• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of nihpaAbout Author manuscriptsSubmit a manuscriptNIH Public Access; Author Manuscript; Accepted for publication in peer reviewed journal;
Curr Biol. Author manuscript; available in PMC Aug 26, 2009.
Published in final edited form as:
PMCID: PMC2583239
NIHMSID: NIHMS77470

let-7: Developmental timing conserved through evolution

Expression of the let-7 heterochronic microRNA is tightly correlated with the onset of adult development in many bilateral species suggesting that it acts as a evolutionarily conserved developmental timer. This hypothesis has now been confirmed by studies in Drosophila

The current excitement surrounding microRNAs (miRNAs) makes it difficult to remember that this field arose from elegant genetic studies of developmental timing. The two founding members of the miRNA family, lin-4 and let-7, were discovered in genetic screens for heterochronic defects in C. elegans [13]. As core components of the heterochronic pathway, lin-4 and let-7 act in genetic switches that regulate progression through stage-specific developmental events [4, 5]. For example, the dramatic up-regulation of let-7 near the end of larval development results in reduced expression of key heterochronic proteins that promote larval-specific cell fates, thereby ensuring the successful transition into adulthood [4, 5]. As might be expected, the timing of let-7 expression is crucial. Precocious let-7 expression leads to the premature onset of adult fates while the absence of let-7 retards exit from the juvenile stage [3, 6].

In a landmark paper, Pasquinelli et al. [2] reported that the let-7 miRNA is not restricted to nematodes but rather is conserved throughout bilateral animals. More importantly, its expression pattern is also conserved, with let-7 induction tightly coordinated with the progression from juvenile to adult fates in all species examined. These observations raised the exciting possibility that let-7 acts as an evolutionarily conserved regulator of adult fates, a hypothesis that has waited nearly a decade for in vivo confirmation. Two recent papers, by Caygill and Johnston in this issue of Current Biology [7] and Sokol et al. [8], have addressed this topic, and defined key roles for let-7 in controlling the juvenile-to-adult transition in several tissues of Drosophila.

let-7 regulates developmental timing in Drosophila

The Drosophila let-7 miRNA is induced at the onset of metamorphosis and processed from an RNA precursor that contains two other conserved miRNAs, miR-100 and miR-125 [911]. Both research teams use gene targeting to specifically delete let-7, although this mutation also inactivates miR-125 in the study by Caygill and Johnson [7, 8]. For the purpose of simplicity, we focus this review on phenotypes that have been characterized using both a loss of let-7 function as well as specific ectopic let-7 expression.

Caygill and Johnston [7] found that their mutant displays widespread defects during metamorphosis, with surviving adults having small wings. They attribute this phenotype to a significant reduction in cell size, in spite of the presence of more cells than wild type. Like other cuticular structures in the adult fly, the wing develops from a population of progenitor cells in the imaginal discs. These disc cells proliferate throughout larval stages until cell divisions cease at the onset of metamorphosis in preparation for terminal differentiation. Since C. elegans let-7 acts to temporally restrict specific larval cell divisions, Caygill and Johnston asked if the extra wing cells result from a delayed exit from the cell cycle. Consistent with this hypothesis, the cells in mutant wing discs continue to divide 24 hrs after puparium formation, a time when divisions have largely ceased in wild-type cells. In the reciprocal experiment, ectopic expression of let-7 during larval development causes wing disc cells to precociously exit the cell cycle. Thus, the onset of let-7 expression at puparium formation temporally restricts the period during which wing disc cells undergo division.

This newfound role for let-7 in the imaginal discs provides an exciting complement to studies of let-7 in other systems. Prior to this discovery, our understanding of let-7 function in vivo has primarily focused on its roles in the development of a specific C. elegans epidermal cell type referred to as seam cells. When let-7 is absent, the seam cells continue to divide and fail to undergo terminal differentation [3]. Although significantly less is known about let-7 in humans and mice, the observation that let-7 is abundantly expressed in differentiated tissues and absent in progenitor cells and certain types of cancer cells suggests a role in regulating cell proliferation [12, 13]. Future studies of let-7 in the Drosophila wing disc, which is an ideal system for detailed characterization of gene function, provides a new opportunity to understand how this miRNA controls the timing of cell divisions in higher animals.

