Biology subjects

Dickson, B. M.

Publications and source records attributed to Dickson, B. M..

3 recordsLinked to original sources

bioRxiv2023

Analysis of histone antibody specificity directly in sequencing data using siQ-ChIP

We previously developed sans spike-in quantitative chromatin immunoprecipitation sequencing (siQ-ChIP), a technique that introduces an absolute quantitative scale to ChIP-seq data without reliance on spike-in normalization approaches. The physical model of siQ-ChIP predicted that the IP step of ChIP would produce a classical binding isotherm when antibody or epitope was titrated. Here, we define experimental conditions in which this titration is observable for antibodies that recognize modified states of histone proteins. We show that minimally sequenced points along an isotherm can reveal differential binding specificities that are associated with on- and off-target epitope interactions. This work demonstrates that the interpretation of histone post-translational modification distribution from ChIP-seq data has a dependence on antibody concentration. Collectively, these studies introduce a simplified and reproducible experimental method to generate quantitative ChIP-seq data without spike-in normalization and demonstrate that histone antibody specificity can be analyzed directly in ChIP-seq experiments.

molecular biology↗

bioRxiv2022

Ketolysis is a metabolic driver of CD8+ T cell effector function through histone acetylation

Environmental nutrient availability influences T cell metabolism, impacting T cell function and shaping immune outcomes. However, the metabolic pathways critical for optimal T cell responses remain poorly understood. Here, we identify ketone bodies (KBs) - including {beta}-hydroxybutyrate ({beta}OHB) and acetoacetate (AcAc) - as essential fuels supporting CD8+ T cell metabolism and effector function. Ketolysis is an intrinsic feature of highly functional CD8+ T effector (Teff) cells and {beta}OHB directly increases CD8+ Teff cell IFN-{gamma} production and cytolytic activity. Using metabolic tracers, we establish that CD8+ Teff cells preferentially use KBs over glucose to fuel the tricarboxylic acid (TCA) cycle in vitro and in vivo. KBs directly boost the respiratory capacity of CD8+ T cells and TCA cycle-dependent metabolic pathways that fuel T cell growth. Mechanistically, we find that {beta}OHB is a major substrate for acetyl-CoA production in CD8+ T cells and regulates effector responses through effects on histone acetylation. Together, our results identify cell-intrinsic ketolysis as a metabolic and epigenetic driver of optimal CD8+ T cell effector responses. One Sentence summaryKetone bodies promote CD8+ T cell metabolism and effector function through regulation of epigenetic programming

immunology↗

bioRxiv2022

Theoretical and practical refinements of sans spike-in quantitative ChIP-seq with application to p300/CBP inhibition

Previously, we introduced an absolute and physical quantitative scale for chromatin immunoprecipitation followed by sequencing. The scale itself was determined directly from measurements routinely made on sequencing samples without additional reagents or spike-ins. We called this approach sans spike-in quantitative ChIP, or siQ-ChIP. In this paper we extend those results in several ways. First, we simplified the calculations defining the quantitative scale. Second, we highlight the normalization constraint implied by the quantitative scale and introduce a new scheme for generating tracks for siQ-ChIP. We next introduce some whole-genome analyses that are unique to siQ-ChIP which allow us, for example, to project the IP mass onto the genome to evaluate how much of any genomic interval was captured in the IP. We apply these analyses to p300/CBP inhibition and demonstrate that response to inhibition is a function of genomic architecture. In particular, active transcription start sites are only weakly perturbed by p300/CBP inhibition while enhancers are strongly perturbed. Similar observations have been reported in the literature, but without a quantitative scale, those observations have been misinterpreted. We discuss how the siQ-ChIP approach precludes such misinterpretations, which stem from the widespread community practice of treating unquantified and unnormalized ChIP-seq tracks as though they are quantitative.

biophysics↗