Adaptor protein RapZ activates endoribonuclease RNase E by protein-protein interaction to cleave a small regulatory RNA

In Escherichia coli endoribonuclease RNase E initiates RNA degradation and represents a hub for post-transcriptional regulation. The tetrameric adapter protein RapZ targets the small regulatory RNA GlmZ to degradation by RNase E. RapZ binds GlmZ through a domain located at the C-terminus and interacts with the N-terminal catalytic domain of RNase E, promoting GlmZ cleavage in the base-pairing region. When necessary, cleavage of GlmZ is counteracted by the homologous small RNA GlmY, which sequesters RapZ through molecular mimicry. In the current study, we addressed the molecular mechanism employed by RapZ. We show that RapZ mutants impaired in RNA-binding but proficient in binding RNase E, are capable to stimulate GlmZ cleavage by RNase E in vivo and in vitro when provided at increased concentrations. In contrast, a truncated RapZ variant retaining RNA-binding activity but incapable to bind RNase E lacks this activity. In agreement, we find that tetrameric RapZ binds the likewise tetrameric RNase E through direct interaction, independent of RNA. Albeit RapZ is capable to stimulate cleavage of at least one non-cognate RNA by RNase E in vitro, its activity is restricted to GlmZ in vivo as revealed by RNA-seq, suggesting that certain features within the RNA substrate are also required for cleavage. In conclusion, RapZ boosts RNase E activity through interaction with its catalytic domain, representing a novel mechanism of RNase E activation. In contrast, RNA-binding has merely a recruiting role increasing the likelihood that productive RapZ/GlmZ/RNase E complexes may form.