Studies in Drosophila may also help to unravel the role of let-7 in other cell types. Sokol et al. [8] observed that let-7 is expressed in a wide range of tissues including the central nervous system, motor neurons, and muscle. Consistent with this expression pattern, both groups observe defects in neuromuscular development [7, 8]. Sokol and colleagues find that the dorsal internal oblique muscles (DIOMs), which are normally destroyed during adult maturation, persist in let-7 mutant adults, while both groups show that the adult-specific dorsal muscles (DM) and their associated neuromuscular junctions (NMJ) appear immature when compared to wild-type controls.

The finding that let-7 induces the destruction of the larval-specific DIOMs is intriguingly reminiscent of a recently described role for let-7 in C. elegans male development, where a single cell known as the linker cell dies at the larval-to-adult transition [14]. The death of this cell is dependent on the expression of let-7 but independent of the known apoptotic machinery. Does let-7 regulate DIOM destruction via a similar mechanism? If so, then the molecular pathways that govern this response may provide important new insights into our understanding of cell death regulation. Moreover, the behavioral defects seen in let-7 mutants and the widespread expression of let-7 in the nervous system of both flies and humans suggests that further studies of fly let-7 will provide insights into the regulation of neuronal maturation during adult development [3, 7, 8].

Discovery of a new let-7 target

The characterization of let-7 in Drosophila provides, for the first time, an opportunity to identify its in vivo targets in an organism other than C. elegans. Caygill and Johnston [7] have taken advantage of this new resource by demonstrating that the BTB-zinc finger transcription factor Abrupt (Ab) is regulated by let-7. Several important characteristics of the abrupt gene define it as a likely direct target for let-7 control. The 3′-untranslated region of the abrupt mRNA contains five let-7 binding sites [15], and Ab protein is down-regulated during metamorphosis in synchrony with the up-regulation of let-7. Furthermore, Ab protein levels are intimately linked to let-7 expression. While ectopic let-7 expression in wing disc cells leads to precocious down-regulation of Ab protein, levels of Ab remain high in let-7 mutants. Moreover, the retarded DM NMJ phenotype observed in let-7 mutants is suppressed by a partial loss of abrupt function, suggesting that this phenotype is due to Ab overexpression. Ab thus appears to be a critical regulator of developmental timing in Drosophila, inhibiting adult fates until it is down-regulated by let-7 during terminal differentiation.

Integrating heterochronic and endocrine timers

As we have discussed previously [16], studies in C. elegans and Drosophila have revealed two distinct aspects of developmental timing. Studies in C. elegans have focused primarily on roles for the heterochronic genes in assigning temporal identity to specific cells within the context of their defined lineages. In contrast, studies in Drosophila have focused on the role of systemic pulses of the steroid hormone 20-hydroxyecdysone (20E) in establishing temporal boundaries that define developmental transitions in the life cycle [16]. The discovery of heterochronic functions for let-7 in Drosophila provides an exciting new opportunity to integrate these two temporal pathways. The fly let-7 miRNA is induced in late third instar larvae, in precise synchrony with known direct targets of 20E and its EcR receptor, as the hormone triggers puparium formation and the onset of adult differentiation [2, 9, 11]. Curiously, however, this induction appears to occur independently of either 20E or EcR, implying the existence of other systemic temporal regulators [9]. This could be provided by the precursor to 20E, ecdysone, which also peaks in late third instar larvae and appears to have distinct hormonal activity [17, 18]. Interestingly, some let-7 target genes also appear to be regulated by 20E. Both abrupt and a lin-41 homolog brat are induced by 20E and potentially down-regulated by let-7 [17, 19]. Future studies of let-7 regulation in Drosophila as well as the characterization of additional let-7 targets should provide a new basis for understanding how the cellular temporal identity conferred by heterochronic genes is integrated with the systemic temporal boundaries established by hormone signaling.

