Enhanced cleavage of RNA mediated by an interaction between substrates andthe arginine-rich domain of E-coli ribonuclease E

Endonucleolytic cutting by the essential Escherichia coil ribonuclease RNas eE has a central role in both the processing and decay of RNA. Previously, it has been shown that an oligoribonucleotide corresponding in sequence to the single-stranded region at the 5' end of RNAI, the antisense regulator o f ColE1-type plasmid replication, is efficiently cut by RNaseE. Combined wi th the knowledge that alteration of the structure of stem-loops within comp lex RNaseE substrates can either increase or decrease the rate of cleavage, this result has led to the notion that stem-loops do not serve as essentia l recognition motifs for RNaseE, but can affect the rate of cleavage indire ctly by, for example, determining the single-strandedness of the site or it s accessibility. We report here, however, that not all oligoribonucleotides corresponding to RNaseE-cleaved segments of complex substrates are suffici ent to direct efficient RNaseE cleavage. We provide evidence using 9 S RNA, a precursor of 5 S rRNA, that binding of structured regions by the arginin e-rich RNA-binding domain (ARRBD) of RNaseE can be required for efficient c leavage. Binding by the ARRBD appears to counteract the inhibitory effects of sub-optimal cleavage site sequence and overall substrate conformation. F urthermore, combined with the results from recent analyses of E. coli mutan ts in which the ARRBD of RNase E is deleted, our findings suggest that subs trate binding by RNaseE is essential for the normal rapid decay of E. coli mRNA. The simplest interpretation of our results is that the ARRBD recruits RNaseE to structured RNAs, thereby increasing the localised concentration of the N-terminal catalytic domain, which in turn leads to an increase in t he rate of cleavage. (C) 2000 Academic Press.