Single mRNP analysis by super-resolution microscopy and fluorescence correlation spectroscopy reveals that small mRNP granules represent mRNA singletons

Mateu-Regué, Àngels; Christiansen, Jan; Bagger, Frederik Otzen; Hellriegel, Christian; Nielsen, Finn Cilius

doi:10.1101/558098

Cited by 1 publication

(1 citation statement)

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“…This targets the protein for proteasomal degradation via the ubiquitin pathway [62]. The fusion of this pest sequence to the C-terminal of yeGFP was previously shown to reduce the half-life of yeGFP from 7 hrs to 30 mins [63]. The GFP expression cassettes were genomically integrated at the LEU2 locus and transcriptionally expressed by a galactose inducible GAL1 promoter.…”

Section: Estimating Single Cell Proteasomal Decay Ratesmentioning

confidence: 99%

Evaluating single-cell variability in proteasomal decay

Das,

Singh,

Shah

2023

Preprint

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Gene expression is a stochastic process that leads to variability in mRNA and protein abundances even within an isogenic population of cells grown in the same environment. This variation, often called gene-expression noise, has typically been attributed to transcriptional and translational processes while ignoring the contributions of protein decay variability across cells. Here we estimate the single-cell protein decay rates of two degron GFPs in Saccharomyces cerevisiae using time-lapse microscopy. We find substantial cell-to-cell variability in the decay rates of the degron GFPs. We evaluate cellular features that explain the variability in the proteasomal decay and find that the amount of 20s catalytic beta subunit of the proteasome marginally explains the observed variability in the degron GFP half-lives. We propose alternate hypotheses that might explain the observed variability in the decay of the two degron GFPs. Overall, our study highlights the importance of studying the kinetics of the decay process at single-cell resolution and that decay rates vary at the single-cell level, and that the decay process is stochastic. A complex model of decay dynamics must be included when modeling stochastic gene expression to estimate gene expression noise.

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Section: Estimating Single Cell Proteasomal Decay Ratesmentioning

confidence: 99%