References

1. Lee RC, Feinbaum RL, Ambros V. The C. elegans heterochronic gene lin-4 encodes small RNAs with antisense complementarity to lin-14. Cell. 1993;75:843–854. [PubMed]
2. Pasquinelli AE, Reinhart BJ, Slack F, Martindale MQ, Kuroda MI, Maller B, Hayward DC, Ball EE, Degnan B, Muller P, Spring J, Srinivasan A, Fishman M, Finnerty J, Corbo J, Levine M, Leahy P, Davidson E, Ruvkun G. Conservation of the sequence and temporal expression of let-7 heterochronic regulatory RNA. Nature. 2000;408:86–89. [PubMed]
3. Reinhart BJ, Slack FJ, Basson M, Pasquinelli AE, Bettinger JC, Rougvie AE, Horvitz HR, Ruvkun G. The 21-nucleotide let-7 RNA regulates developmental timing in Caenorhabditis elegans. Nature. 2000;403:901–906. [PubMed]
4. Moss EG. Heterochronic genes and the nature of developmental time. Curr Biol. 2007;17:R425–434. [PubMed]
5. Rougvie AE. Intrinsic and extrinsic regulators of developmental timing: from miRNAs to nutritional cues. Development (Cambridge, England) 2005;132:3787–3798. [PubMed]
6. Hayes GD, Ruvkun G. Misexpression of the Caenorhabditis elegans miRNA let-7 is sufficient to drive developmental programs. Cold Spring Harb Symp Quant Biol. 2006;71:21–27. [PubMed]
7. Caygill EE, Johnston LA. Temporal regulation of metamorphic processes in Drosophila by the let-7 and miR-125 heterochronic microRNAs. Curr Biol 2008 [PMC free article] [PubMed]
8. Sokol NS, Xu P, Jan YN, Ambros V. Drosophila let-7 microRNA is required for remodeling of the neuromusculature during metamorphosis. Genes Dev. 2008;22:1591–1596. [PMC free article] [PubMed]
9. Bashirullah A, Pasquinelli AE, Kiger AA, Perrimon N, Ruvkun G, Thummel CS. Coordinate regulation of small temporal RNAs at the onset of Drosophila metamorphosis. Dev Biol. 2003;259:1–8. [PubMed]
10. Sempere LF, Dubrovsky EB, Dubrovskaya VA, Berger EM, Ambros V. The expression of the let-7 small regulatory RNA is controlled by ecdysone during metamorphosis in Drosophila melanogaster. Dev Biol. 2002;244:170–179. [PubMed]
11. Sempere LF, Sokol NS, Dubrovsky EB, Berger EM, Ambros V. Temporal regulation of microRNA expression in Drosophila melanogaster mediated by hormonal signals and broad-Complex gene activity. Dev Biol. 2003;259:9–18. [PubMed]
12. Ibarra I, Erlich Y, Muthuswamy SK, Sachidanandam R, Hannon GJ. A role for microRNAs in maintenance of mouse mammary epithelial progenitor cells. Genes Dev. 2007;21:3238–3243. [PMC free article] [PubMed]
13. Cho WC. OncomiRs: the discovery and progress of microRNAs in cancers. Mol Cancer. 2007;6:60. [PMC free article] [PubMed]
14. Abraham MC, Lu Y, Shaham S. A morphologically conserved nonapoptotic program promotes linker cell death in Caenorhabditis elegans. Dev Cell. 2007;12:73–86. [PubMed]
15. Burgler C, Macdonald PM. Prediction and verification of microRNA targets by MovingTargets, a highly adaptable prediction method. BMC Genomics. 2005;6:88. [PMC free article] [PubMed]
16. Thummel CS. Molecular mechanisms of developmental timing in C. elegans and Drosophila. Dev Cell. 2001;1:453–465. [PubMed]
17. Beckstead RB, Lam G, Thummel CS. The genomic response to 20-hydroxyecdysone at the onset of Drosophila metamorphosis. Genome Biol. 2005;6:R99. [PMC free article] [PubMed]
18. Champlin DT, Truman JW. Ecdysteroid control of cell proliferation during optic lobe neurogenesis in the moth Manduca sexta. Development (Cambridge, England) 1998;125:269–277. [PubMed]
19. Slack FJ, Basson M, Liu Z, Ambros V, Horvitz HR, Ruvkun G. The lin-41. RBCC gene acts in the C. elegans heterochronic pathway between the let-7 regulatory RNA and the LIN-29 transcription factor. Mol Cell. 2000;5:659–669. [PubMed]
PubReader format: click here to try

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...