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. 2023 Jan 12;13(1):jkac296.
doi: 10.1093/g3journal/jkac296.

A synthetic genetic array screen for interactions with the RNA helicase DED1 during cell stress in budding yeast

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A synthetic genetic array screen for interactions with the RNA helicase DED1 during cell stress in budding yeast

Sara B Carey et al. G3 (Bethesda). .

Abstract

During cellular stress it is essential for cells to alter their gene expression to adapt and survive. Gene expression is regulated at multiple levels, but translation regulation is both a method for rapid changes to the proteome and, as one of the most energy-intensive cellular processes, a way to efficiently redirect cellular resources during stress conditions. Despite this ideal positioning, many of the specifics of how translation is regulated, positively or negatively, during various types of cellular stress remain poorly understood. To further assess this regulation, we examined the essential translation factor Ded1, an RNA helicase that has been previously shown to play important roles in the translational response to cellular stress. In particular, ded1 mutants display an increased resistance to growth inhibition and translation repression induced by the TOR pathway inhibitor, rapamycin, suggesting that normal stress responses are partially defective in these mutants. To gain further insight into Ded1 translational regulation during stress, synthetic genetic array analysis was conducted in the presence of rapamycin with a ded1 mutant and a library of nonessential genes in Saccharomyces cerevisiae to identify positive and negative genetic interactions in an unbiased manner. Here, we report the results of this screen and subsequent network mapping and Gene Ontology-term analysis. Hundreds of candidate interactions were identified, which fell into expected categories, such as ribosomal proteins and amino acid biosynthesis, as well as unexpected ones, including membrane trafficking, sporulation, and protein glycosylation. Therefore, these results provide several specific directions for further comprehensive studies.

Keywords: helicase; rapamycin; stress; translation; yeast.

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Conflict of interest statement

None declared.

Figures

Fig. 1.
Fig. 1.
Identification of genetic interactions with ded1-ΔCT following rapamycin treatment. a) The synthetic interaction scores with ded1-ΔCT for growth on rapamycin for all genes tested (4,799) are shown in ascending order. The inner dashed blue lines represent 1 SD from the mean score, and the outer dashed red lines represent 1.5 SD. Genes falling outside of these thresholds were considered “hits” and used for further analysis. b) Growth of single (fis1Δ, hsp30Δ, and sbp1Δ) and ded1-ΔCT double mutants for 3 representative hits on rich media (YPD) and rich media plus rapamycin are shown. Five-fold serial dilutions were grown at 30°C for 2 (YPD) or 4 (Rapamycin) days. Synthetic interactions were observed consistent with the screen results.
Fig. 2.
Fig. 2.
Network cluster maps of interacting genes. Network maps were generated using STRING for the synthetic negative/suppressors of ded1-ΔCT (a) and the synthetic positive/enhancers of ded1-ΔCT (b) hits that exceeded the 1.5 SD threshold below and above the mean interaction score, respectively. Thickness of the edges between genes signifies the strength of data support for the interaction. Disconnected nodes/genes are not shown. Clusters were generated via k-means clustering. The identity of each cluster (no. 1–5) is labeled below, and corresponding colors were used in Tables 1 and 2.

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References

    1. Al-Hadid Q, Roy K, Chanfreau G, Clarke SG.. Methylation of yeast ribosomal protein Rpl3 promotes translational elongation fidelity. RNA. 2016;22(4):489–498. - PMC - PubMed
    1. Aryanpur PP, Mittelmeier TM, Bolger TA.. The RNA helicase Ded1 regulates translation and granule formation during multiple phases of cellular stress responses. Mol Cell Biol. 2022;42(1):e0024421. - PMC - PubMed
    1. Aryanpur PP, Regan CA, Collins JM, Mittelmeier TM, Renner DM, Vergara AM, Brown NP, Bolger TA.. Gle1 regulates RNA binding of the DEAD-box helicase Ded1 in its complex role in translation initiation. Mol Cell Biol. 2017;37(21): - PMC - PubMed
    1. Aryanpur PP, Renner DM, Rodela E, Mittelmeier TM, Byrd A, Bolger TA.. The DEAD-box RNA helicase Ded1 has a role in the translational response to TORC1 inhibition. Mol Biol Cell. 2019;30(17):2171–2184. - PMC - PubMed
    1. Baryshnikova A, Costanzo M, Kim Y, Ding H, Koh J, Toufighi K, Youn JY, Ou J, San Luis BJ, Bandyopadhyay S, et al. . Quantitative analysis of fitness and genetic interactions in yeast on a genome scale. Nat Methods. 2010;7(12):1017–1024. - PMC - PubMed

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