Ottilie S, Miller P J, Johnson D I, Creasy C L, Sells M A, Bagrodia S, Forsburg S L, Chernoff J
Molecular Biology and Virology Laboratory, Salk Institute, San Diego, CA 92186, USA.
EMBO J. 1995 Dec 1;14(23):5908-19. doi: 10.1002/j.1460-2075.1995.tb00278.x.
A STE20/p65pak homolog was isolated from fission yeast by PCR. The pak1+ gene encodes a 72 kDa protein containing a putative p21-binding domain near its amino-terminus and a serine/threonine kinase domain near its carboxyl-terminus. The Pak1 protein autophosphorylates on serine residues and preferentially binds to activated Cdc42p both in vitro and in vivo. This binding is mediated through the p21 binding domain on Pak1p and the effector domain on Cdc42p. Overexpression of an inactive mutant form of pak1 gives rise to cells with markedly abnormal shape with mislocalized actin staining. Pak1 overexpression does not, however, suppress lethality associated with cdc42-null cells or the morphologic defeat caused by overexpression of mutant cdc42 alleles. Gene disruption of pak1+ establishes that, like cdc42+, pak1+ function is required for cell viability. In budding yeast, pak1+ expression restores mating function to STE20-null cells and, in fission yeast, overexpression of an inactive form of Pak inhibits mating. These results indicate that the Pak1 protein is likely to be an effector for Cdc42p or a related GTPase, and suggest that Pak1p is involved in the maintenance of cell polarity and in mating.
通过聚合酶链反应（PCR）从裂殖酵母中分离出一种STE20/p65pak同源物。pak1⁺基因编码一种72 kDa的蛋白质，在其氨基末端附近含有一个假定的p21结合结构域，在其羧基末端附近含有一个丝氨酸/苏氨酸激酶结构域。Pak1蛋白在丝氨酸残基上进行自身磷酸化，并且在体外和体内都优先结合活化的Cdc42p。这种结合是通过Pak1p上的p21结合结构域和Cdc42p上的效应结构域介导的。pak1无活性突变形式的过表达会导致细胞形状明显异常，肌动蛋白染色定位错误。然而，Pak1过表达并不能抑制与cdc42缺失细胞相关的致死性或由突变cdc42等位基因过表达引起的形态缺陷。pak1⁺的基因破坏表明，与cdc42⁺一样，pak1⁺功能是细胞活力所必需的。在芽殖酵母中，pak1⁺表达可恢复STE20缺失细胞的交配功能，而在裂殖酵母中，无活性形式的Pak过表达会抑制交配。这些结果表明，Pak1蛋白可能是Cdc42p或相关GTP酶的效应物，并表明Pak1p参与细胞极性的维持和交配过程。
