Proteotoxicity From Aberrant Ribosome Biogenesis Compromises Cell Fitness

Abstract

Proteins are the workhorses of the cell, carrying out much of the structural and enzymatic work of life. Many proteins function as part of greater assemblies, or complexes, with other proteins and/or biomolecules. As the cell grows and divides, it must double its proteome, which requires synthesis and folding of individual proteins, assembly into complexes, and proper subcellular localization. This is repeated for millions of proteins molecules compromising thousands of potential assemblies, making up a rather tricky set of tasks for cells to carry out simultaneously. The very machine that carries out protein synthesis—the ribosome—is among the most challenging complexes to assemble and one of the most abundant. Each ribosome requires ~80 unique proteins and 4 unique RNAs to be stitched together in an assembly line that spans the nucleus to the cytosol. Intriguingly, there exist many extracellular insults and genetic mutations that compromise the integrity of ribosome biogenesis, but that impact different cell types and cell states with a range of severities. This represents an intriguing paradox, as the null expectation would be that ribosomes would be critically important to all cells. In this work, I sought to identify the mechanism by which disrupting the integrity of ribosome assembly compromises cell fitness using budding yeast as a model organism with exceptional chemical and genetic tools. By using these tools in yeast to perturb diverse stages of ribosome assembly one at a time on very short timescales, I was able to study the most proximal consequences. I found that cells experience a collapse of protein folding homeostasis in conditions that give rise to excess newly-synthesized ribosomal proteins relative to what can be assembled. In detailing this response, I found that cells specifically activate the conserved proteostasis restoration response directed by the transcription factor Heat Shock Factor 1 (Hsf1). In doing so, this work identifies the first such endogenous proteins capable of eliciting Hsf1 activation, and I further explored the possibility that the fate of newly-synthesized proteins may more generally be compromised by other proteotoxic conditions that drive Hsf1 activation. This work implicates protein synthesis in general as a risk for the proteome that may be compromised by various insults, and suggests that the ribosome in particular may be troublesome for rapidly-proliferating cells with high ribosome production.