Drosophila let-7 microRNA is required for remodeling of the neuromusculature during metamorphosis

Abstract

The Drosophila let-7-Complex (let-7-C) is a polycistronic locus encoding three ancient microRNAs: let-7, miR-100, and fly lin-4 (miR-125). We find that the let-7-C locus is principally expressed in the pupal and adult neuromusculature. let-7-C knockout flies appear normal externally but display defects in adult behaviors (e.g., flight, motility, and fertility) as well as clear juvenile features in their neuromusculature. We find that the function of let-7-C to ensure the appropriate remodeling of the abdominal neuromusculature during the larval-toadult transition is carried out predominantly by let-7 alone. This heterochronic role of let-7 is likely just one of the ways in which let-7-C promotes adult behavior.Supplemental material is available at http://www.genesdev.org.Received March 10, 2008; revised version accepted April 11, 2008. Mutations in heterochronic genes in Caenorhabditis elegans cause cells in particular lineages to express their stage-specific fates earlier or later than normal (Ambros and Horvitz 1984). Detailed analysis of these genes has revealed a regulatory pathway of heterochronic genes that specifies the timing of cellular development in diverse cell types and thereby ensures a coordinated schedule of developmental events throughout the worm (for review, see Rougvie 2005; Moss 2007). The existence of the heterochronic gene pathway in worms and the conservation of some of its components through animal evolution suggest that functionally analogous pathways could also coordinate developmental timing in higher organisms . Two of these highly conserved components of the heterochronic pathway, let-7 and lin-4, are microRNAs (miRNAs), a class of small RNAs that post-transcriptionally modulate the expression of target transcripts (for review, see Jackson and Standart 2007). The sequences and developmentally regulated expression profiles of let-7 and lin-4 are conserved among diverse bilaterians Sempere et al. 2003). For example, Drosophila let-7 and miR-125 (fly lin-4) are robustly up-regulated during metamorphosis, as is another highly conserved miRNA, miR-100 Sempere et al. 2002Sempere et al. , 2003Bashirullah et al. 2003). All three of these ancient miRNAs are encoded in a 1-kb region of the Drosophila genome ( Fig. 1; Sempere et al. 2003), and their clustered organization has been conserved and duplicated in vertebrates (Supplemental Fig. S1; Sempere et al. 2003;Prochnik et al. 2007). These findings suggest that miR-100, let-7, and miR-125 coordinately control gene expression to regulate developmental timing in animals.To test this hypothesis, we analyzed the roles of miR-100, let-7, and miR-125 in Drosophila and find that these miRNAs are required for normal adult behavior, suggesting roles in neural development and/or function. let-7 in particular is required for remodeling of the fly neuromusculature during the larval-to-adult transition, confirming that a general developmental timing function of let-7 has been evolutionarily conserved from worms to flies. Results and DiscussionThe clus...

Differential expression of miRNAs related to caste differentiation in the honey bee, Apis mellifera

“…The loss of one or multiple of them in Drosophila induced the adult behavioral deficiencies in oviposition, fertility, and oogenesis (Iovino et al 2009;Sokol et al 2008). Since there are obverse discrepancies between workers and queens in adult behaviors such as flight, motility, oviposition, and fertility, we could hypothesize that miRNA let-7, miR-100, and miR-184 may play miRNAs in caste differentiation of honey bee crucial roles in the caste switch in bees as JHrelated miRNAs.…”

“…Female mouse Dicer knock-out mutants are infertile (Escalier 2008;Hong et al 2008;Luense et al 2009). In Drosophila , knock-out mutants of the gene cluster comprising the miR-100, let-7, and miR-125 genes display reduced fertility, motility, and flight (Sokol et al 2008). The microRNA bantam regulates cell growth and proliferation, organ and body size, and germ cell fate (Brennecke et al 2003;Hipfner et al 2002;Moberg and Hariharan 2003;Yang et al 2009).…”

Regulation of Drosophila circadian rhythms by miRNA let-7 is mediated by a regulatory cycle

“…To exclude the possibility that the behavioural phenotype was caused by ectopic induction of let-7 in neurons where it is usually not expressed, we utilized a knock-in Gal4 in the let-7-C locus ( Supplementary Fig. 1b) 33 to drive expression of let-7-C, and showed the miRNA levels of let-7, miR-125 and miR-100 were more abundant ( Supplementary Fig. 2a).…”

“…Single let-7, miR-125, miR-100 mutant strains were generated by deletions of rescuing transgenes, in which the expression of let-7, miR-100 or miR-125 had been separately eliminated, introduced into strains with deletions of the entire endogenous let-7-C locusnamed nlet-7 (KO1/GKI;pP{W8,let-7-C nlet-7 }/ þ ), nmiR-100 (KO1/GKI;pP{W8,let-7-C nmiR-100 }/ þ ) and nmiR-125 (KO1/ GKI;pP{W8,let-7-C nmiR-125 }/ þ ) ( Supplementary Fig. 1b) 33 . In this study, PCR and real time quantitative reverse transcriptase PCR analysis were performed to confirm the deletion of the let-7-C locus and individual miRNAs ( Supplementary Fig.…”

“…Deletions of the let-7-C and let-7,miR-125 had been generated by homologous recombination methods 32,33 . Single let-7, miR-125, miR-100 mutant strains were generated by deletions of rescuing transgenes, in which the expression of let-7, miR-100 or miR-125 had been separately eliminated, introduced into strains with deletions of the entire endogenous let-7-C locusnamed nlet-7 (KO1/GKI;pP{W8,let-7-C nlet-7 }/ þ ), nmiR-100 (KO1/GKI;pP{W8,let-7-C nmiR-100 }/ þ ) and nmiR-125 (KO1/ GKI;pP{W8,let-7-C nmiR-125 }/ þ ) ( Supplementary Fig.…”