Full metadata record
| DC Field | Value | Language |
|---|---|---|
| dc.contributor | Department of Health Technology and Informatics | en_US |
| dc.contributor.advisor | Leung, Polly (HTI) | en_US |
| dc.contributor.advisor | Chow, Franklin (HTI) | en_US |
| dc.creator | Gebrelibanos, Daniel Kahase | - |
| dc.identifier.uri | https://theses.lib.polyu.edu.hk/handle/200/14448 | - |
| dc.language | English | en_US |
| dc.publisher | Hong Kong Polytechnic University | en_US |
| dc.rights | All rights reserved | en_US |
| dc.title | Unraveling the regulation mechanism of non-coding small RNA00203 on biofilm formation in Acinetobacter baumannii | en_US |
| dcterms.abstract | Acinetobacter baumannii is the most prevalent and clinically relevant member of the A. baumannii complex and a priority pathogen for the search for new antimicrobials. Biofilm formation and antimicrobial resistance are the key attributes that enable the bacterium to circulate within healthcare institutions. Moreover, biofilm-related infections usually tend to be chronic and refractory to antibiotic treatments and immune responses, complicating patient management. To disarm the biofilm, a deep understanding of its regulatory dynamics is crucial. In our previous work, we demonstrated that a novel regulatory small RNA00203 (sRNA00203) plays a major role in biofilm formation and biofilm-related antimicrobial resistance in A. baumannii ST1894. However, the underlying regulatory networks remain unexplored. Thus, the current study aims to (1) identify genome-wide influenced genes and pathways by sRNA00203; (2) identify and validate the primary target mRNAs of sRNA00203 in biofilm cells, and (3) identify proteins that potentially interact with sRNA00203. | en_US |
| dcterms.abstract | We conducted comparative transcriptomics and proteomics studies on the sRNA00203 deletion mutant and wild-type A. baumannii ST1894 strains to identify genes that are potentially influenced by sRNA00203. The results revealed that a total of 816 genes and 600 proteins were significantly influenced in biofilm cells (absolute value of log2 fold change ≥ 1, q < 0.05). Of the 816 differentially expressed genes in the sRNA00203-deleted strain, 406 were upregulated and 410 were downregulated. Functional annotation showed that major facilitator superfamily efflux pump, type IV pili(T4P), quorum sensing, and iron acquisition systems were repressed. In contrast, genes related to ribosomal proteins, Csu pili, Ade IJK efflux pumps, and porins (CarO type IV, OmpW, and OmpH) were induced. Whole-cell proteome analysis reaffirmed consistent findings, with the exception of the ribosomal proteins and Csu pili. Remarkably, the genes that encode the protein translocase subunit SecE and UDP-3-O-(3-hydroxymyristoyl) glucosamine N-acyltransferase (lpxD) were found to be influenced at the translation level. Phenotypically, the majority of the sRNA-deleted strain displayed a reduction of T4P under transmission electron microscope. | en_US |
| dcterms.abstract | The combined omics studies uncovered the genome-wide potential targets of sRNA00203, but could not distinguish the base-pairing primary targets. To address this gap, we designed a tag-based RNA-RNA interactome capture assay. The 24-nucleotide tag was integrated into sRNA00203 through PCR, and a tagged sRNA00203-expressing strain was constructed using Gibson assembly. Then, using tag antisense sequences as biotin-labelled riboprobes, we were able to retrieve the bound RNA from the tagged sRNA00203-expressing strain and sRNA00203-overexpressing (control) biofilm cell lysate. RNA-seq of the retrieved RNA revealed significant enrichment of 219 genes. Of these genes, genes encoding Class I SAM-dependent methyltransferase (E5A72_RS16120), preprotein translocase subunit SecE, and lipid A hydroxylase LpxO were confirmed to bind at predicted sites with sRNA00203 by electrophoretic mobility shift assay (EMSA). | en_US |
| dcterms.abstract | To further investigate the involvement of chaperone proteins in the regulatory process of sRNA00203, we performed an RNA-protein pulldown assay using A. baumannii biofilm cell lysate. The retrieved proteins were subjected to label-free proteomics. We observed that 32 of the 155 identified proteins were differentially expressed (DEPs) between sRNA00203 and control RNA. Predominantly, proteins such as ribosomal proteins, RNA polymerase subunit, and elongation factors were included in the DEPs, which are annotated as RNA-binding proteins. The known sRNA-modulating chaperone proteins like Hfq and CsrA were not significantly enriched. Instead, UPF0234 protein and chaperone protein DnaK were identified as putative sRNA00203 binding proteins. | en_US |
| dcterms.abstract | Collectively, this study explores regulatory circuits of the novel sRNA00203 for the first time. The sRNA promotes biofilm formation in A. baumannii by inducing the formation of pili, a secretion system, cell wall components, and iron acquisition pathways. Besides, sRNA00203 expression also negatively influences the expression of efflux pumps and porins, which may in turn alter susceptibility of A. baumannii to antimicrobial agents in the biofilm state. The biofilm-related genes such as E5A72_RS16120, secE, and lpxO have been confirmed to interact directly with the sRNA. Furthermore, the regulatory process mediated by the sRNA00203 could potentially be independent of the Hfq chaperone protein. However, new candidate interacting proteins identified in this study could facilitate the sRNA to act on the target mRNAs. | en_US |
| dcterms.extent | xvi, 152 pages : color illustrations | en_US |
| dcterms.isPartOf | PolyU Electronic Theses | en_US |
| dcterms.issued | 2026 | en_US |
| dcterms.educationalLevel | Ph.D. | en_US |
| dcterms.educationalLevel | All Doctorate | en_US |
| dcterms.accessRights | open access | en_US |
Copyright Undertaking
As a bona fide Library user, I declare that:
- I will abide by the rules and legal ordinances governing copyright regarding the use of the Database.
- I will use the Database for the purpose of my research or private study only and not for circulation or further reproduction or any other purpose.
- I agree to indemnify and hold the University harmless from and against any loss, damage, cost, liability or expenses arising from copyright infringement or unauthorized usage.
By downloading any item(s) listed above, you acknowledge that you have read and understood the copyright undertaking as stated above, and agree to be bound by all of its terms.
Please use this identifier to cite or link to this item:
https://theses.lib.polyu.edu.hk/handle/200/14448