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Massive expression of cysteine-containing proteins is cytotoxic, manifesting as abnormal morphology in yeast cells

ABSTRACT: The enhanced green fluorescent protein (EGFP) is considered to be a harmless protein because the critical expression level that causes growth defects is higher than that of other proteins. Here we found that overexpression of EGFP, but not a glycolytic protein Gpm1, triggered the cell elongation phenotype in the budding yeast Saccharomyces cerevisiae. By the morphological analysis of the cell overexpressing each of five fluorescent proteins harboring different properties, we revealed that cysteine content was associated with the cell elongation phenotype. We further confirmed this finding by mutational analysis of an EGFP with additional cysteines, a cysteine-containing glycolytic enzyme Tpi1, and a cysteine-less glycolytic enzyme Gpm1. The abnormal cell morphology triggered by overexpression of EGFP was also observed in the fission yeast Schizosaccharomyces pombe. Overexpression of cysteine-containing protein was toxic, especially at high-temperature, while the toxicity could be modulated by additional protein characteristics. The morphological abnormality seemed to be triggered by the S-S bond-triggered protein aggregation, and by the interaction with a chaperone Ssa1 and the heat shock transcription factor Hsf1. Evolutionary analysis of duplicated genes showed that cysteine toxicity may be an evolutionary bias to exclude cysteine from highly expressed proteins. The overexpression of cysteine-less moxGFP, the least toxic protein revealed in this study, would be a good model system to understand the physiological state of protein burden triggered by ultimate overexpression of harmless proteins.

ORGANISM(S): Saccharomyces cerevisiae

PROVIDER: GSE178244 | GEO | 2022/03/31

REPOSITORIES: GEO

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Project description:Aim. Hyperthyroidism reduces the oxidative capacity of skeletal muscle (SKM) and promotes development of a glycolytic phenotype by altering the expression of metabolic proteins. Our previous results showed that smoothelin-like protein 1 (SMTNL1), a transcriptional coregulator, promotes insulin sensitivity in SKM. Our aim was to elucidate the role of SMTNL1 in SKM under physiological and pathological 3,3′,5-Triiodo-L-thyronine (T3) concentrations. Methods. Human hyper- and euthyroid SKM biopsies were used for microarray analysis and phospho-kinase arrays. Physiological and hyperthyroid concentrations of T3 were applied on differentiated C2C12 cells upon SMTNL1 overexpression to assess the activity and expression level of the element of insulin signaling. Real-time metabolic measurements of oxygen consumption rate and extracellular acidification rate were measured. Results. Expression of genes related to energy production, nucleic acid- and lipid metabolism were changed significantly in hyperthyroid samples. SMTNL1 expression decreased in the hyperthyroid samples and in T3 treated C2C12 cells. SMTNL1 regulated muscle cell differentiation and compensated for the glycolytic shift in SKM triggered by pathological T3 exposure. Moreover, it selectively regulated TR expression at the transcriptional level. T3 excess triggered the development of insulin resistance while overexpression of SMTNL1 induced insulin sensitivity through the inhibition of JNK activity and hampered the non-genomic effects of T3 by decreasing the activity of ERK1/2 through PKC. SMTNL1 overexpression induced GLUT4 expression in hyperthyroid model to restore the normal responsiveness of glucose transport to insulin. SMTNL1 regulated glucose phosphorylation and balances glycolysis and glycogen synthesis via the downregulation of hexokinase II. T3 overload strongly increased the rate of lactate production, while SMTNL1 overexpression antagonizes the T3 effects. Conclusion. These lines of evidence suggest that SMTNL1 potentially prevents hyperthyroidism-induced changes in SKM and it holds great promise as a novel therapeutic target. Smoothelin-like Protein 1 in Hyperthyroidism

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Massive expression of cysteine-containing proteins is cytotoxic, manifesting as abnormal morphology in yeast cells

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