miR-335 promotes mesendodermal lineage segregation and shapes a transcription factor gradient in the endoderm

Abstract

Transcription factors (TFs) pattern developing tissues and determine cell fates; however, how spatio-temporal TF gradients are generated is ill defined. Here we show that miR-335 fine-tunes TF gradients in the endoderm and promotes mesendodermal lineage segregation. Initially, we identified miR-335 as a regulated intronic miRNA in differentiating embryonic stem cells (ESCs). miR-335 is encoded in the mesoderm-specific transcript (Mest) and targets the 3′-UTRs of the endoderm-determining TFs Foxa2 and Sox17. Mest and miR-335 are co-expressed and highly accumulate in the mesoderm, but are transiently expressed in endoderm progenitors. Overexpression of miR-335 does not affect initial mesendoderm induction, but blocks Foxa2-and Sox17-mediated endoderm differentiation in ESCs and ESC-derived embryos. Conversely, inhibition of miR-335 activity leads to increased Foxa2 and Sox17 protein accumulation and endoderm formation. Mathematical modeling predicts that transient miR-335 expression in endoderm progenitors shapes a TF gradient in the endoderm, which we confirm by functional studies in vivo. Taken together, our results suggest that miR-335 targets endoderm TFs for spatio-temporal gradient formation in the endoderm and to stabilize lineage decisions during mesendoderm formation.KEY WORDS: Foxa2, Sox17, Endoderm, Mesendoderm, miR-335, Mir335, Gastrulation, Mouse INTRODUCTIONThe first lineage decision during mouse development occurs when the morula develops to the blastocyst stage at embryonic day (E) 2.5-3.5 (Rossant and Tam, 2009). During this time, the inner cell mass (ICM) of the blastocyst segregates from the trophectoderm (TE) that will form the placenta. The ICM further develops to the epiblast epithelium that will give rise to all differentiated cell types in the mammalian body. At E6.5, gastrulation starts and pluripotent Oct4 + epiblast cells undergo epithelial-mesenchymal transition (EMT) to ingress into the posterior primitive streak (PS) region to form mesoderm and definitive endoderm (DE), whereas the remaining epiblast cells will form the ectoderm (Beddington and Robertson, 1999;Tam and Loebel, 2007;Zorn and Wells, 2009). Both EMT and mesendoderm differentiation are induced by Wnt/β-catenin and Nodal/TGFβ signaling in the mouse embryo Arnold and Robertson, 2009). Wnt/β-catenin signaling leads to the activation of target genes in the epiblast, such as the Forkhead transcription factor a2 (Foxa2) (Sasaki and Hogan, 1993;Monaghan et al., 1993;Sawada et al., 2005) and the T-box transcription factor Brachyury (T; Herrmann, 1991;Yamaguchi et al., 1999;Arnold et al., 2000), which mark distinct mesendodermal progenitor cell populations in the posterior epiblast (Burtscher and Lickert, 2009). Genetic lineage tracing experiments revealed that Foxa2 + and T + mesendoderm progenitors give rise to anterior and posterior mesendoderm populations, respectively (Uetzmann et al., 2008;Horn et al., 2012; Kumar et al., 2007;Verheyden et al., 2005); however, how lineageinappropriate TF expression is prevented after...

MiR‐335 overexpression impairs insulin secretion through defective priming of insulin vesicles

“…For instance, miR‐335 directly targets the transcription factors FOXA2 and SOX17 and miR‐335 thereby promotes mesendodermal lineage segregation (Yang et al. ). FOXA2 is not only involved in developmental β ‐cell differentiation, but the protein also impact β ‐cell function.…”

“…It is possible that other targets of miR-335 contribute to the regulation of insulin secretion, resulting in a net cumulative negative effect of miR-335 on insulin secretion. For instance, miR-335 directly targets the transcription factors FOXA2 and SOX17 and miR-335 thereby promotes mesendodermal lineage segregation (Yang et al 2014). FOXA2 is not only involved in developmental b-cell differentiation, but the protein also impact b-cell function.…”

miR-221 attenuates the osteogenic differentiation of human annulus fibrosus cells

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SimiRa: A tool to identify coregulation between microRNAs and RNA-binding proteins

“…The top miRNAs showing similar functional categories as TAF15 are miR-590-3p (J D 0.33) and miR-495-3p (J D 0.29). MiR-495 has been shown to inhibit differentiation of human mesenchymal stem cells 58 and mouse embryonic stem cells, 36 pointing toward a similar regulatory loop as for miR-17-5p/miR-20a-5p. MiR-590-3p, on the other hand, is so far not associated with cell cycle progression and is thus a highly interesting candidate for functional studies in combination with TAF15.…”

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