Project description:Smooth muscle guides morphogenesis of epithelia during development of several organs, including the mammalian lung. However, it remains unclear how airway smooth-muscle differentiation is spatiotemporally patterned and whether it originates from distinct mesenchymal progenitors. Using single-cell RNA-sequencing of embryonic mouse lungs, we show that the pulmonary mesenchyme contains a continuum of cell identities, but no distinct progenitors. Transcriptional variability correlates with sub-epithelial and sub-mesothelial mesenchymal compartments that are regulated by Wnt signaling. Live-imaging and tension sensors reveal patterned migratory behaviors and cortical forces in each compartment, and show that sub-epithelial mesenchyme gives rise to airway smooth muscle. Differentiation trajectory reconstruction reveals that cytoskeleton, adhesion, and Wnt signaling pathways are activated early in differentiation. Finally, we show that Wnt activation stimulates the earliest stages of differentiation and induces local accumulation of mesenchymal F-actin, which influences epithelial morphology. Our work provides the first single-cell view of pulmonary mesenchymal patterning during branching morphogenesis.
Project description:To identify proteins phosphorylated by Akt downstream of PI3K-mediated PDGFRalpha signaling, we immunoprecipitated Akt phosphorylation substrates from PDGF-AA-treated primary mouse embryonic palatal mesenchyme (MEPM) lysates and analyzed the peptides by nano LC-MS/MS.
Project description:Lymphangioleiomyomatosis (LAM) is a progressive cystic lung disease caused by tuberous sclerosis complex 1/2 (TSC1/2) gene mutations resulting in activation of the mechanistic target of rapamycin complex 1 (mTORC1). A subset of LAM patients develops pulmonary vascular remodeling and pulmonary hypertension. To model LAM disease, we utilized an mTORC1 gain-of-function mouse model with a Tsc2 knock-out (Tsc2KO) specific to lung mesenchyme (Tbx4LME-CreTsc2fl/fl), similar to the mesenchyme specific genetic alterations seen in human disease. As early as 8 weeks of age, ECs from Tbx4LME-CreTsc2fl/fl mice exhibited marked transcriptomic changes despite absence of morphological changes to the distal lung microvasculature. In contrast, 1 year old Tbx4LME-CreTsc2fl/fl mice spontaneously developed pulmonary vascular remodeling with increased medial thickness. We subsequently performed single cell RNA-sequencing of 1 year old mouse lung and identified paracrine ligands originating from Tsc2KO mesenchyme impacting arterial endothelial cells. These cells contained transcriptionally altered genes including those in pathways associated with blood vessel remodeling, highlighting the pathogenic importance of the mesenchymal-endothelial cell axis.
Project description:Purpose: The aim of this study is to study the expression and function of axon guidance genes in the developing portal tract. Here, the RNA-Sequencing data were collected from purified embryonic mesenchyme at two developmental stages (E16 and E18).
Project description:Identification of genes differentially expressed between epithelium and mesenchyme of embryonic lung. Embryonic lung are harvested from time pregnant mice
Project description:Makorins are an evolutionary conserved family of proteins that contain C3H-type zinc finger modules and a RING E3 ubiquitin ligase domain. Previous analysis indicated a maternal role for Makorin 1 (Mkrn1) in Drosophila embryonic patterning and germ cell specification, but the underlying mechanism has remained elusive. Here, we show that Mkrn1 is specifically required for translational activation of oskar, which encodes a critical regulator of axis specification and germ plasm assembly. We demonstrate that Mkrn1 interacts with poly(A) binding protein (pAbp) and specifically binds osk 3’ UTR adjacent to A-rich sequences. The binding of Mkrn1 to osk 3’UTR occurs in a region that overlaps with Bruno responsive elements (BRE), previously shown to have a dual role in regulating osk translation. We observe an increased association of the translational repressor Bruno (Bru) with osk mRNA upon depletion of Mkrn1, implying that both proteins compete with each other for osk binding. Consistently, reducing Bru dosage is sufficient to partially rescue osk translation and the embryonic lethality associated with Mkrn1 alteration. Thus, we conclude that Mkrn1 controls embryonic patterning and germ cell formation by specifically activating osk translation via displacing Bru from its 3’ UTR
Project description:During organogenesis of the intestine, reciprocal crosstalk between the endodermally-derived epithelium and the underlying mesenchyme is required for regional patterning and proper differentiation. Though both of these tissue layers participate in patterning, the mesenchyme is thought to play a prominant role in the determination of epithelial phenotype during development and in adult life. However, the molecular basis of this instructional dominance is unclear. In fact, surprisingly little is known about the cellular origins of many of the critical signaling molecules and the gene transcriptional events that they impact. Here, we profile genes that are expressed in separated mesenchymal and epithelial compartments of the perinatal mouse intestine. The data indicate that the vast majority of soluble modulators of signaling pathways such as Hedgehog, Bmp, Wnt, Fgf and Igf are expressed predominantly or exclusively by the mesenchyme, accounting for its ability to dominate instructional crosstalk. We also catalog the most highly enriched transcription factors in both compartments and find evidence for a major role for Hnf4alpha and Hnf4 gamma in the regulation of epithelial genes. Finally, we find that while epithelially enriched genes tend to be highly tissue-restricted in their expression, mesenchymally-enriched genes tend to be broadly expressed in multiple tissues. Thus, the unique tissue-specific signature that characterizes the intestinal epithelium is instructed and supported by a mesenchyme that itself expresses genes that are largely non-tissue specific. Keywords: comparative genomic hybridization: epithelium vs. mesenchyme