Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.601621
Title: Regulation of potassium channels by microRNA : targeting of Kir2.1 and down-regulation of inward rectifier potassium current by miR-212
Author: Goldini , Dana
Awarding Body: Queen's University Belfast
Current Institution: Queen's University Belfast
Date of Award: 2013
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Abstract:
Cardiac remodelling is associated with differential expression of microRNAs (miRNAs ) and down-regulation of inward rectifier K+ current (lKl) , of which Kir2.1 is the predominant molecular component . miR-212, miR-132 and miR-320 have been reported to be pro-hypertrophic miRNAs. Furthermore they are predicted to bind Kir2 .1 3' untranslated region (3'UTR) by miRNA target prediction algorithms, indicating a potential role in IKI down-regulation. The aims of this study were to examine which miRNAs overexpressed in hearts undergoing remodelling, en route to failure, are involved in the downregulation of Kir2.1 and IKI. MiRNAs up- regulated in failing and hypertrophic hearts were cross-referenced to miRNAs predicted to target Kir2 .1 by prediction algorithms . The interaction of the shortlisted miRNAs, miR-132, miR-212 and miR-320, with Kir2.1 3'UTR was investigated by a duallight luciferase assay expressing a luciferase gene with fragments of Kir2.1 3'UTR. Co - transfection of HEK293T cells with a luciferase construct, a Bgalactosidase construct and a plasmid encoding miR-212 (pmiR-212) reduced luciferase activity compared to control. MiR-132 and miR-320 did not show any significant reduction of luciferase activity. The HeLa cell line was chosen as an experimental model to study Kir2.1 expression under the regulation of miR- 212 . HeLa cells were confirmed to express Kir2.1 with a negligible contribution by Kir2 .2 at mRNA and protein levels. Transfection of HeLa with miR-212 or control indicated a strong down-regulation of IKl, confirmed by protein decrease. HL-l cells were characterized for the first time to pass IKI and to express Kir2.1, Kir2.2 and Kir2 .3. However, Kir2.1 expression was apparent l y dependent on the serum used for cell culture . miR-212 was transfected into HL-l expression was very low Kir2.1 compared to Kir2.2 and Kir2 . 3. IKI was not reduced, suggesting that miR- 212 reduces IKI by down - regulating Kir2.1 expression, suggesting miR-212 to be a regulator of IKI in cardiac hypertrophy and heart failure.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.601621  DOI: Not available
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