Structure, evolution and function of the bi-directionally transcribed iab-4/iab-8 microRNA locus in arthropods
Nucleic Acids Research ,
Mar 2013
Hui, Jerome H. L. , Marco, Antonio , Hunt, Suzanne , Melling, Janet , Griffiths-Jones, Sam , Ronshaugen, Matthew
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Structure, evolution and function of the bi-directionally transcribed iab-4/iab-8 microRNA locus in arthropods
Nucleic Acids Research
Structure, evolution and function of the bi-directionally transcribed iab-4/iab-8 microRNA locus in arthropods
Jerome H. L. Hui 0 1
Antonio Marco 0
Suzanne Hunt 0
Janet Melling 0
Sam Griffiths-Jones 0
Matthew Ronshaugen 0
0 Faculty of Life Sciences, University of Manchester , Michael Smith Building, Oxford Road, Manchester M13 9PT , UK
1 School of Life Sciences, Chinese University of Hong Kong , Shatin , Hong Kong
In Drosophila melanogaster, the iab-4/iab-8 locus encodes bi-directionally transcribed microRNAs that regulate the function of flanking Hox transcription factors. We show that bi-directional transcription, temporal and spatial expression patterns and Hox regulatory function of the iab-4/iab-8 locus are conserved between fly and the beetle Tribolium castaneum. Computational predictions suggest iab-4 and iab-8 microRNAs can target common sites, and cell-culture assays confirm that iab-4 and iab-8 function overlaps on Hox target sites in both fly and beetle. However, we observe key differences in the way Hox genes are targeted. For instance, abd-A transcripts are targeted only by iab-8 in Drosophila, whereas both iab-4 and iab-8 bind to Tribolium abd-A. Our evolutionary and functional characterization of a bi-directionally transcribed microRNA establishes the iab-4/iab-8 system as a model for understanding how multiple products from sense and antisense microRNAs target common sites.
INTRODUCTION
Specification of segmental identity on the head–tail axis of
all animals is accomplished, in large part, by the highly
coordinated expression of an ancient conserved complex
of related transcription factors collectively known as Hox
genes (
1
). Since the first molecular characterization of the
Drosophila Hox complex, it has been recognized that
numerous non-coding RNAs are interspersed between
the protein coding genes (
2,3
). Subsequent work in other
animals has found that non-coding RNAs are a prevalent
feature of many animal Hox complexes (4). Insight into
the function of a subset of these RNAs was facilitated by
the discovery of microRNAs (miRNAs): 22-bp RNAs
that direct translational repression. The Hox gene
complex contains several miRNAs, which are themselves
ancient genes, and their conserved positions within the
complex suggest that this association arose near the time
of the origin of the Hox complex itself (
1
). Only a small
number of metazoan miRNAs conserved between
distantly related metazoan animals date from this time
( 600 million years ago). Indeed, the oldest shared
conserved animal miRNA is miR-100, which was duplicated
in an ancestor of bilaterians to give rise to the Hox miR-10
miRNA (
5
). Studies of Hox complex miRNA function
find that they seem to act in part by modulating the
translation of Hox genes themselves. As translational
repression occurs after processing in the cytoplasm, there is no
obvious reason why a miRNA should target genes located
nearby in the genome.
Recent advances in sequencing technologies have
enabled large-scale identification of miRNAs in animals
(
6,7
). The available data suggest that only a tiny
proportion of miRNA loci are transcribed in both sense and
antisense directions. Indeed this bi-directional
transcription has been shown for only a handful of loci in a
variety of animals [e.g. Drosophila melanogaster (8) and
human (
9
)]. In these cases, the two distinct primary
transcripts are independently processed by Drosha and Dicer
to produce distinct mature miRNA sequences. The origin
and conservation of bi-directional miRNA transcription
and the functional relationships of products from sense
and antisense transcription have not been extensively
studied.
The miRNAs produced from the D. melanogaster iab-4
locus in the Hox complex are currently the only
functionally characterized sense/antisense miRNAs (
10–13
). The
bi-directionally transcribed iab-4 locus produces two
primary transcripts that fold into two distinct hairpin
precursors, named dme-iab-4 and dme-iab-8 (the latter is also
sometimes referred to as dme-iab-4-as). The spatial and
temporal expression of the sense and antisense transcripts
differ during embryogenesis (
10,12,13
). The mature
miRNA products regulate translation of the Hox genes
Ubx and abd-A in the fly (
10–13
). Interestingly, the
vertebrate mir-196 and the insect iab-4 miRNAs are located in
analogous regions in the Hox cluster and regulate
homologous Hox genes, yet there is no evidence that the two
miRNAs are themselves homologues (
14
). This has been
suggested as an example of ‘genomic parallelism’, where
selective pressure acts on miRNAs that have
independently arisen in this locus, resulting in functional
convergence to target the flanking Hox gene (
15
). Here, we
investigate the evolution of bi-directional transcription
at the iab-4/iab-8 miRNA locus. We use computational
analysis and in vivo assays to identify and confirm iab-4
and iab-8 target sites in insect Hox UTRs (unt (...truncated)
This is a preview of a remote PDF: https://academic.oup.com/nar/article-pdf/41/5/3352/16943491/gks1445.pdf
Hui, Jerome H. L., Marco, Antonio, Hunt, Suzanne, Melling, Janet, Griffiths-Jones, Sam, Ronshaugen, Matthew.
Structure, evolution and function of the bi-directionally transcribed iab-4/iab-8 microRNA locus in arthropods ,
Nucleic Acids Research,
2013, pp. 3352-3361, Volume 41, Issue 5, DOI: 10.1093/nar/gks1445