MicroRNA-dependent metamorphosis in hemimetabolan insects

Abstract

How does a juvenile insect transform into an adult? This question, which sums up the wonder of insect metamorphosis, has fascinated mankind since ancient times. Modern physiology has established the endocrine basis regulating these transformations, which mainly depend on two hormone types: ecdysteroids, which promote molts, and juvenile hormones, which repress the transformation into the adult stage. The interplay of these two hormones regulates the genes involved in juvenile and adult programs and the shift from one to the other. microRNAs (miRNAs) are small noncoding RNAs, which participate in many biological processes, and we wondered whether they might be also involved in insect metamorphosis. In insects, Dicer-1 ribonuclease transforms miRNA precursors into mature miRNAs. Thus, using systemic RNA interference (RNAi) to silence the expression of Dicer-1 in the hemimetabolan insect Blattella germanica, we depleted miRNA contents in the last instar nymph. This practically inhibited metamorphosis after the next molt, as the resulting specimens showed nymphoid features and were able to molt again. The experiments show that miRNAs play a key role in hemimetabolan metamorphosis, perhaps regulating genes that are juvenile hormone targets.Blattella ͉ dicer ͉ microRNAs R esearch into insect metamorphosis has traditionally focused on morphological aspects by differentiating two basic modes: the hemimetabolan and the holometabolan. Hemimetabolan species grow gradually, with each successive nymphal instar increasingly resembling the adult form until the final transition from nymph to adult, which is characterized by the displaying of functional wings and the appearance of external genitalia. Holometabolan species grow gradually through larval instars, which are very different from the adult, until the transitions from larva to pupa and from pupa to adult, which are characterized by dramatic morphological and functional changes (1). Since the 1940s, and mainly thanks to the contributions of Sir Vincent B. Wigglesworth, research in this field has focused on the endocrine aspects, particularly the actions of juvenile hormones (JHs) and ecdysteroids, and the gene cascades activated or repressed by them (2). Only a few reports considered the role of miRNAs, and all of which were based on the extremely modified holometabolan species Drosophila melanogaster (3, 4). miRNAs play a critical role in many biological processes, by modulating gene expression at the posttranscriptional level through binding at the 3Ј-untranslated region of the target mRNA (5, 6). Dicer ribonucleases are important in the biogenesis of miRNAs as they are involved in the production of mature miRNAs from miRNA precursors (premiRNAs), and of small interfering RNAs (siRNAs) in the RNA interference (RNAi) pathway (7). However, whereas a single Dicer ribonuclease is involved in both miRNA and siRNA production in the nematode Caenorhabditis elegans and in vertebrates, two of them, known as Dicer-1 and Dicer-2, which act in the miRNA and siRNA pathways, respect...

MiR-2 family regulates insect metamorphosis by controlling the juvenile hormone signaling pathway

“…We had observed in B. germanica that depletion of Dicer-1 and consequent reduction of miRNA levels prevented metamorphosis (24). The results presented herein show that Dicer-1 depletion results in increased levels of Kr-h1 mRNA, and that RNAi of Kr-h1 in Dicer-1 knockdown individuals rescues metamorphosis.…”

“…To gain a first insight into whether Kr-h1 might be a promising candidate, we tested whether depletion of Dicer-1, impairing mature miRNA formation, would affect Kr-h1 mRNA levels. We proceeded as in our previous work (24) and treated penultimate instar female nymphs (N5) of B. germanica with double-stranded RNA targeting Dicer-1 (dsDicer-1). As expected, Dicer-1 mRNA levels became depleted and mature miRNA levels were reduced (Fig.…”

“…The detailed procedures for RNAi have been reported previously (24). Primers to prepare the dsRNAs are described in Table S2.…”

“…Our approach involved silencing the expression of Dicer-1, the ribonuclease that produces mature miRNAs from miRNA precursors (22), following a method that had been used successfully in zebrafish (23). The approach was also effective for B. germanica, because depletion of Dicer-1 mRNA levels resulted in reduced levels of mature miRNAs and, importantly, prevented metamorphosis; last instar nymphs molted to supernumerary nymphs or to nymph-adult intermediates instead of normal adults (24).…”

Multiple miRNAs jointly regulate the biosynthesis of ecdysteroid in the holometabolous insects, Chilo suppressalis

“…It should be noticed that 1351 miRNAs were from 12 Drosophila species, which accounted for about half of known insect miRNAs (Kozomara and Griffiths-Jones 2014). Many studies to elucidate the functions of miRNAs in regulating a variety of insect physiological processes have been carried out, such as in metamorphosis development (Sokol et al 2008;Gomez-Orte and Belles 2009;Ling et al 2014;Belles 2017;Wu et al 2017), cell growth (Wang et al 2010), behavior (Cohen et al 2017), sex determination (McJunkin and Ambros 2017), oogenesis , embryogenesis (Jannot et al 2016), immunity (Zhu et al 2013), insect-pathogen interactions (Hussain and Asgari 2010;Asgari 2013;Wu et al 2016), etc. Several miRNAs have been reported to regulate insect metamorphosis by targeting genes either in the ecdysone cascade or the JH cascade (Zhang et al 2009;Belles 2017).…”