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. 2019 Oct 25;9:15336. doi: 10.1038/s41598-019-51703-x

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© The Author(s) 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Let-7d levels oscillate when cells undergo cell divisions. (A–E) A side-by-side comparison of horizontal sections from mouse cortex, at E13.5, analyzed by in situ hybridization for let-7d and with immunostaining for progenitor and cell cycle markers. (A,B) In situ hybridization for let-7d (purple) and immunostaining for Pax6 (green in A’) or Tbr2 (green in B’). Merged images in A” and B”. Let-7 levels vary (compare above and below yellow dashed line in A; arrowheads) within the Pax6+ domain (brackets in A,B). (C) In situ hybridization for let-7d (purple) and immunostaining for PH3 (green in C’). Merged image in C”. Let-7d levels are low in PH3+ cells (arrowheads). (D,E) In situ hybridization for let-7d (purple in D) shows that cells with the highest level of let-7d reside in the region where EdU+ cells in S-phase are present (red in E; see arrowheads). DAPI nuclear staining shown in blue (E). (F) Schematic of interkinetic nuclear migration. Progenitors migrate to the apical surface to divide. (G) Schematic highlighting the dynamic changes in let-7d expression pattern over time in the mouse cortex. (H–J) In situ hybridization for let-7d in the retina at E11.5 (purple in H) showed that let-7d levels vary within retinal progenitors (compare arrowheads on either side of yellow dashed line in H). Immunostaining for Tuj1 (red in I) and staining with DAPI (blue in I) showed that fluctuations in let-7d levels do not correlate with newly-generated, Tuj1+, post-mitotic cells (white arrowheads). (J) EdU+ cells in S-phase (red), immunostaining for PH3 (green), and DAPI nuclear staining (blue) suggest that cells having a high level of let-7d reside in the vicinity of EdU+ cells. (K) Schematic highlighting the dynamic changes in let-7d expression pattern over time in the mouse retina.