Gene replacement strategies validate the use of functional tags on centromeric chromatin and invalidate an essential role for CENP-AK124ub

Cell Reports

Daniele Fachinetti¹  Jennine M. Dawicki-McKenna²  Craig W. Gambogi²  Praveen Kumar Allu²  Glennis A. Logsdon³  Catalina Salinas-Luypaert⁴  Ben E. Black⁴ 

[1] Graduate Program in Biochemistry and Molecular Biophysics, University of Pennsylvania, Philadelphia, PA 19104, USA;Department of Biochemistry and Biophysics, Penn Center for Genome Integrity, Epigenetics Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA;Department of Genome Sciences, University of Washington School of Medicine, Seattle, WA 98195, USA;Institut Curie, PSL University, CNRS, UMR 144, 26 rue d’Ulm, 75005, Paris, France;

关键词: centromere;    genome editing;    CENP-A;    histone variant;    mitosis;    functional protein tags;   

DOI  :  

来源: DOAJ

【 摘 要 】

Summary: Functional tags are ubiquitous in cell biology, and for studies of one chromosomal locus, the centromere, tags have been remarkably useful. The centromere directs chromosome inheritance at cell division. The location of the centromere is defined by a histone H3 variant, CENP-A. The regulation of the chromatin assembly pathway essential for centromere inheritance and function includes posttranslational modification (PTM) of key components, including CENP-A itself. Others have recently called into question the use of functional tags, with the claim that at least two widely used tags obscured the essentiality of one particular PTM, CENP-AK124 ubiquitination (ub). Here, we employ three independent gene replacement strategies that eliminate large, lysine-containing tags to interrogate these claims. Using these approaches, we find no evidence to support an essential function of CENP-AK124ub. Our general methodology will be useful to validate discoveries permitted by powerful functional tagging schemes at the centromere and other cellular locations.

【 授权许可】

Unknown