Cargando…

Unknown Areas of Activity of Human Ribonuclease Dicer: A Putative Deoxyribonuclease Activity

The Dicer ribonuclease plays a crucial role in the biogenesis of small regulatory RNAs (srRNAs) by processing long double-stranded RNAs and single-stranded hairpin RNA precursors into small interfering RNAs (siRNAs) and microRNAs (miRNAs), respectively. Dicer-generated srRNAs can control gene expres...

Descripción completa

Detalles Bibliográficos
Autores principales: Wojnicka, Marta, Szczepanska, Agnieszka, Kurzynska-Kokorniak, Anna
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7144382/
https://www.ncbi.nlm.nih.gov/pubmed/32244942
http://dx.doi.org/10.3390/molecules25061414
_version_ 1783519828332511232
author Wojnicka, Marta
Szczepanska, Agnieszka
Kurzynska-Kokorniak, Anna
author_facet Wojnicka, Marta
Szczepanska, Agnieszka
Kurzynska-Kokorniak, Anna
author_sort Wojnicka, Marta
collection PubMed
description The Dicer ribonuclease plays a crucial role in the biogenesis of small regulatory RNAs (srRNAs) by processing long double-stranded RNAs and single-stranded hairpin RNA precursors into small interfering RNAs (siRNAs) and microRNAs (miRNAs), respectively. Dicer-generated srRNAs can control gene expression by targeting complementary transcripts and repressing their translation or inducing their cleavage. Human Dicer (hDicer) is a multidomain enzyme comprising a putative helicase domain, a DUF283 domain, platform, a PAZ domain, a connector helix, two RNase III domains (RNase IIIa and RNase IIIb) and a dsRNA-binding domain. Specific, ~20-base pair siRNA or miRNA duplexes with 2 nucleotide (nt) 3’-overhangs are generated by Dicer when an RNA substrate is anchored within the platform-PAZ-connector helix (PPC) region. However, increasing number of reports indicate that in the absence of the PAZ domain, binding of RNA substrates can occur by other Dicer domains. Interestingly, truncated variants of Dicer, lacking the PPC region, have been found to display a DNase activity. Inspired by these findings, we investigated how the lack of the PAZ domain, or the entire PPC region, would influence the cleavage activity of hDicer. Using immunopurified 3xFlag-hDicer produced in human cells and its two variants: one lacking the PAZ domain, and the other lacking the entire PPC region, we show that the PAZ domain deletion variants of hDicer are not able to process a pre-miRNA substrate, a dsRNA with 2-nt 3ʹ-overhangs, and a blunt-ended dsRNA. However, the PAZ deletion variants exhibit both RNase and DNase activity on short single-stranded RNA and DNAs, respectively. Collectively, our results indicate that when the PAZ domain is absent, other hDicer domains may contribute to substrate binding and in this case, non-canonical products can be generated.
format Online
Article
Text
id pubmed-7144382
institution National Center for Biotechnology Information
language English
publishDate 2020
publisher MDPI
record_format MEDLINE/PubMed
spelling pubmed-71443822020-04-13 Unknown Areas of Activity of Human Ribonuclease Dicer: A Putative Deoxyribonuclease Activity Wojnicka, Marta Szczepanska, Agnieszka Kurzynska-Kokorniak, Anna Molecules Article The Dicer ribonuclease plays a crucial role in the biogenesis of small regulatory RNAs (srRNAs) by processing long double-stranded RNAs and single-stranded hairpin RNA precursors into small interfering RNAs (siRNAs) and microRNAs (miRNAs), respectively. Dicer-generated srRNAs can control gene expression by targeting complementary transcripts and repressing their translation or inducing their cleavage. Human Dicer (hDicer) is a multidomain enzyme comprising a putative helicase domain, a DUF283 domain, platform, a PAZ domain, a connector helix, two RNase III domains (RNase IIIa and RNase IIIb) and a dsRNA-binding domain. Specific, ~20-base pair siRNA or miRNA duplexes with 2 nucleotide (nt) 3’-overhangs are generated by Dicer when an RNA substrate is anchored within the platform-PAZ-connector helix (PPC) region. However, increasing number of reports indicate that in the absence of the PAZ domain, binding of RNA substrates can occur by other Dicer domains. Interestingly, truncated variants of Dicer, lacking the PPC region, have been found to display a DNase activity. Inspired by these findings, we investigated how the lack of the PAZ domain, or the entire PPC region, would influence the cleavage activity of hDicer. Using immunopurified 3xFlag-hDicer produced in human cells and its two variants: one lacking the PAZ domain, and the other lacking the entire PPC region, we show that the PAZ domain deletion variants of hDicer are not able to process a pre-miRNA substrate, a dsRNA with 2-nt 3ʹ-overhangs, and a blunt-ended dsRNA. However, the PAZ deletion variants exhibit both RNase and DNase activity on short single-stranded RNA and DNAs, respectively. Collectively, our results indicate that when the PAZ domain is absent, other hDicer domains may contribute to substrate binding and in this case, non-canonical products can be generated. MDPI 2020-03-20 /pmc/articles/PMC7144382/ /pubmed/32244942 http://dx.doi.org/10.3390/molecules25061414 Text en © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Wojnicka, Marta
Szczepanska, Agnieszka
Kurzynska-Kokorniak, Anna
Unknown Areas of Activity of Human Ribonuclease Dicer: A Putative Deoxyribonuclease Activity
title Unknown Areas of Activity of Human Ribonuclease Dicer: A Putative Deoxyribonuclease Activity
title_full Unknown Areas of Activity of Human Ribonuclease Dicer: A Putative Deoxyribonuclease Activity
title_fullStr Unknown Areas of Activity of Human Ribonuclease Dicer: A Putative Deoxyribonuclease Activity
title_full_unstemmed Unknown Areas of Activity of Human Ribonuclease Dicer: A Putative Deoxyribonuclease Activity
title_short Unknown Areas of Activity of Human Ribonuclease Dicer: A Putative Deoxyribonuclease Activity
title_sort unknown areas of activity of human ribonuclease dicer: a putative deoxyribonuclease activity
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7144382/
https://www.ncbi.nlm.nih.gov/pubmed/32244942
http://dx.doi.org/10.3390/molecules25061414
work_keys_str_mv AT wojnickamarta unknownareasofactivityofhumanribonucleasediceraputativedeoxyribonucleaseactivity
AT szczepanskaagnieszka unknownareasofactivityofhumanribonucleasediceraputativedeoxyribonucleaseactivity
AT kurzynskakokorniakanna unknownareasofactivityofhumanribonucleasediceraputativedeoxyribonucleaseactivity