Distinct Ire1-driven transcriptional responses control morphogenesis in Candida albicans

The ability of pathogenic yeast Candida albicans to cause infections in humans depends on morphogenesis, the ability to switch from a spherical yeast to a filamentous hyphal form. Morphogenesis in C. albicans requires Ire1, a transmembrane protein responsible for binding misfolded proteins and initiating the Unfolded Protein Response (UPR). The UPR is a signaling pathway activated in response to the accumulation of misfolded secretory proteins in the endoplasmic reticulum (ER). Ire1 activation in the ER membrane leads to splicing of cytosolic HAC1 mRNA and subsequent UPR target gene upregulation by transcription factor Hac1. In C. albicans, the specific UPR target genes required for morphogenesis are unknown. We utilized RNA-Sequencing to assess the transcriptome of UPR-deficient cells in misfolded protein- and morphogenesis-inducing conditions. Surprisingly, we found little overlap between the datasets, suggesting that a specific Ire1 transcriptional signature drives filamentous growth. Moreover, Hac1 is only partially required for morphogenesis, suggesting a role of Hac1-independent Ire1 function in filamentation. We propose that the cell wall stress response and regulated Ire1-dependent decay (RIDD) are potential Hac1-independent mechanisms connecting Ire1 to morphogenesis in C. albicans. Overall design: To characterize the UPR in C. albicans and its role in morphogenesis, we used a control strain (DAY286) and a strain with dimished expression of UPR activator Ire1 (ire1DX). Strains were grown with the proteotoxic stressor tunicamycin at 30°C for two hours and with morphogenesis-inducing fetal bovine serum at 37°C for four hours. Control samples were untreated and grown at 30°C for two hours. Each condition was performed in triplicate.

致病性酵母白色念珠菌（Candida albicans）引发人类感染的能力，依赖于形态发生——即从球形酵母态转换为丝状菌丝形态的能力。白色念珠菌的形态发生需要Ire1蛋白，这是一种负责结合错误折叠蛋白并启动未折叠蛋白反应（Unfolded Protein Response, UPR）的跨膜蛋白。未折叠蛋白反应是一类响应内质网（endoplasmic reticulum, ER）内错误折叠分泌蛋白积累而激活的信号通路。内质网膜上的Ire1激活会介导胞质HAC1 mRNA的剪接，并通过转录因子Hac1上调UPR靶基因的表达。在白色念珠菌中，参与形态发生的特异性UPR靶基因目前仍未明确。我们借助RNA测序（RNA-Sequencing）技术，评估了错误折叠蛋白诱导与形态发生诱导培养条件下UPR缺陷细胞的转录组特征。出乎意料的是，两类数据集的重叠度极低，这提示存在一类特异性的Ire1转录特征驱动丝状生长。此外，Hac1仅部分参与形态发生，提示Ire1存在不依赖Hac1的功能，该功能在菌丝形成中发挥作用。

我们提出，细胞壁应激反应与调控性Ire1依赖降解（regulated Ire1-dependent decay, RIDD）是连接Ire1与白色念珠菌形态发生的潜在不依赖Hac1的机制。

总体实验设计：为表征白色念珠菌的未折叠蛋白反应及其在形态发生中的作用，我们采用了对照菌株（DAY286）以及UPR激活因子Ire1表达降低的菌株（ire1DX）。将菌株分别置于30℃下，用蛋白毒性应激剂衣霉素处理2小时；同时在37℃下，用形态发生诱导物胎牛血清处理4小时。对照样本未经处理，于30℃下培养2小时。每个实验条件均设置三次重复。