Khdrbs1 stimulates TMEJ by suppressing poison exon inclusion in Polq mRNA
DNA polymerase Theta-Mediated End-Joining (TMEJ) is a stand-alone mutagenic DNA double-strand break repair pathway that becomes critical when high-fidelity repair is compromised. Although the enzymatic mechanism of TMEJ has been extensively studied in recent years, its regulation remains poorly understood. Here, we identify and characterize the RNA-binding protein Khdrbs1 (also known as Sam68) as a key modulator of TMEJ in mammalian cells. We demonstrate that Khdrbs1 is required for proper expression of Polq, the gene encoding polymerase theta (Polq), the central factor in TMEJ. Loss of Khdrbs1 results in reduced TMEJ at CRISPR-induced DSBs and increased sensitivity to ionizing radiation. Mechanistically, Khdrbs1 promotes correct splicing of Polq mRNA by suppressing the inclusion of a conserved poison exon that introduces a premature termination codon in the transcript. This function depends on the RNA-binding domain of Khdrbs1. Genetic deletion of the poison exon restores Polq expression and rescues TMEJ activity in Khdrbs1-depleted cells. Together, these findings establish alternative splicing as a direct mechanism controlling TMEJ capacity and reveal a conserved regulatory module that tunes mutagenic DSB repair through modulation of Polq abundance.
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