Yeast RNA exosome activity is necessary for maintaining cell wall stability through proper protein glycosylation

Abstract

Nuclear RNA exosome is the main 3'→5' RNA degradation and processing complex in eukaryotic cells and its dysregulation therefore impacts gene expression and viability. In this work we show that RNA exosome activity is necessary for maintaining cell wall stability in yeast Saccharomyces cerevisiae. While the essential RNA exosome catalytic subunit Dis3 provides exoribonuclease catalytic activity, the second catalytic subunit Rrp6 has a non-catalytic role in this process. RNA exosome cofactors Rrp47 and Air1/2 are also involved. RNA exosome mutants undergo osmoremedial cell lysis at high temperature or at physiological temperature upon treatment with cell wall stressors. Finally, we show that a defect in protein glycosylation is a major reason for cell wall instability of RNA exosome mutants. Genes encoding enzymes that act in the early steps of the protein glycosylation pathway are downregulated at high temperature in cells lacking Rrp6 protein or Dis3 exoribonuclease activity and overexpression of the essential enzyme Psa1, that catalyzes synthesis of the mannosylation precursor, suppresses temperature sensitivity and aberrant morphology of these cells. Furthermore, this defect is connected to a temperature-dependent increase in accumulation of non-coding RNAs transcribed from loci encoding relevant glycosylation-related genes.

Interplay of the RNA Exosome Complex and RNA-Binding Protein Ssd1 in Maintaining Cell Wall Stability in Yeast

“…We recently identified the nuclear RNA exosome complex as an important yeast cell wall regulator ( 19 ). The absence of the RNA exosome catalytic subunit Rrp6 leads to cell wall instability, which manifests as cell lysis and inviability under conditions of high temperature or other forms of cell wall stress ( 19 ).…”

“…We recently identified the nuclear RNA exosome complex as an important yeast cell wall regulator ( 19 ). The absence of the RNA exosome catalytic subunit Rrp6 leads to cell wall instability, which manifests as cell lysis and inviability under conditions of high temperature or other forms of cell wall stress ( 19 ). The temperature sensitivity of rrp6 Δ mutant cells can be completely suppressed at high temperatures by providing the cells with osmotic support (e.g., by adding 1 M sorbitol to the growth medium), and this suppression was found to be consistent for rrp6 Δ mutants from different genetic backgrounds ( 19 ).…”

“…The absence of the RNA exosome catalytic subunit Rrp6 leads to cell wall instability, which manifests as cell lysis and inviability under conditions of high temperature or other forms of cell wall stress ( 19 ). The temperature sensitivity of rrp6 Δ mutant cells can be completely suppressed at high temperatures by providing the cells with osmotic support (e.g., by adding 1 M sorbitol to the growth medium), and this suppression was found to be consistent for rrp6 Δ mutants from different genetic backgrounds ( 19 ). However, it was striking that the temperature-sensitive phenotype of rrp6 Δ mutant cells was most pronounced in the W303 genetic background, making it the primary choice for studying Rrp6-related phenotypes ( 18 , 21 ).…”

“…The roles of the core subunits and Rrp6 also include stimulation of the exoribonuclease activity of the Dis3 subunit ( 18 ). We have recently shown that the exoribonuclease activity of Dis3 and a noncatalytic function of Rrp6 are involved in maintaining cell wall stability in yeast, which is why RNA exosome mutant cells display temperature-sensitive phenotypes ( 19 ). We now show that the reason that the temperature sensitivity of RNA exosome mutant cells is most pronounced in the W303 genetic background is the presence of the nonfunctional ssd1-d allele.…”

Regulation of the conserved 3′‐5′ exoribonuclease EXOSC10/Rrp6 during cell division, development and cancer

“…Despite playing important and highly diverse cellular roles and evolutionary conservation, budding yeast RRP6 is not essential for mitotic cell division at permissive temperature; however, rrp6 mutant cells display temperature‐sensitive ( ts ) phenotypes (Briggs, Burkard & Butler, 1998; Assenholt et al ., 2008). Recent publications reveal a novel role for Rrp6 in regulating protein glycosylation and maintenance of cell wall stability; these findings elegantly explain why rrp6 mutant cells fail to divide at elevated temperatures (Wang et al ., 2020; Novacic et al ., 2021). Furthermore, rrp6 mutant cells show decreased growth rates as compared to wild‐type cells even at the permissive temperature; a phenotype that is exacerbated on media containing ethanol or glycerol as carbon sources and on synthetic complete medium (Briggs et al ., 1998; Qian et al ., 2012).…”