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SRX23818567: GSM8123313: arcZ_mutant-pJV300-1; Klebsiella pneumoniae subsp. pneumoniae ATCC 43816; RNA-Seq
1 ILLUMINA (Illumina NovaSeq 6000) run: 9.9M spots, 3G bases, 810.3Mb downloads

External Id: GSM8123313_r1
Submitted by: Shanghai Institute of Immunity and Infection, Chinese Academy of Sciences
Study: RNA interactome profiling in hypervirulent Klebsiella pneumoniae reveals small RNA ArcZ as an inhibitor of mucoviscosity and virulence
show Abstracthide Abstract
Hypervirulent Klebsiella pneumoniae (HvKP) is an emerging human pathogen causing invasive infection in immune-competent hosts. The hypervirulence is strongly linked to the overproduction of hypermucovisous capsule, but the underlining regulatory mechanism of hypermucoviscosity (HMV) has been elusive, especially at the post-transcriptional level mediated by small RNAs (sRNAs). Using a recently developed RNA interactome profiling approach, we have investigated the Hfq-associated sRNA regulatory network and established the first in vivo RNA-RNA interactome in HvKP. Our data reveal numerous interactions between sRNAs and HMV-related mRNAs, and identify a plethora of sRNA that inhibit or promote HMV. One of the strongest repressors of HMV was ArcZ, a conserved sRNA in the Enterobacteriaceae family. We found that ArcZ is activated by the master regulator of catabolite repression Crp, and down-regulates the expression of mlaA encoding an outer-membrane lipoprotein, leading to decreased HMV and virulence attenuation in mice. ArcZ significantly reduced HMV in several carbapenem-resistant and hypervirulent clinical isolates with diverse genetic background, suggesting it is an antisense RNA inhibitor of HMV with therapeutic potential. In summary, our work provides a comprehensive map of the RNA-RNA interaction network of HvKP and identifies ArcZ as a conserved repressor of HMV, providing novel insights into the mechanisms of posttranscriptional regulations of virulence. Overall design: To assess the the effect of expression of ArcZ and MlaA on transcriptome, we have first applied RNA-seq to biological triplicates of K. pneumoniae WT, ?mlaA, ?arcZ, ?arcZ+pZE12-ArcZ and ?arcZ+pZE12-ArcZ-M5. WT and ?arcZ carry the empty vector. Strains were grown in LB media containing IPTG as sRNA expression inducer. Cells at OD600 3 were harvested to construct the strand-specific cDNA library and to be sequenced.
Sample: arcZ_mutant-pJV300-1
SAMN40239619 • SRS20640990 • All experiments • All runs
Library:
Name: GSM8123313
Instrument: Illumina NovaSeq 6000
Strategy: RNA-Seq
Source: TRANSCRIPTOMIC
Selection: cDNA
Layout: PAIRED
Construction protocol: Cells at OD600 3 were harvested by addition of 0.2 volumes of stop mix (95% ethanol, 5% (v/v) phenol) and snap-frozen in liquid nitrogen. Total RNA was used as input material for the RNA sample preparations. mRNA was purified from total RNA using probes to remove rRNA. Fragmentation was carried out using divalent cations under elevated temperature. After fragmentation, the first strand cDNA was synthesized using random hexamer primers. Then the second strand cDNA was synthesized using dUTP, instead of dTTP. The directional library was ready after end repair, A-tailing, adapter ligation, size selection, USER® enzyme digestion, amplification, and purification.
Runs: 1 run, 9.9M spots, 3G bases, 810.3Mb
Run# of Spots# of BasesSizePublished
SRR282062899,935,3713G810.3Mb2024-07-17

ID:
32120197

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