Mechanistic and kinetic insights into transcription factor biology via acute protein depletion.

Genetic downregulation of the BCL11A transcription factor (TF) reverses the switch from fetal to adult hemoglobin and is effective in treating β-hemoglobinopathies. Genetic ablation results in a gradual reduction in protein abundance and does not lend itself to the analysis of the immediate consequences of protein loss or the determination of the direct interactors/targets of the protein of interest. We achieved acute degradation of the largely disordered and 'undruggable' BCL11A protein by fusing it with a conditional degradation (degron) tag, FKBP12 F36V , called degradable tags (dTAG). Small molecules then depleted the BCL11A-dTAG through endogenous proteolytic pathways. By integrating acute depletion with nascent transcriptomics and cell cycle separation techniques, we demonstrate the necessity of BCL11A occupancy at the target chromatin for sustained transcriptional repression in erythroid cells. We advocate for expanding the exploration of TF function to include acute depletion, which holds the potential to unveil unprecedented kinetic insights into TF mechanisms of action.