Cargando…
Human cytomegalovirus microRNAs are carried by virions and dense bodies and are delivered to target cells
Human cytomegalovirus (HCMV) infection results in the production of virions, dense bodies (DBs) and non-infectious enveloped particles, all of which incorporate proteins and RNAs that can be transferred to host cells. Here, we investigated whether virions and DBs also carry microRNAs (miRNAs) and as...
Autores principales: | , , , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Microbiology Society
2017
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5656795/ https://www.ncbi.nlm.nih.gov/pubmed/28589873 http://dx.doi.org/10.1099/jgv.0.000736 |
_version_ | 1783273764837916672 |
---|---|
author | Mohammad, Abdul-Aleem Costa, Helena Landázuri, Natalia Lui, Weng-Onn Hultenby, Kjell Rahbar, Afsar Yaiw, Koon-Chu Söderberg-Nauclér, Cecilia |
author_facet | Mohammad, Abdul-Aleem Costa, Helena Landázuri, Natalia Lui, Weng-Onn Hultenby, Kjell Rahbar, Afsar Yaiw, Koon-Chu Söderberg-Nauclér, Cecilia |
author_sort | Mohammad, Abdul-Aleem |
collection | PubMed |
description | Human cytomegalovirus (HCMV) infection results in the production of virions, dense bodies (DBs) and non-infectious enveloped particles, all of which incorporate proteins and RNAs that can be transferred to host cells. Here, we investigated whether virions and DBs also carry microRNAs (miRNAs) and assessed their delivery and functionality in cells. Human lung fibroblasts (MRC-5) were infected with the HCMV strain AD169, and conditioned cell culture medium was collected and centrifuged. The pellets were treated with RNase-ONE, and the virions and DBs were purified with a potassium tartrate–glycerol gradient and dialysed. The virions and DBs were incubated with micrococcal nuclease, DNA and RNA were extracted and then analysed with TaqMan PCR assays, while the proteins were examined with Western blots. To assess the delivery of miRNAs to cells and their functionality, virions and DBs were irradiated with UV light. The purity of the virions and DBs was confirmed by typical morphology, the presence of the structural protein pp65 and the HCMV genome, the ability to infect MRC-5 cells and the absence of the host genome. RNA analysis revealed the presence of 14 HCMV-encoded miRNAs (UL22A-5p, US25-1-5p, UL22A-3p, US5-2-3p, UL112-3p, US25-2-3p, US25-2-5p, US33-3p, US5-1, UL36-5p, US4-5p, UL36-3p, UL70-5p and US25-1-3p), HCMV immediate-early mRNA and long non-coding RNA2.7, moreover, two host-encoded miRNAs (hsa-miR-218-5p and hsa-miR-21-5p) and beta-2-microglobulin RNA. UV-irradiated virions and DBs delivered viral miRNAs (US25-1-5p and UL112-3p) to the host cells, and miR-US25-1-5p was functional in a luciferase reporter assay. We conclude that virions and DBs carry miRNAs that are biologically functional and can be delivered to cells, which may affect cellular processes. |
format | Online Article Text |
id | pubmed-5656795 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2017 |
publisher | Microbiology Society |
record_format | MEDLINE/PubMed |
spelling | pubmed-56567952017-11-14 Human cytomegalovirus microRNAs are carried by virions and dense bodies and are delivered to target cells Mohammad, Abdul-Aleem Costa, Helena Landázuri, Natalia Lui, Weng-Onn Hultenby, Kjell Rahbar, Afsar Yaiw, Koon-Chu Söderberg-Nauclér, Cecilia J Gen Virol Research Article Human cytomegalovirus (HCMV) infection results in the production of virions, dense bodies (DBs) and non-infectious enveloped particles, all of which incorporate proteins and RNAs that can be transferred to host cells. Here, we investigated whether virions and DBs also carry microRNAs (miRNAs) and assessed their delivery and functionality in cells. Human lung fibroblasts (MRC-5) were infected with the HCMV strain