性解释了为什么收缩环组装需要formin蛋白和脯氨酰肌动蛋白结合蛋白，以及为什么细胞活力取决于脯氨酰肌动蛋白结合蛋白同时结合肌动蛋白和聚-L-脯氨酸的能力。
