CRISPR-Cas-mediated tethering recruits the yeast HMR mating-type locus to the nuclear periphery but fails to silence gene expression

To investigate the relationship between genome structure and function, we have developed a programmable CRISPR-Cas system for nuclear peripheral recruitment in yeast. We benchmarked this system at the HMR and GAL2 loci, both well-characterized model systems for localization to the nuclear periphery....

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Bibliographic Details
Published inbioRxiv
Main Authors Cliff, Emily R, Kirkpatrick, Robin L, Cunningham-Bryant, Daniel, Fernandez, Brianna, Zalatan, Jesse G
Format Paper
LanguageEnglish
Published Cold Spring Harbor Cold Spring Harbor Laboratory Press 01.05.2021
Subjects
CRISPR
Gene expression
Gene silencing
Genomes
Localization
Phenotypes
Protein structure
Reporter gene
Structure-function relationships
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Summary:To investigate the relationship between genome structure and function, we have developed a programmable CRISPR-Cas system for nuclear peripheral recruitment in yeast. We benchmarked this system at the HMR and GAL2 loci, both well-characterized model systems for localization to the nuclear periphery. Using microscopy and gene silencing assays, we demonstrate that CRISPR-Cas-mediated tethering can recruit the HMR locus but does not silence reporter gene expression. A previously reported Gal4-mediated tethering system does silence gene expression, and we demonstrate that the silencing phenotype has an unexpected dependence on the structure of the protein tether. The CRISPR-Cas system was unable to recruit GAL2 to the nuclear periphery. Our results reveal potential challenges for synthetic genome structure perturbations and suggest that distinct functional effects can arise from subtle structural differences in how genes are recruited to the periphery. Competing Interest Statement The authors have declared no competing interest.
DOI:10.1101/2021.04.30.442211