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Proteomic and transcriptomic experiments reveal an essential role of RNA degradosome complexes in shaping the transcriptome of Mycobacterium tuberculosis.
journal contributionposted on 2020-01-06, 17:41 authored by Przemysław Płociński, Maria Macios, Joanna Houghton, Emilia Niemiec, Renata Płocińska, Anna Brzostek, Marcin Słomka, Jarosław Dziadek, Douglas Young, Andrzej Dziembowski
The phenotypic adjustments of Mycobacterium tuberculosis are commonly inferred from the analysis of transcript abundance. While mechanisms of transcriptional regulation have been extensively analysed in mycobacteria, little is known about mechanisms that shape the transcriptome by regulating RNA decay rates. The aim of the present study is to identify the core components of the RNA degradosome of M. tuberculosis and to analyse their function in RNA metabolism. Using an approach involving cross-linking to 4-thiouridine-labelled RNA, we mapped the mycobacterial RNA-bound proteome and identified degradosome-related enzymes polynucleotide phosphorylase (PNPase), ATP-dependent RNA helicase (RhlE), ribonuclease E (RNase E) and ribonuclease J (RNase J) as major components. We then carried out affinity purification of eGFP-tagged recombinant constructs to identify protein-protein interactions. This identified further interactions with cold-shock proteins and novel KH-domain proteins. Engineering and transcriptional profiling of strains with a reduced level of expression of core degradosome ribonucleases provided evidence of important pleiotropic roles of the enzymes in mycobacterial RNA metabolism highlighting their potential vulnerability as drug targets.
DEAD-box RNA HelicasesEndoribonucleasesEscherichia coliEscherichia coli ProteinsMultienzyme ComplexesMycobacterium smegmatisMycobacterium tuberculosisPolyribonucleotide NucleotidyltransferaseProteomeProteomicsRNARNA HelicasesRNA StabilityRNA, BacterialRibonuclease IIIRibonucleasesThiouridineTranscriptomeYoung FC001222Developmental Biology05 Environmental Sciences06 Biological Sciences08 Information and Computing Sciences