Sengar A S, Markley N A, Marini N J, Young D
Department of Medical Biochemistry, University of Calgary Health Science Centre, Alberta, Canada.
Mol Cell Biol. 1997 Jul;17(7):3508-19. doi: 10.1128/MCB.17.7.3508.
We have identified a Schizosaccharomyces pombe gene, mkh1, that encodes a MEK kinase (MEKK) homolog. The coding region of mkh1 is contained within a single exon encoding a 1,116-amino-acid protein. The putative catalytic domain of Mkh1 is 54% identical to the catalytic domain of S. cerevisiae Bck1, the most closely related protein. Deletion of mkh1 did not significantly affect cell growth or division under standard conditions. However, mkh1delta cell growth was inhibited by high KCl or NaCl concentrations. mkh1delta cells required a longer time to reenter the cell cycle after prolonged stationary-phase arrest. Also, mkh1delta cells exhibited a round cell shape, while overexpression of Mkh1 resulted in an elongated cell shape. mkh1delta cells exhibited a more dramatic phenotype when grown in nutrient-limiting conditions at high temperature or in hyperosmotic medium. In such conditions, completion of cytokinesis was inhibited, resulting in the growth of pseudohyphal filaments with multiple septa and nuclei. Also, mkh1delta cells were hypersensitive to beta-glucanase treatment. Together these results suggest that Mkh1 regulates cell morphology, cell wall integrity, salt resistance, cell cycle reentry from stationary-phase arrest, and filamentous growth in response to stress. These phenotypes are essentially identical to those exhibited by cells lacking Pmk1/Spm1, a recently identified mitogen-activated protein kinase. Our evidence suggests that Pmk1/Spm1 acts downstream from Mkh1 in a common pathway. Our results also suggest that Mkh1 and Pck2 act independently to maintain cell wall integrity, cell morphology, and salt resistance but act in opposition to regulate filamentous growth.
我们鉴定出了粟酒裂殖酵母的一个基因mkh1，它编码一种MEK激酶（MEKK）同源物。mkh1的编码区包含在一个单一外显子中，该外显子编码一个1116个氨基酸的蛋白质。Mkh1的推定催化结构域与最密切相关的蛋白质——酿酒酵母Bck1的催化结构域有54%的同一性。在标准条件下，缺失mkh1对细胞生长或分裂没有显著影响。然而，mkh1Δ细胞的生长受到高浓度KCl或NaCl的抑制。mkh1Δ细胞在长时间静止期停滞后重新进入细胞周期需要更长时间。此外，mkh1Δ细胞呈现圆形细胞形态，而Mkh1的过表达导致细胞形态拉长。当在高温或高渗培养基中的营养限制条件下生长时，mkh1Δ细胞表现出更显著的表型。在这种条件下，胞质分裂的完成受到抑制，导致具有多个隔膜和细胞核的假菌丝丝生长。此外，mkh1Δ细胞对β-葡聚糖酶处理高度敏感。这些结果共同表明，Mkh1调节细胞形态、细胞壁完整性、耐盐性、从静止期停滞重新进入细胞周期以及对应激的丝状生长。这些表型与缺乏Pmk1/Spm1（一种最近鉴定的丝裂原活化蛋白激酶）的细胞所表现出的表型基本相同。我们的证据表明，Pmk1/Spm1在一条共同途径中作用于Mkh1的下游。我们的结果还表明，Mkh1和Pck2独立作用以维持细胞壁完整性、细胞形态和耐盐性，但在调节丝状生长方面作用相反。