AD169, and conditioned cell culture medium was collected and centrifuged. The pellets were treated with RNase-ONE, and the virions and DBs were purified with a potassium tartrate–glycerol gradient and dialysed. The virions and DBs were incubated with micrococcal nuclease, DNA and RNA were extracted and then analysed with TaqMan PCR assays, while the proteins were examined with Western blots. To assess the delivery of miRNAs to cells and their functionality, virions and DBs were irradiated with UV light. The purity of the virions and DBs was confirmed by typical morphology, the presence of the structural protein pp65 and the HCMV genome, the ability to infect MRC-5 cells and the absence of the host genome. RNA analysis revealed the presence of 14 HCMV-encoded miRNAs (UL22A-5p, US25-1-5p, UL22A-3p, US5-2-3p, UL112-3p, US25-2-3p, US25-2-5p, US33-3p, US5-1, UL36-5p, US4-5p, UL36-3p, UL70-5p and US25-1-3p), HCMV immediate-early mRNA and long non-coding RNA2.7, moreover, two host-encoded miRNAs (hsa-miR-218-5p and hsa-miR-21-5p) and beta-2-microglobulin RNA. UV-irradiated virions and DBs delivered viral miRNAs (US25-1-5p and UL112-3p) to the host cells, and miR-US25-1-5p was functional in a luciferase reporter assay. We conclude that virions and DBs carry miRNAs that are biologically functional and can be delivered to cells, which may affect cellular processes. Microbiology Society 2017-05 2017-06-05 /pmc/articles/PMC5656795/ /pubmed/28589873 http://dx.doi.org/10.1099/jgv.0.000736 Text en © 2017 The Authors http://creativecommons.org/licenses/by/4.0/ This is an open access article under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution and reproduction in any medium, provided the original author and source are credited. |
spellingShingle | Research Article Mohammad, Abdul-Aleem Costa, Helena Landázuri, Natalia Lui, Weng-Onn Hultenby, Kjell Rahbar, Afsar Yaiw, Koon-Chu Söderberg-Nauclér, Cecilia Human cytomegalovirus microRNAs are carried by virions and dense bodies and are delivered to target cells |
title | Human cytomegalovirus microRNAs are carried by virions and dense bodies and are delivered to target cells |
title_full | Human cytomegalovirus microRNAs are carried by virions and dense bodies and are delivered to target cells |
title_fullStr | Human cytomegalovirus microRNAs are carried by virions and dense bodies and are delivered to target cells |
title_full_unstemmed | Human cytomegalovirus microRNAs are carried by virions and dense bodies and are delivered to target cells |
title_short | Human cytomegalovirus microRNAs are carried by virions and dense bodies and are delivered to target cells |
title_sort | human cytomegalovirus micrornas are carried by virions and dense bodies and are delivered to target cells |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5656795/ https://www.ncbi.nlm.nih.gov/pubmed/28589873 http://dx.doi.org/10.1099/jgv.0.000736 |
work_keys_str_mv | AT mohammadabdulaleem humancytomegalovirusmicrornasarecarriedbyvirionsanddensebodiesandaredeliveredtotargetcells AT costahelena humancytomegalovirusmicrornasarecarriedbyvirionsanddensebodiesandaredeliveredtotargetcells AT landazurinatalia humancytomegalovirusmicrornasarecarriedbyvirionsanddensebodiesandaredeliveredtotargetcells AT luiwengonn humancytomegalovirusmicrornasarecarriedbyvirionsanddensebodiesandaredeliveredtotargetcells AT hultenbykjell humancytomegalovirusmicrornasarecarriedbyvirionsanddensebodiesandaredeliveredtotargetcells AT rahbarafsar humancytomegalovirusmicrornasarecarriedbyvirionsanddensebodiesandaredeliveredtotargetcells AT yaiwkoonchu humancytomegalovirusmicrornasarecarriedbyvirionsanddensebodiesandaredeliveredtotargetcells AT soderbergnauclercecilia humancytomegalovirusmicrornasarecarriedbyvirionsanddensebodiesandaredeliveredtotargetcells |