TY - JOUR
T1 - Global identification of RsmA/N binding sites in Pseudomonas aeruginosa by in vivo UV CLIP-seq
AU - Chihara, Kotaro
AU - Barquist, Lars
AU - Takasugi, Kenichi
AU - Noda, Naohiro
AU - Tsuneda, Satoshi
N1 - Publisher Copyright:
© 2021 Informa UK Limited, trading as Taylor & Francis Group.
PY - 2021
Y1 - 2021
N2 - Pseudomonas aeruginosa harbours two redundant RNA-binding proteins RsmA/RsmN (RsmA/N), which play a critical role in balancing acute and chronic infections. However, in vivo binding sites on target transcripts and the overall impact on the physiology remains unclear. In this study, we applied in vivo UV crosslinking immunoprecipitation followed by RNA-sequencing (UV CLIP-seq) to detect RsmA/N-binding sites at single-nucleotide resolution and mapped more than 500 binding sites to approximately 400 genes directly bound by RsmA/N in P. aeruginosa. This also verified the ANGGA sequence in apical loops skewed towards 5ʹUTRs as a consensus motif for RsmA/N binding. Genetic analysis combined with CLIP-seq results suggested previously unrecognized RsmA/N targets involved in LPS modification. Moreover, the RsmA/N-titrating RNAs RsmY/RsmZ may be positively regulated by the RsmA/N-mediated translational repression of their upstream regulators, thus providing a possible mechanistic explanation for homoeostasis of the Rsm system. Thus, our study provides a detailed view of RsmA/N-RNA interactions and a resource for further investigation of the pleiotropic effects of RsmA/N on gene expression in P. aeruginosa.
AB - Pseudomonas aeruginosa harbours two redundant RNA-binding proteins RsmA/RsmN (RsmA/N), which play a critical role in balancing acute and chronic infections. However, in vivo binding sites on target transcripts and the overall impact on the physiology remains unclear. In this study, we applied in vivo UV crosslinking immunoprecipitation followed by RNA-sequencing (UV CLIP-seq) to detect RsmA/N-binding sites at single-nucleotide resolution and mapped more than 500 binding sites to approximately 400 genes directly bound by RsmA/N in P. aeruginosa. This also verified the ANGGA sequence in apical loops skewed towards 5ʹUTRs as a consensus motif for RsmA/N binding. Genetic analysis combined with CLIP-seq results suggested previously unrecognized RsmA/N targets involved in LPS modification. Moreover, the RsmA/N-titrating RNAs RsmY/RsmZ may be positively regulated by the RsmA/N-mediated translational repression of their upstream regulators, thus providing a possible mechanistic explanation for homoeostasis of the Rsm system. Thus, our study provides a detailed view of RsmA/N-RNA interactions and a resource for further investigation of the pleiotropic effects of RsmA/N on gene expression in P. aeruginosa.
KW - CLIP-seq
KW - Pseudomonas aeruginosa
KW - RsmA
KW - RsmN
KW - post-transcriptional regulation
UR - http://www.scopus.com/inward/record.url?scp=85105156024&partnerID=8YFLogxK
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U2 - 10.1080/15476286.2021.1917184
DO - 10.1080/15476286.2021.1917184
M3 - Article
C2 - 33866926
AN - SCOPUS:85105156024
SN - 1547-6286
VL - 18
SP - 2401
EP - 2416
JO - RNA Biology
JF - RNA Biology
IS - 12
ER -