Abstract
The mechanism, physiological relevance and evolutionary implication of selective RNA processing and stabilization (SRPS) remain elusive. Here we report the genome-wide maps of transcriptional start sites (TSs) and post-transcriptional processed sites (PSs) for Clostridium cellulolyticum. The PS-associated genes are preferably associated with subunits of heteromultimeric protein complexes, and the intergenic PSs (iPSs) are enriched in operons exhibiting highly skewed transcript-abundance landscape. Stem-loop structures associated with those iPSs located at 3′ termini of highly transcribed genes exhibit folding free energy negatively correlated with transcript-abundance ratio of flanking genes. In the cellulosome-encoding cip-cel operon, iPSs and stem-loops precisely regulate structure and abundance of the subunit-encoding transcripts processed from a primary polycistronic RNA, quantitatively specifying cellulosome stoichiometry. Moreover, cellulosome evolution is shaped by the number, position and biophysical nature of TSs, iPSs and stem-loops. Our findings unveil a genome-wide RNA-encoded strategy controlling in vivo stoichiometry of protein complexes.
Highlights
The mechanism, physiological relevance and evolutionary implication of selective RNA processing and stabilization (SRPS) remain elusive
To resolve the potential conflict between the equimolar stoichiometry of transcripts within an operon and the non-equimolar stoichiometry of subunits frequently necessitated, one strategy employed by the cell is selective RNA processing and stabilization (SRPS)[4], where the primary mRNA transcribed as an operon is processed by nucleases into segments first and variation in stability among the segments contribute to differential gene expression[4]
C. cellulolyticum ATCC 35319 genome harbours a large number of glycosylhydrolase genes, and its ‘Cellulose Degradome’ consists of a ‘core’ set of 48 CAZymes required for degrading cellulose-containing substrates and an ‘accessory’ set of 76 CAZymes required for specific noncellulose substrates[26]
Summary
The mechanism, physiological relevance and evolutionary implication of selective RNA processing and stabilization (SRPS) remain elusive. In the cellulosomeencoding cip-cel operon, iPSs and stem-loops precisely regulate structure and abundance of the subunit-encoding transcripts processed from a primary polycistronic RNA, quantitatively specifying cellulosome stoichiometry. In the 12-gene cip-cel operon that encodes cellulosomal enzymes, in vivo and in vitro analyses of the primary and processed transcripts validated the predicted TSs and iPSs and unraveled a mechanism where a combination of five iPSs and four stem-loops precisely determine the stoichiometry of cellulosome in vivo. This mechanism is phylogenetically conserved and drives cellulosome evolution by specifying the structure and abundance of cellulosomal-subunit-encoding transcripts via varying the number, positioning and biophysical nature of TSs, iPSs and stem-loops. Our findings unveil a genome-wide, RNA-encoded mechanism linking sequences and in vivo stoichiometry of protein complexes, and have general implications in designing and engineering protein complex assembly
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