Pathway of intramolecular signal transduction for Chi hotspot control of RecBCD helicase-nuclease

Life-saving repair of broken DNA by homologous recombination requires coordinated enzymatic reactions to prepare it for interaction with intact DNA. The multiple activities of enterobacterial RecBCD helicase-nuclease are coordinated by Chi recombination hotspots (5′ GCTGGTGG 3′) recognized during DNA unwinding. Our previous studies led to a signal transduction model in which the Chi-bound RecC subunit signals RecD, a fast helicase, to stop; RecD then signals RecB, a slower helicase, to swing its nuclease domain into position to cut DNA at Chi and load RecA protein onto the newly generated 3′-ended single-stranded DNA. Here, we report a set of mutants blocked at each step in this pathway. The mutants have helicase and degradative nuclease activities, but most have lost or reduced coordination by Chi and have reduced recombination and DNA repair proficiency. The mutant alterations are widely scattered throughout the three subunits of RecBCD; each alters the intimate contact between two subunits in the crystal or cryoEM structures, supporting the signal transduction model. Similar large conformational changes in other vital enzymes, such as DNA and RNA polymerases, may make RecBCD a paradigm for elucidating their control by the nucleic acid sites they recognize.