Biology Reference
In-Depth Information
Chapter 13
Inhibition of miRNA Maturation by Peptide Nucleic Acids
Concetta Avitabile , Enrica Fabbri , Nicoletta Bianchi , Roberto Gambari ,
and Alessandra Romanelli
Abstract
Molecules able to interfere in miRNA genesis and function are potent tools to unravel maturation and
processing pathways. Antisense oligonucleotides or analogs are actually employed for the inhibition of
miRNA function. Here we illustrate how Peptide Nucleic Acids oligomers targeting pre-miRNA are
exploited to inhibit miRNA maturation.
Key words
Peptide nucleic acids, Pre-miR, miR, Peptide, FACS, RT-PCR
1
Introduction
Since the discovery of the small noncoding RNAs miRNAs as
posttranscriptional regulators of gene expression, enormous efforts
have been made to identify miRNA's biogenesis, targets and func-
tions [
1
,
2
]. MiRNAs are originated from precursors, pre-miRNAs.
In humans a miR loading complex (miRLC) formed by Ago2 and
Dicer has been identifi ed as responsible for the miRNA maturation;
pre-miRs are loaded and processed within this complex by the
nuclease Dicer to generate miRNA duplexes [
3
,
4
]. The duplexes
are loaded onto the Ago proteins and successively unwinded; the
guide strand is retained by Ago, while the passenger strand is
degraded. In an alternative pathway, independent by Dicer, pre-
miRs beginning with 5
A associate to Ago2 in a miRNA
precursor deposit complex (miDPC); Ago2 both cleaves the pre-
cursor and binds the mature miRNA [
5
]. MiRNAs are involved in
several biological processes, including cell proliferation, differentia-
tion and apoptosis; also several miRNAs have been demonstrated to
be associated to pathological conditions [
6
]. In this context, the
inhibition of miRNA function has become a priority task.
Here we describe a strategy for the inhibition of miRNA matu-
ration using Peptide Nucleic Acid oligomers designed to target the
′
U or 5
′
