Systematic identification of factors mediating accelerated mRNA degradation in response to changes in environmental nitrogen

PLoS Genetics
Darach MillerDavid Gresham

Abstract

Cellular responses to changing environments frequently involve rapid reprogramming of the transcriptome. Regulated changes in mRNA degradation rates can accelerate reprogramming by clearing or stabilizing extant transcripts. Here, we measured mRNA stability using 4-thiouracil labeling in the budding yeast Saccharomyces cerevisiae during a nitrogen upshift and found that 78 mRNAs are subject to destabilization. These transcripts include Nitrogen Catabolite Repression (NCR) and carbon metabolism mRNAs, suggesting that mRNA destabilization is a mechanism for targeted reprogramming of the transcriptome. To explore the molecular basis of destabilization we implemented a SortSeq approach to screen the pooled deletion collection library for trans factors that mediate rapid GAP1 mRNA repression. We combined low-input multiplexed Barcode sequencing with branched-DNA single-molecule mRNA FISH and Fluorescence-activated cell sorting (BFF) to identify the Lsm1-7p/Pat1p complex and general mRNA decay machinery as important for GAP1 mRNA clearance. We also find that the decapping modulators EDC3 and SCD6, translation factor eIF4G2, and the 5' UTR of GAP1 are factors that mediate rapid repression of GAP1 mRNA, suggesting that translational co...Continue Reading

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Citations

Aug 23, 2019·International Journal of Molecular Sciences·Yoshika JanapalaNikolay E Shirokikh
Jan 5, 2020·The Journal of Experimental Biology·Pavan AgrawalLoren L Looger
Jul 3, 2020·Development·Karen LinnemannstönsJulia Christina Gross
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Apr 17, 2021·PLoS Computational Biology·Kaiwen ShiXiaofei Zheng

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Methods Mentioned

BETA
RNAseq
PCA
ESR
fluorescence activated cell sorting
flow cytometry
FACS
PCR
Illumina sequencing

Software Mentioned

SortSeq
pcaMethods
DECOD
tophat2
tools
cutadapt
samtools
R
GNU Makefile
AME

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