Pledger, E. S. | bioRxiv Science

bioRxiv2025

Translational Downregulation of 5' TOP mRNAs During T Cell Exhaustion

T cell exhaustion is a dysfunctional state that arises during chronic infections and cancer, characterized by impaired effector functions and sustained expression of inhibitory receptors. While transcriptional, epigenetic, and metabolic rewiring have been well documented in exhausted T cells, a comprehensive understanding of how translation is regulated in this state remains incomplete. To address this gap, we performed ribosome profiling and RNA sequencing on exhausted human CD8+ T cells to globally assess translational control. Our analyses reveal a marked repression of 5 terminal oligopyrimidine (TOP) mRNAs during exhaustion. Unexpectedly, we demonstrate that this translational repression occurs despite evidence of elevated mTOR activity in exhausted T cells. These findings uncover a previously unknown layer of translational control in exhausted T cells.

molecular biology↗

bioRxiv2019

CUT&Tag for efficient epigenomic profiling of small samples and single cells

Many chromatin features play critical roles in regulating gene expression. A complete understanding of gene regulation will require the mapping of specific chromatin features in small samples of cells at high resolution. Here we describe Cleavage Under Targets and Tagmentation (CUT&Tag), an enzyme-tethering strategy that provides efficient high-resolution sequencing libraries for profiling diverse chromatin components. In CUT&Tag, a chromatin protein is bound in situ by a specific antibody, which then tethers a protein A-Tn5 transposase fusion protein. Activation of the transposase efficiently generates fragment libraries with high resolution and exceptionally low background. All steps from live cells to sequencing-ready libraries can be performed in a single tube on the benchtop or a microwell in a high-throughput pipeline, and the entire procedure can be performed in one day. We demonstrate the utility of CUT&Tag by profiling histone modifications, RNA Polymerase II and transcription factors on low cell numbers and single cells.

genomics↗