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High throughput RNA interference based screen for the identification of genes implicated in listeria monocytogenes infection
Includes bibliographical references.
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| Format: | Thesis |
| Language: | English |
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Department of Medicine
2014
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| _version_ | 1867613373198237696 |
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| access_status_str | Open Access |
| author | Tantoh, Asongwe Lionel Ateh |
| author2 | Brombacher, Frank |
| author_browse | Brombacher, Frank Tantoh, Asongwe Lionel Ateh |
| author_facet | Brombacher, Frank Tantoh, Asongwe Lionel Ateh |
| author_sort | Tantoh, Asongwe Lionel Ateh |
| collection | Thesis |
| description | Includes bibliographical references. |
| format | Thesis |
| id | oai:open.uct.ac.za:11427/9533 |
| institution | University of Cape Town (South Africa) |
| language | eng |
| last_indexed | 2026-06-10T12:35:06.996Z |
| license_str | Not specified — see source repository |
| provenance_str_mv | Harvested via OAI-PMH from UCTD — University of Cape Town Open Access Repository |
| publishDate | 2014 |
| publishDateRange | 2014 |
| publishDateSort | 2014 |
| publisher | Department of Medicine |
| publisherStr | Department of Medicine |
| record_format | dspace |
| source_str | UCTD — University of Cape Town Open Access Repository |
| spelling | oai:open.uct.ac.za:11427/9533 High throughput RNA interference based screen for the identification of genes implicated in listeria monocytogenes infection Tantoh, Asongwe Lionel Ateh Brombacher, Frank Mhlanga, N Emans M Includes bibliographical references. The facultative intracellular pathogen L. monocytogenes has acquired a versatile arsenal of adaptor proteins. It uses these proteins to invade and undermine the host immune surveillance system, as well as to survive and propagate within the host cell. Our understanding of how L. monocytogenes infects cells points towards an interplay between bacterial proteins and human host factors. It has become apparent that this interplay is critical for the establishment and spread of infection. Here we describe a high throughput screening approach to assay for potential human host factors involved in L. monocytogenes infection. In this methodology, we applied a high-density siRNA array comprising of the kinome, phosphatome, ubiquitome and protease encoding human genes that enabled rapid parallel screening for human host factors through reverse transfection into overlaid HeLa cells. The siRNA transfected HeLa cells were exposed to L. monocytogenes expressing the green fluorescent protein. The high content screen comprised a total of 15 replicate screens or 47, 250 individual siRNA experiments. Using a combination of image-based analysis and the data-mining tool of Principal Component Analysis, genes whose silencing visually impaired or enhanced bacteria invasion/proliferation were selected as candidate genes. The 65 strongest genes were subjected to a secondary screening for validation. Among the 15 confirmed hits, we recognized some cellular membrane proteins that might function as cell receptors for bacteria entry and others that may be related to calcium release triggered by bacteria during cell invasion. In addition, we identified a transcription factor bromo adjacent homology domain-containing 1(BAHD1) protein, whose interplay with the bacterial virulence factor LntA has recently been shown to modulate IFN-λ-mediated immune response to control bacterial colonization of the host cell. Also, we identified the inositol polyphosphate-5-phosphatase (INPP5B) that may regulate bacterial internalization by modulating actin dynamics at bacterial internalization sites as shown with oculocerebrorenal syndrome of Lowe (OCRL), whose OCRL gene shares an identical sequence and overlapping enzyme activities with INPP5B. We also identified the large tumor suppressor kinase 2 (LATS2) that may play a potential role in bacterial colonization by modulating the expression of the host E-cad receptor gene. This study represents a significant step toward unveiling the key molecular requirements for host cell invasion and undoubtedly provides an avenue of new therapeutic drugs against L. monocytogenes infection. 2014-11-11T06:57:49Z 2014-11-11T06:57:49Z 2014 Doctoral Thesis Doctoral PhD http://hdl.handle.net/11427/9533 eng application/pdf Department of Medicine Faculty of Health Sciences University of Cape Town |
| spellingShingle | Tantoh, Asongwe Lionel Ateh High throughput RNA interference based screen for the identification of genes implicated in listeria monocytogenes infection |
| thesis_degree_str | Doctoral |
| title | High throughput RNA interference based screen for the identification of genes implicated in listeria monocytogenes infection |
| title_full | High throughput RNA interference based screen for the identification of genes implicated in listeria monocytogenes infection |
| title_fullStr | High throughput RNA interference based screen for the identification of genes implicated in listeria monocytogenes infection |
| title_full_unstemmed | High throughput RNA interference based screen for the identification of genes implicated in listeria monocytogenes infection |
| title_short | High throughput RNA interference based screen for the identification of genes implicated in listeria monocytogenes infection |
| title_sort | high throughput rna interference based screen for the identification of genes implicated in listeria monocytogenes infection |
| url | http://hdl.handle.net/11427/9533 |
| work_keys_str_mv | AT tantohasongwelionelateh highthroughputrnainterferencebasedscreenfortheidentificationofgenesimplicatedinlisteriamonocytogenesinfection |