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. 2022 Apr 29;20(4):e3001631.
doi: 10.1371/journal.pbio.3001631. eCollection 2022 Apr.

Expression of Concern: Targeted Inhibition of miRNA Maturation with Morpholinos Reveals a Role for miR-375 in Pancreatic Islet Development

PLOS Biology Editors

Expression of Concern: Targeted Inhibition of miRNA Maturation with Morpholinos Reveals a Role for miR-375 in Pancreatic Islet Development

PLOS Biology Editors. PLoS Biol. .
No abstract available

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Figures

Fig 1
Fig 1. Morpholinos Targeting the Mature miRNA Deplete the Zebrafish Embryo of Specific miRNAs.
(A) Removed panel previously showing Northern blot for miR-206 in wild-type and MO miR-206–injected embryos at 24, 48, and 72 hpf. (B) In situ analysis of miR-206 and miR-124 expression in different stage embryos after injection of MO miR-206. (C) Effect of a morpholino targeting miR-30c on a silencing assay with miR-30c and a responsive GFP sensor construct. (D) In vivo and in vitro effects of a morpholino on the stability and RNA extraction of a synthetic miR-206 duplex. miR-205 serves as a loading control.
Fig 2
Fig 2. Morpholinos Targeting the Precursor miRNA Interfere with miRNA Maturation.
(A) Design of morpholinos targeting the precursor miRNA. (B) Northern blot analysis of miR-205 in 30-h-old embryos injected with different morpholinos against pri-miR-205. 5S RNA serves as a loading control. (C) Time series of miR-205 expression after injection of mature, no lap loop, and drosha star morpholinos against pri-miR-205. (D) Northern blot analysis of miR-205 derived from embryos injected with a GFP-pri-miR-205 transcript and four different morpholinos targeting pri-miR-205. Co-injected miR-206 serves as an injection and loading control. Embryos were collected 8 h after injection. (E) GFP expression in 24-h embryos injected with morpholinos and a GFP-pri-miR-205 construct as used in (C). Pri-miR-205 is positioned just upstream of the polyA signal in the 3′ UTR of the GFP mRNA. Red fluorescent protein (RFP) serves as an injection control. (F) RT-PCR analysis of injected GFP-pri-miR-205 mRNA with (+) and without (−) co-injected morpholinos. Luciferase serves as an injection control. Embryos were collected 8 h after injection. (G) Northern analysis of the effect of morpholinos on an injected miR-205 precursor. Embryos were collected 8 h after injection. WT, wild type.
Fig 5
Fig 5. Knockdown of miR-375 Results in Aberrant Migration of Pancreatic Islet Cells.
(A) In situ analysis of miR-375 knockdown in MO miR-375–injected embryos and noninjected controls at 24, 48, 72, and 120 hpf. Arrowheads indicate the pituitary gland and the pancreatic islet. (B) In situ analysis of the pancreatic islet (insulin staining) and the pituitary gland (pit1 staining) in miR-375 morphants and noninjected controls. Arrowheads indicate the pituitary gland and the pancreatic islet. (C) In situ analysis of pancreatic islet development in wild-type and morphant embryos using insulin, somatostatin, and glucagon as markers. (D) Time series of insulin expression in wild-type and morphant embryos injected with MO miR-375. The WT 72h panel of Fig 5D is identical to the WT insulin panel of Fig 6A, due to these panels representing the same experimental condition. (E) Insulin expression in 72-hpf embryos injected with MO miR-375 and a complementary morpholino.
Fig 6
Fig 6. Specific Effects of miR-375 Knockdown on the Development of the Endocrine Pancreas.
(A) In situ analysis of miR-375 and insulin expression in 72-hpf embryos injected with morpholinos against the miR-375 precursor and negative control morpholinos for let-7 and miR-124. The WT insulin panel of Fig 6A is identical to the WT 72h panel of Fig 5D, due to these panels representing the same experimental condition. (B) Expression of islet1, foxa2, and ptf1a in wild-type and miR-375 knockdown embryos. Arrows indicate the pancreatic islet. WT, wild type.
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References

    1. Kloosterman WP, Lagendijk AK, Ketting RF, Moulton JD, Plasterk RHA (2007) Targeted Inhibition of miRNA Maturation with Morpholinos Reveals a Role for miR-375 in Pancreatic Islet Development. PLoS Biol 5(8): e203. 10.1371/journal.pbio.0050203 - DOI - PMC - PubMed
    1. Kloosterman Wigard P., Steiner Florian A., Berezikov Eugene, Ewart de Bruijn, Jose van de Belt, Mark Verheul, Edwin Cuppen, Ronald H.A. Plasterk, Cloning and expression of new microRNAs from zebrafish, Nucleic Acids Research, Volume 34, Issue 9, 1 May 2006, Pages 2558–2569, 10.1093/nar/gkl278 - DOI - PMC - PubMed

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