HelD is a global transcription factor enhancing gene expression in rapidly growing mycobacteria.

HelD protein, also named HelR (encoded by MSMEG_2174 in Mycobacterium smegmatis), interacts with mycobacterial RNA polymerase (RNAP) and affects rifampicin resistance in Mycobacterium abscessus. Here, we provide data on rifampicin resistance and helD presence in the genomes of other clinically relevant nontuberculous mycobacteria. We show that helD is primarily found in rapidly growing mycobacteria, such as M. smegmatis, where we detected HelD at a subset of promoters that can also associate with CarD and RbpA. Transcriptome analysis of a helD deletion strain revealed that HelD has a dual role in gene regulation: it enhances transcription during exponential growth but reduces it in stationary phase. We show that transcription downregulation in stationary phase is accompanied by decreased amounts of the transcription initiation factors RbpA and CarD. In addition, we characterized HelD-containing RNAP complexes using native cryo-EM. Based on these data, we propose that HelD acts as a general transcription factor that releases stalled RNAP complexes and dissociates RNAP from promoters. HelD demonstrates how a single protein can perform fundamentally different tasks, mediating both antibiotic resistance and efficient transcription, and thereby shaping bacterial adaptability.