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. 2016 Sep 21;5(4):e1238560.
doi: 10.1080/21624054.2016.1238560. eCollection 2016.

sta-1 is repressed by mir-58 family in Caenorhabditis elegans

Encarnación Lozano  1 , María Pilar de Lucas  1 , Alberto G Sáez  1
Affiliations

sta-1 is repressed by mir-58 family in Caenorhabditis elegans

Encarnación Lozano et al. Worm. .

Abstract

The miR-58 family comprises 6 microRNAs with largely shared functions, and with an overall high expression, because one of its members, miR-58, is the most abundant microRNA in Caenorhabditis elegans. We recently found that 2 TGF-β signaling pathways, Sma/Mab and Dauer, responsible for body size and dauer formation respectively, among other phenotypes, are downregulated by the miR-58 family. Here, we further explore this family by showing that it also acts through the sta-1 3'UTR. sta-1 encodes a transcription factor, homologous to mammalian STATs, that inhibits dauer formation in association with the TGF-β Dauer pathway. We also observe that mutants with a constitutively active TGF-β Dauer pathway express higher levels of sta-1 mRNA. Our results reinforce the view of the miR-58 family and STA-1 as regulators of dauer formation in coordination with the TGF-β Dauer pathway.

Keywords: Dauer; TGF-β Dauer; microRNAs; mir-58; sta-1.

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Figures

Figure 1.
Figure 1.
sta-1 mRNA levels are upregulated in the mir-58 family mutant and in worms with a constitutively active TGF-β Dauer pathway. mRNA levels of sta-1 were measured in mixed-staged populations of wild-type strain N2, the quadruple mutant mir-58f(−) (MT15563) and 3 additional strains with constitutively active TGF-β Dauer signaling: daf-3(ok3610) (RB2589), daf-4(++) (pwIs922), and daf-8(++) (DR2490). Measurements were carried out by quantitative Real-Time PCR with a specific TaqMan probe for sta-1. mRNA levels of mutant strains (color bars) were normalized to those of N2 (white bars). Each value represents the average from 4 independent biological replicates. Error bars indicate standard deviations. Significant statistical differences between each mutant and N2 are indicated as *(P < 0.001).
Figure 2.
Figure 2.
Luciferase-reporter assays show that miR-58 family members repress gene expression through the sta-1 3′UTR. Human HeLa cells were transiently transfected with psiCHECK-2 vector containing either the wild-type (white) or mutated (gray) 3′UTR from sta-1, along with miR-58 family mimics of miR-58.1, miR-80, miR-81 and miR-58.2 (i.e. miR-1834), or the unrelated miR-67 as negative control. The luciferase activity for each mimic was normalized to the value obtained with miR-67 using the same sta-1 3′UTR. Data shown are representative from 2 independent experiments. Error bars indicate standard deviations. *(P < 0.005), comparing luciferase activities for each inhibitory miRNA to miR-67s (white bars), and between wild-type and mutated sta-1 3′UTRs for each microRNA (pairs of white and gray bars).
Figure 3.
Figure 3.
Model of genetic interactions between the family of miR-58, TGF-β Dauer, sta-1 and the dauer phenotype. Based on our results (this work and reference 5), and on the sta-1 and TGF-β Dauer literature. The thin line between sta-1 and dauer illustrates the weaker inhibitory role of sta-1 on dauer compared to the stronger effect from the TGF-β Dauer pathway.
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