Transcription of the C. elegans let-7 microRNA is temporally regulated by one of its targets, hbl-1

Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 06520, USA.
Developmental Biology (Impact Factor: 3.55). 08/2009; 334(2):523-34. DOI: 10.1016/j.ydbio.2009.07.012
Source: PubMed


The let-7 family of microRNAs (miRNAs) are important regulators of developmental timing and cell differentiation and are often misexpressed in human cancer. In C. elegans, let-7 controls cell fate transitions from larval stage 4 (L4) to adulthood by post-transcriptionally down-regulating lineage-abnormal 41 (lin-41) and hunchback-like 1 (hbl-1). Primary let-7 (pri-let-7) transcripts are up-regulated in the L3, yet little is known about what controls this transcriptional up-regulation. We sought factors that either turn on let-7 transcription or keep it repressed until the correct time. Here we report that one of let-7's targets, the transcription factor Hunchback-like 1 (HBL-1), is responsible for inhibiting the transcription of let-7 in specific tissues until the L3. hbl-1 is a known developmental timing regulator and inhibits adult development in larval stages. Therefore, one important function of HBL-1 in maintaining larval stage fates is inhibition of let-7. Indeed, our results reveal let-7 as the first known target of the HBL-1 transcription factor in C. elegans and suggest a negative feedback loop mechanism for let-7 and HBL-1 regulation.

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    • "The expression of HMGA2 is regulated via micro RNAs (miRNA) of the let-7 family by binding to sequences located in the 3′ untranslated region (UTR) of the transcript [31]–[35], all of which are conserved in rodents, dog, and chicken [36]–[38]. Binding of let-7 miRNAs to complementary sequences regulates post-transcriptionally the expression of HMGA2 in a negative way [31], [35], [39], [40]. Recently a deregulated let-7 expression was associated with lung [41], [42], breast [43] and prostate cancer [44]. "
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    ABSTRACT: The architectural transcription factor HMGA2 is abundantly expressed during embryonic development. In several malignant neoplasias including prostate cancer, high re-expression of HMGA2 is correlated with malignancy and poor prognosis. The let-7 miRNA family is described to regulate HMGA2 negatively. The balance of let-7 and HMGA2 is discussed to play a major role in tumour aetiology. To further analyse the role of HMGA2 in prostate cancer a stable and highly reproducible in vitro model system is precondition. Herein we established a canine CT1258-EGFP-HMGA2 prostate cancer cell line stably overexpressing HMGA2 linked to EGFP and in addition the reference cell line CT1258-EGFP expressing solely EGFP to exclude EGFP-induced effects. Both recombinant cell lines were characterised by fluorescence microscopy, flow cytometry and immunocytochemistry. The proliferative effect of ectopically overexpressed HMGA2 was determined via BrdU assays. Comparative karyotyping of the derived and the initial CT1258 cell lines was performed to analyse chromosome consistency. The impact of the ectopic HMGA2 expression on its regulator let-7a was analysed by quantitative real-time PCR. Fluorescence microscopy and immunocytochemistry detected successful expression of the EGFP-HMGA2 fusion protein exclusively accumulating in the nucleus. Gene expression analyses confirmed HMGA2 overexpression in CT1258-EGFP-HMGA2 in comparison to CT1258-EGFP and native cells. Significantly higher let-7a expression levels were found in CT1258-EGFP-HMGA2 and CT1258-EGFP. The BrdU assays detected an increased proliferation of CT1258-HMGA2-EGFP cells compared to CT1258-EGFP and native CT1258. The cytogenetic analyses of CT1258-EGFP and CT1258-EGFP-HMGA2 resulted in a comparable hyperdiploid karyotype as described for native CT1258 cells. To further investigate the impact of recombinant overexpressed HMGA2 on CT1258 cells, other selected targets described to underlie HMGA2 regulation were screened in addition. The new fluorescent CT1258-EGFP-HMGA2 cell line is a stable tool enabling in vitro and in vivo analyses of the HMGA2-mediated effects on cells and the development and pathogenesis of prostate cancer.
    PLoS ONE 06/2014; 9(6):e98788. DOI:10.1371/journal.pone.0098788 · 3.23 Impact Factor
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    • "Expression of hbl-1 is, in turn, repressed by let-7 and its sister miRNAs, which then relieves hbl-1 inhibition of let-7 transcription by the L3 stage (Abbott et al., 2005; Abrahante et al., 2003; Lin et al., 2003). The HBL-1 responsive element was predicted to reside downstream of the TRE before the A start site (Roush and Slack, 2009). This sequence contains an A-rich element similar to the binding site defined for Hunchback in other organisms (Stanojevic et al., 1989; Treisman and Desplan, 1989). "
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    ABSTRACT: The let-7 microRNA (miRNA) is highly conserved across animal phyla and generally regulates cellular differentiation and developmental timing pathways. In C. elegans, the mature let-7 miRNA starts to accumulate in the last stages of larval development where it directs cellular differentiation programs required for adult fates. Here we show that expression of the let-7 gene in C. elegans is under complex transcriptional control. The onset of let-7 transcription begins as early as the first larval stage in some tissues, and as late as the third larval stage in others, and is abrogated at the gravid adult stage. Transcription from two different start sites in the let-7 promoter oscillates during each larval stage. We show that transcription is regulated by two distinct cis-elements in the promoter of let-7, the previously described temporal regulatory element (TRE), and a novel element downstream of the TRE that we have named the let-7 transcription element (LTE). These elements play distinct and redundant roles in regulating let-7 expression in specific tissues. In the absence of the TRE and LTE, transcription of let-7 is undetectable and worms exhibit the lethal phenotype characteristic of let-7 null mutants. We also identify several genes that affect the transcription of let-7 generally and tissue-specifically. Overall, spatio-temporal regulation of let-7 transcription is orchestrated by multiple cis- and trans-acting factors to ensure appropriate expression of this essential miRNA during worm development.
    Developmental Biology 11/2012; 374(1). DOI:10.1016/j.ydbio.2012.11.021 · 3.55 Impact Factor
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    • "Student's t test) had completed this process (Fig. 4d–e and S4a). Similar delays were observed in strains containing two independent hbl-1 alleles (mg285 and ve18), both of which reduce but do not eliminate hbl-1 gene activity (Abrahante et al., 2003; Lin et al., 2003; Roush and Slack, 2009). The hbl-1 delayed remodeling defect was rescued by a transgene containing the F13D11 cosmid (which spans the hbl-1 gene; Fig. 4e). "
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    ABSTRACT: During development, circuits are refined by the dynamic addition and removal of synapses; however, little is known about the molecular mechanisms that dictate where and when synaptic refinement occurs. Here we describe transcriptional mechanisms that pattern remodeling of C. elegans neuromuscular junctions (NMJs). The embryonic GABAergic DD motor neurons remodel their synapses, whereas the later born VD neurons do not. This specificity is mediated by differential expression of a transcription factor (HBL-1), which is expressed in DD neurons but is repressed in VDs by UNC-55/COUP-TF. DD remodeling is delayed in hbl-1 mutants whereas precocious remodeling is observed in mutants lacking the microRNA mir-84, which inhibits hbl-1 expression. Mutations increasing and decreasing circuit activity cause corresponding changes in hbl-1 expression, and corresponding shifts in the timing of DD plasticity. Thus, convergent regulation of hbl-1 expression defines a genetic mechanism that patterns activity-dependent synaptic remodeling across cell types and across developmental time.
    Neuron 02/2012; 73(3):453-65. DOI:10.1016/j.neuron.2011.11.025 · 15.05 Impact Factor
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