Department of Microbiology and Immunology, Witebsky Center for Microbial Pathogenesis and Immunology, Jacobs School of Medicine and Biomedical Sciences, University at Buffalo, Buffalo, New York, USA.
mBio. 2023 Apr 25;14(2):e0019623. doi: 10.1128/mbio.00196-23. Epub 2023 Apr 5.
In response to the host environment, the human pathogen Cryptococcus neoformans must rapidly reprogram its translatome from one which promotes growth to one which is responsive to host stress. In this study, we investigate the two events which comprise translatome reprogramming: the removal of abundant, pro-growth mRNAs from the translating pool, and the regulated entry of stress-responsive mRNAs into the translating pool. Removal of pro-growth mRNAs from the translating pool is controlled primarily by two regulatory mechanisms, repression of translation initiation via Gcn2, and decay mediated by Ccr4. We determined that translatome reprogramming in response to oxidative stress requires both Gcn2 and Ccr4, whereas the response to temperature requires only Ccr4. Additionally, we assessed ribosome collision in response to host-relevant stress and found that collided ribosomes accumulated during temperature stress but not during oxidative stress. The phosphorylation of eIF2α that occurred as a result of translational stress led us to investigate the induction of the integrated stress response (ISR). We found that eIF2α phosphorylation varied in response to the type and magnitude of stress, yet all tested conditions induced translation of the ISR transcription factor Gcn4. However, Gcn4 translation did not necessarily result in canonical Gcn4-dependent transcription. Finally, we define the ISR regulon in response to oxidative stress. In conclusion, this study begins to reveal the translational regulation in response to host-relevant stressors in an environmental fungus which is capable of adapting to the environment inside the human host. Cryptococcus neoformans is a human pathogen capable of causing devastating infections. It must rapidly adapt to changing environments as it leaves its niche in the soil and enters the human lung. Previous work has demonstrated a need to reprogram gene expression at the level of translation to promote stress adaptation. In this work, we investigate the contributions and interplay of the major mechanisms that regulate entry of new mRNAs into the pool (translation initiation) and the clearance of unneeded mRNAs from the pool (mRNA decay). One result of this reprogramming is the induction of the integrated stress response (ISR) regulon. Surprisingly, all stresses tested led to the production of the ISR transcription factor Gcn4, but not necessarily to transcription of ISR target genes. Furthermore, stresses result in differential levels of ribosome collisions, but these are not necessarily predictive of initiation repression as has been suggested in the model yeast.
针对宿主环境，人类病原体新型隐球菌必须迅速将其翻译组从促进生长的翻译组重新编程为对宿主应激反应的翻译组。在这项研究中，我们研究了组成翻译组重编程的两个事件：从翻译池去除丰富的促生长 mRNA，以及应激响应 mRNA 有调控地进入翻译池。翻译组从翻译池中的重新编程主要由两种调节机制控制，通过 Gcn2 抑制翻译起始，以及 Ccr4 介导的衰减。我们确定，氧化应激响应中的翻译组重编程需要 Gcn2 和 Ccr4，而温度响应仅需要 Ccr4。此外，我们评估了宿主相关应激下的核糖体碰撞，并发现核糖体碰撞在温度应激时积累，但在氧化应激时不积累。翻译应激导致的 eIF2α 磷酸化促使我们研究整合应激反应 (ISR) 的诱导。我们发现，eIF2α 磷酸化因应激的类型和强度而异，但所有测试条件均诱导 ISR 转录因子 Gcn4 的翻译。然而，Gcn4 翻译并不一定导致典型的 Gcn4 依赖性转录。最后，我们定义了氧化应激下的 ISR 调控组。总之，这项研究开始揭示环境真菌对宿主相关应激因子的翻译调控，这种真菌能够适应人类宿主内的环境。新型隐球菌是一种能够引起毁灭性感染的人类病原体。它必须迅速适应环境变化，因为它离开土壤中的栖息地并进入人类肺部。先前的工作已经证明需要在翻译水平上重新编程基因表达以促进应激适应。在这项工作中，我们研究了调节新 mRNA 进入翻译池的主要机制（翻译起始）和从翻译池清除不需要的 mRNA 的相互作用和相互作用（mRNA 衰减）。这种重编程的一个结果是诱导了整合应激反应 (ISR) 调控组。令人惊讶的是，所有测试的应激都导致了 ISR 转录因子 Gcn4 的产生，但不一定导致 ISR 靶基因的转录。此外，应激导致核糖体碰撞的水平不同，但这些与模型酵母中提出的起始抑制不一定相关。
