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Harper, N. W.

Publications and source records attributed to Harper, N. W..

2 recordsLinked to original sources

p53 controls choice between apoptotic and non-apoptotic death following DNA damage

DNA damage can activate apoptotic and non-apoptotic forms of cell death; however, it remains unclear what features dictate which type of cell death is activated. We report that p53 controls the choice between apoptotic and non-apoptotic death following exposure to DNA damage. In contrast to the conventional model, which suggests that p53-deficient cells should be resistant to DNA damage-induced cell death, we find that p53-deficient cells die at high rates following DNA damage, but exclusively using non-apoptotic mechanisms. Our experimental data and computational modeling reveal that non-apoptotic death in p53-deficient cells has not been observed due to use of assays that are either insensitive to cell death, or that specifically score apoptotic cells. Using functional genetic screening - with an analysis that enables computational inference of the drug-induced death rate - we find in p53-deficient cells that DNA damage activates a mitochondrial respiration-dependent form of cell death, called MPT-driven necrosis. Cells deficient for p53 have high basal respiration, which primes MPT-driven necrosis. Finally, using metabolite profiling, we identified mitochondrial activity-dependent metabolic vulnerabilities that can be targeted to potentiate the lethality of DNA damage specifically in p53-deficient cells. Our findings reveal how the dual functions of p53 in regulating mitochondrial activity and the DNA damage response combine to facilitate the choice between apoptotic and non-apoptotic death.

cell biology↗

Simultaneous profiling of multiple chromatin proteins in the same cells

Methods derived from CUT&RUN and CUT&Tag enable genome-wide mapping of the localization of proteins on chromatin from as few as one cell. These and other mapping approaches focus on one protein at a time, preventing direct measurements of colocalization of different chromatin proteins in the same cells and requiring prioritization of targets where samples are limiting. Here we describe multi-CUT&Tag, an adaptation of CUT&Tag that overcomes these hurdles by using antibody-specific barcodes to simultaneously map multiple proteins in the same cells. Highly specific multi-CUT&Tag maps of histone marks and RNA Polymerase II uncovered sites of co-localization in the same cells, active and repressed genes, and candidate cis-regulatory elements. Single-cell multi-CUT&Tag profiling facilitated identification of distinct cell types from a mixed population and characterization of cell type-specific chromatin architecture. In sum, multi-CUT&Tag increases the information content per cell of epigenomic maps, facilitating direct analysis of the interplay of different proteins on chromatin.

genomics↗