Colledge of Medicine, School of Medicine and Health Sciences, University of Tsukuba, Tsukuba, Japan.
Department of Molecular Cell Biology, Graduate School of Comprehensive Human Sciences and Faculty of Medicine, University of Tsukuba, Tsukuba, Japan.
PLoS Genet. 2022 Jul 29;18(7):e1010340. doi: 10.1371/journal.pgen.1010340. eCollection 2022 Jul.
Puf5, a Puf-family RNA-binding protein, binds to 3´ untranslated region of target mRNAs and negatively regulates their expression in Saccharomyces cerevisiae. The puf5Δ mutant shows pleiotropic phenotypes including a weakened cell wall, a temperature-sensitive growth, and a shorter lifespan. To further analyze a role of Puf5 in cell growth, we searched for a multicopy suppressor of the temperature-sensitive growth of the puf5Δ mutant in this study. We found that overexpression of CLB2 encoding B-type cyclin suppressed the temperature-sensitive growth of the puf5Δ mutant. The puf5Δ clb2Δ double mutant displayed a severe growth defect, suggesting that Puf5 positively regulates the expression of a redundant factor with Clb2 in cell cycle progression. We found that expression of CLB1 encoding a redundant B-type cyclin was decreased in the puf5Δ mutant, and that this decrease of the CLB1 expression contributed to the growth defect of the puf5Δ clb2Δ double mutant. Since Puf5 is a negative regulator of the gene expression, we hypothesized that Puf5 negatively regulates the expression of a factor that represses CLB1 expression. We found such a repressor, Ixr1, which is an HMGB (High Mobility Group box B) protein. Deletion of IXR1 restored the decreased expression of CLB1 caused by the puf5Δ mutation and suppressed the growth defect of the puf5Δ clb2Δ double mutant. The expression of IXR1 was negatively regulated by Puf5 in an IXR1 3´ UTR-dependent manner. Our results suggest that IXR1 mRNA is a physiologically important target of Puf5, and that Puf5 and Ixr1 contribute to the cell cycle progression through the regulation of the cell cycle-specific expression of CLB1.
Puf5 是 Puf 家族 RNA 结合蛋白，它与靶 mRNA 的 3´非翻译区结合，并负调控其在酿酒酵母中的表达。puf5Δ 突变体表现出多种表型，包括细胞壁减弱、温度敏感生长和寿命缩短。为了进一步分析 Puf5 在细胞生长中的作用，我们在这项研究中搜索了 puf5Δ 突变体的温度敏感生长的多个拷贝抑制子。我们发现，编码 B 类细胞周期蛋白的 CLB2 的过表达抑制了 puf5Δ 突变体的温度敏感生长。puf5Δ clb2Δ 双突变体表现出严重的生长缺陷，表明 Puf5 正向调控细胞周期进程中与 Clb2 冗余的因子的表达。我们发现，编码冗余的 B 类细胞周期蛋白的 CLB1 的表达在 puf5Δ 突变体中降低，并且这种 CLB1 表达的降低导致了 puf5Δ clb2Δ 双突变体的生长缺陷。由于 Puf5 是基因表达的负调控因子，我们假设 Puf5 负调控抑制 CLB1 表达的因子的表达。我们发现了这样一个抑制剂，Ixr1，它是一个 HMGB（高迁移率族 box B）蛋白。删除 IXR1 恢复了 puf5Δ 突变引起的 CLB1 表达降低，并抑制了 puf5Δ clb2Δ 双突变体的生长缺陷。IXR1 的表达受 Puf5 以 IXR1 3´UTR 依赖的方式负调控。我们的结果表明，IXR1mRNA 是 Puf5 的一个生理上重要的靶标，并且 Puf5 和 Ixr1 通过调节 CLB1 的细胞周期特异性表达促进细胞周期进程。
