Park Jong-In, Collinson Emma J, Grant Chris M, Dawes Ian W
Ramaciotti Centre for Gene Function Analysis, School of Biotechnology and Biomolecular Sciences, University of New South Wales, Sydney 2052, Australia.
J Biol Chem. 2005 Jan 28;280(4):2529-35. doi: 10.1074/jbc.M407900200. Epub 2004 Nov 15.
The Ras-cyclic AMP pathway is connected to other nutrient-regulated signaling pathways and mediates the global stress responses of Saccharomyces cerevisiae. Here, we show that Rom2p, the Rho1 GTP/GDP exchange factor, can mediate stress responses and cell growth via the Ras-cAMP pathways. ROM2 was isolated as a suppresser of heat and NaCl sensitivity caused by the lack of the Ras-GTPase activator Ira2p or of cAMP phosphodiesterases. Subsequent analysis of strains with a rom2 deletion showed that Rom2p is essential for resistance to a variety of stresses caused by freeze-thawing, oxidants, cycloheximide, NaCl, or cobalt ions. Stress sensitivity and the growth defect caused by the rom2 deletion could be suppressed by depleting Ras or protein kinase A (PKA) activity or by overexpressing the high affinity cAMP phosphodiesterase Pde2p. In addition, overexpression of ROM2 could not rescue cells lacking the regulatory subunit of PKA, indicating that the Ras-adenylate, cyclase-PKA cascade is essential for Rom2p-mediated stress responses and cell growth. Deletion of IRA2 exacerbated the freeze-thaw sensitivity and growth defect of the rom2 mutant, indicating that Rom2p signaling may control Ras independently of IRA2. Increases in cAMP levels were detected in the rom2 deletion mutants, and these were comparable with the effects of an ira2 mutation. The effects of the deletion of ROM2 on sensitivity to hydrogen peroxide, paraquat, and cobalt ions, but not to caffeine, were reduced when a constitutive allele of RHO1 was introduced on a single copy plasmid. However, the effects of the deletion of ROM2 on sensitivity to diamide and NaCl were exacerbated. Taken together, our data indicate that Rom2p can regulate PKA activity by controlling cAMP levels via the Ras-cAMP pathway and that for those stresses related to oxidative stress, this cross-talk is probably mediated via the Rho1p-activated MAPK pathway.
Ras-环磷酸腺苷（cAMP）信号通路与其他营养物质调节的信号通路相连，并介导酿酒酵母的全局应激反应。在此，我们表明Rho1 GTP/GDP交换因子Rom2p可通过Ras-cAMP信号通路介导应激反应和细胞生长。ROM2是作为因缺乏Ras-GTP酶激活剂Ira2p或cAMP磷酸二酯酶而导致的热敏感性和对氯化钠敏感性的抑制子被分离出来的。随后对rom2缺失菌株的分析表明，Rom2p对于抵抗由冻融、氧化剂、环己酰亚胺、氯化钠或钴离子引起的多种应激至关重要。rom2缺失导致的应激敏感性和生长缺陷可通过降低Ras或蛋白激酶A（PKA）的活性，或通过过表达高亲和力的cAMP磷酸二酯酶Pde2p来抑制。此外，ROM2的过表达无法拯救缺乏PKA调节亚基的细胞，这表明Ras-腺苷酸环化酶-PKA级联反应对于Rom2p介导的应激反应和细胞生长至关重要。IRA2的缺失加剧了rom2突变体的冻融敏感性和生长缺陷，这表明Rom2p信号传导可能独立于IRA2控制Ras。在rom2缺失突变体中检测到cAMP水平升高，这些升高与ira2突变的影响相当。当在单拷贝质粒上引入RHO1的组成型等位基因时，ROM2缺失对过氧化氢、百草枯和钴离子敏感性的影响降低，但对咖啡因的敏感性影响未降低。然而，ROM2缺失对二硫苏糖醇和氯化钠敏感性的影响加剧。综上所述，我们的数据表明，Rom2p可通过Ras-cAMP信号通路控制cAMP水平来调节PKA活性，并且对于那些与氧化应激相关的应激，这种相互作用可能是通过Rho1p激活的丝裂原活化蛋白激酶（MAPK）信号通路介导的。
