D'Agostino, Federico
(2023)
Characterisation of RNase Y in H. pylori: a non-essential enzyme involved in the processing of cag-PAI non coding
RNA 1 (CncR1) sRNA, [Dissertation thesis], Alma Mater Studiorum Università di Bologna.
Dottorato di ricerca in
Biologia cellulare e molecolare, 35 Ciclo. DOI 10.48676/unibo/amsdottorato/10887.
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Abstract
The importance of Helicobacter pylori as a human pathogen is underlined by the plethora of diseases it is responsible for. The capacity of H. pylori to adapt to the restricted host-associated environment andto evade the host immune response largely depends on a streamlined signalling network. The peculiar H. pylori small genome size combined with its paucity of transcriptional regulators highlights the relevance of post-transcriptional regulatory mechanisms as small non-coding RNAs (sRNAs). However, among the 8 RNases represented in H. pylori genome, a regulator guiding sRNAs metabolism is still not well studied.
We investigated for the first time the physiological role in H. pylori G27 strain of the RNase Y enzyme.
In the first line of research we provide a comprehensive characterization of the RNase Y activity by analysing its genomic organization and the factors that orchestrate its expression. Then, based on bioinformatic prediction models, we depict the most relevant determinants of RNase Y function, demonstrating a correlation of both structure and domain organization with orthologues represented in Gram-positive bacteria. To unveil the post-transcriptional regulatory effect exerted by the RNase Y, we compared the transcriptome of an RNase Y knock-out mutant to the parental wild type strain by RNA-seq approach.
In the second line of research we characterized the activity of this single strand specific endoribonuclease on cag-PAI non coding RNA 1 (CncR1) sRNA. We found that deletion or inactivation of RNase Y led to the accumulation of a 3’-extended CncR1 (CncR1-L) transcript over time. Moreover, beneath its increased half-life, CncR1-L resembled a CncR1 inactive phenotype.
Finally, we focused on the characterization of the in vivo interactome of CncR1. We set up a preliminary MS2-affinity purification coupled with RNA-sequencing (MAPS) approach and we evaluated the enrichment of specific targets, demonstrating the suitability of the technique in the H. pylori G27 strain.
Abstract
The importance of Helicobacter pylori as a human pathogen is underlined by the plethora of diseases it is responsible for. The capacity of H. pylori to adapt to the restricted host-associated environment andto evade the host immune response largely depends on a streamlined signalling network. The peculiar H. pylori small genome size combined with its paucity of transcriptional regulators highlights the relevance of post-transcriptional regulatory mechanisms as small non-coding RNAs (sRNAs). However, among the 8 RNases represented in H. pylori genome, a regulator guiding sRNAs metabolism is still not well studied.
We investigated for the first time the physiological role in H. pylori G27 strain of the RNase Y enzyme.
In the first line of research we provide a comprehensive characterization of the RNase Y activity by analysing its genomic organization and the factors that orchestrate its expression. Then, based on bioinformatic prediction models, we depict the most relevant determinants of RNase Y function, demonstrating a correlation of both structure and domain organization with orthologues represented in Gram-positive bacteria. To unveil the post-transcriptional regulatory effect exerted by the RNase Y, we compared the transcriptome of an RNase Y knock-out mutant to the parental wild type strain by RNA-seq approach.
In the second line of research we characterized the activity of this single strand specific endoribonuclease on cag-PAI non coding RNA 1 (CncR1) sRNA. We found that deletion or inactivation of RNase Y led to the accumulation of a 3’-extended CncR1 (CncR1-L) transcript over time. Moreover, beneath its increased half-life, CncR1-L resembled a CncR1 inactive phenotype.
Finally, we focused on the characterization of the in vivo interactome of CncR1. We set up a preliminary MS2-affinity purification coupled with RNA-sequencing (MAPS) approach and we evaluated the enrichment of specific targets, demonstrating the suitability of the technique in the H. pylori G27 strain.
Tipologia del documento
Tesi di dottorato
Autore
D'Agostino, Federico
Supervisore
Co-supervisore
Dottorato di ricerca
Ciclo
35
Coordinatore
Settore disciplinare
Settore concorsuale
Parole chiave
RNase Y, sRNA, Helicobacter pylori, CncR1, small non-coding RNAs
URN:NBN
DOI
10.48676/unibo/amsdottorato/10887
Data di discussione
19 Giugno 2023
URI
Altri metadati
Tipologia del documento
Tesi di dottorato
Autore
D'Agostino, Federico
Supervisore
Co-supervisore
Dottorato di ricerca
Ciclo
35
Coordinatore
Settore disciplinare
Settore concorsuale
Parole chiave
RNase Y, sRNA, Helicobacter pylori, CncR1, small non-coding RNAs
URN:NBN
DOI
10.48676/unibo/amsdottorato/10887
Data di discussione
19 Giugno 2023
URI
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