Publicação
Functional validation of novel candidate microRNA-like molecules encoded in HIV genomes
| datacite.subject.fos | Departamento de Química e Bioquímica | pt_PT |
| dc.contributor.advisor | Carvalho, Margarida Henriques da Gama, 1972- | |
| dc.contributor.author | Luta, Raquel Alexandra Gaspar | |
| dc.date.accessioned | 2025-04-03T17:08:38Z | |
| dc.date.embargo | 2028-04-10 | |
| dc.date.issued | 2025 | |
| dc.date.submitted | 2024 | |
| dc.description | Tese de mestrado, Bioquímica e Biomedicina, 2025, Universidade de Lisboa, Faculdade de Ciências | pt_PT |
| dc.description.abstract | MicroRNAs (miRNAs) are crucial regulators of gene expression, impacting translation and mRNA stability across several biological processes, including viral infections. Our lab previously identified two viral small non-coding RNAs (sncRNAs) encoded in HIV genomes — one from HIV-1 and one from HIV2 — that can follow the canonical microRNA biogenesis pathway. Prediction of miRNA-mRNA target interactions suggested these sequences might aid viral infection by inhibiting host proteins detrimental to the virus. The work developed in this dissertation aimed to assess the potential of these novel sequences to function like bona fide miRNAs. We conducted functional characterisation of the candidate HIV-derived miRNAs to assess their role in regulating host gene expression and influencing HIV replication. To investigate seed sequence dependency, seed-disruptive mutant constructs of the sncRNAs were created and compared with wild-type counterparts. Both wild-type and mutant miRNAs’ impact on predicted host mRNA targets was evaluated with RT-qPCR and Western blot following transient expression of their precursor transcripts in HEK-293 cells. Additionally, flow cytometry and RT-qPCR on J-lat 6.3 cells assessed HIV reactivation and candidate v-miRNA production, following TNF-α stimulation. HEK-293 RT-qPCR results showed increased mRNA levels of the selected target genes in cells transfected with the mutant constructs, indicating seed dependency in miRNA-mediated repression. Western blot analysis confirmed that wild-type viral sncRNas reduced protein levels, while seed mutants showed partial activity. We demonstrated sncRNA 1_2 expression in latent infected J-lat cells, indicating its potential relevance in HIV infection. However, the effects of the sncRNAs on HIV latency reversal were subtle, making it difficult to draw definitive conclusions. Overall, our findings support the hypothesis that HIV-1 and HIV-2 encode microRNA-like sequences capable of regulating host-virus interactions and provide new insights into v-miRNA-dependent regulatory mechanisms involved in the pathobiology of HIV-1 and HIV-2, highlighting the need for further in-depth studies. | pt_PT |
| dc.identifier.tid | 203946863 | pt |
| dc.identifier.uri | http://hdl.handle.net/10400.5/99992 | |
| dc.language.iso | eng | pt_PT |
| dc.subject | v-microRNAs | pt_PT |
| dc.subject | VIH-1 | pt_PT |
| dc.subject | VIH-2 | pt_PT |
| dc.subject | sequência seed | pt_PT |
| dc.subject | interação vírus-hospedeiro | pt_PT |
| dc.subject | Teses de mestrado - 2025 | pt_PT |
| dc.title | Functional validation of novel candidate microRNA-like molecules encoded in HIV genomes | pt_PT |
| dc.type | master thesis | |
| dspace.entity.type | Publication | |
| rcaap.rights | embargoedAccess | pt_PT |
| rcaap.type | masterThesis | pt_PT |
| thesis.degree.name | Tese de mestrado em Bioquímica e Biomedicina | pt_PT |