Mesodermal Expression Research Articles

Nodal signaling is required for mesodermal and ventral but not for dorsal fates in the indirect developing hemichordate, Ptychodera flava.

ABSTRACTNodal signaling plays crucial roles in vertebrate developmental processes such as endoderm and mesoderm formation, and axial patterning events along the anteroposterior, dorsoventral and left-right axes. In echinoderms, Nodal plays an essential role in the establishment of the dorsoventral axis and left-right asymmetry, but not in endoderm or mesoderm induction. In protostomes, Nodal signaling appears to be involved only in establishing left-right asymmetry. Hence, it is hypothesized that Nodal signaling has been co-opted to pattern the dorsoventral axis of deuterostomes and for endoderm, mesoderm formation as well as anteroposterior patterning in chordates. Hemichordata, together with echinoderms, represent the sister taxon to chordates. In this study, we analyze the role of Nodal signaling in the indirect developing hemichordate Ptychodera flava. In particular, we show that during gastrulation nodal transcripts are detected in a ring of cells at the vegetal pole that gives rise to endomesoderm and in the ventral ectoderm at later stages of development. Inhibition of Nodal function disrupts dorsoventral fates and also blocks formation of the larval mesoderm. Interestingly, molecular analysis reveals that only mesodermal, apical and ventral gene expression is affected while the dorsal side appears to be patterned correctly. Taken together, this study suggests that the co-option of Nodal signaling in mesoderm formation and potentially in anteroposterior patterning has occurred prior to the emergence of chordates and that Nodal signaling on the ventral side is uncoupled from BMP signaling on the dorsal side, representing a major difference from the molecular mechanisms of dorsoventral patterning events in echinoderms.

Mesodermal expression of Moz is necessary for cardiac septum development

Ventricular septal defects (VSDs) are the most commonly occurring congenital heart defect. They are regularly associated with complex syndromes, including DiGeorge syndrome and Holt–Oram syndrome, which are characterised by haploinsufficiency for the T-box transcription factors TBX1 and TBX5, respectively. The histone acetyltransferase monocytic leukaemia zinc finger protein, MOZ (MYST3/KAT6A), is required for the expression of the Tbx1 and Tbx5 genes. Homozygous loss of MOZ results in DiGeorge syndrome-like defects including VSD. The Moz gene is expressed in the ectodermal, mesodermal and endodermal aspects of the developing pharyngeal apparatus and heart; however it is unclear in which of these tissues MOZ is required for heart development. The role of MOZ in the activation of Tbx1 would suggest a requirement for MOZ in the mesoderm, because deletion of Tbx1 in the mesoderm causes VSDs. Here, we investigated the tissue-specific requirements for MOZ in the mesoderm. We demonstrate that Mesp1-cre-mediated deletion of Moz results in high penetrance of VSDs and overriding aorta and a significant decrease in MOZ-dependant Tbx1 and Tbx5 expression. Together, our data suggest that the molecular pathogenesis of VSDs in Moz germline mutant mice is due to loss of MOZ-dependant activation of mesodermal Tbx1 and Tbx5 expression.

Mesodermal gene expression during the embryonic and larval development of the articulate brachiopod Terebratalia transversa.

BackgroundBrachiopods undergo radial cleavage, which is distinct from the stereotyped development of closely related spiralian taxa. The mesoderm has been inferred to derive from the archenteron walls following gastrulation, and the primary mesoderm derivative in the larva is a complex musculature. To investigate the specification and differentiation of the mesoderm in the articulate brachiopod Terebratalia transversa, we have identified orthologs of genes involved in mesoderm development in other taxa and investigated their spatial and temporal expression during the embryonic and larval development of T. transversa.ResultsOrthologs of 17 developmental regulatory genes with roles in the development of the mesoderm in other bilaterian animals were found to be expressed in the developing mesoderm of T. transversa. Five genes, Tt.twist, Tt.GATA456, Tt.dachshund, Tt.mPrx, and Tt.NK1, were found to have expression throughout the archenteron wall at the radial gastrula stage, shortly after the initiation of gastrulation. Three additional genes, Tt.Pax1/9, Tt.MyoD, and Tt.Six1/2, showed expression at this stage in only a portion of the archenteron wall. Tt.eya, Tt.FoxC, Tt.FoxF, Tt.Mox, Tt.paraxis, Tt.Limpet, and Tt.Mef2 all showed initial mesodermal expression during later gastrula or early larval stages. At the late larval stage, Tt.dachshund, Tt.Limpet, and Tt.Mef2 showed expression in nearly all mesoderm cells, while all other genes were localized to specific regions of the mesoderm. Tt.FoxD and Tt.noggin both showed expression in the ventral mesoderm at the larval stages, with gastrula expression patterns in the archenteron roof and blastopore lip, respectively.ConclusionsExpression analyses support conserved roles for developmental regulators in the specification and differentiation of the mesoderm during the development of T. transversa. Expression of multiple mesodermal factors in the archenteron wall during gastrulation supports previous morphological observations that this region gives rise to larval mesoderm. Localized expression domains during gastrulation and larval development evidence early regionalization of the mesoderm and provide a basis for hypotheses regarding the molecular regulation underlying the complex system of musculature observed in the larva.Electronic supplementary materialThe online version of this article (doi:10.1186/s13227-015-0004-8) contains supplementary material, which is available to authorized users.

Lmx1b‐Mediated Emx2‐ Associated Regulatory Region Active During Limb Development

Lmx1b is a LIM-homeodomain transcription factor that has dorsally restricted expression in limb mesoderm and is responsible for limb dorsalization. Mice lacking functional Lmx1b exhibit ventral-ven...

Claudin-10 is required for relay of left–right patterning cues from Hensen’s node to the lateral plate mesoderm

Species-specific symmetry-breaking events at the left–right organizer (LRO) drive an evolutionarily-conserved cascade of gene expression in the lateral plate mesoderm that is required for the asymmetric positioning of organs within the body cavity. The mechanisms underlying the transfer of the left and right laterality information from the LRO to the lateral plate mesoderm are poorly understood. Here, we investigate the role of Claudin-10, a tight junction protein, in facilitating the transfer of left–right identity from the LRO to the lateral plate mesoderm. Claudin-10 is asymmetrically expressed on the right side of the chick LRO, Hensen’s node. Gain- and loss-of-function studies demonstrated that right-sided expression of Claudin-10 is essential for normal rightward heart tube looping, the first morphological asymmetry during organogenesis. Manipulation of Claudin-10 expression did not perturb asymmetric gene expression at Hensen’s node, but did disrupt asymmetric gene expression in the lateral plate mesoderm. Bilateral expression of Claudin-10 at Hensen’s node prevented expression of Nodal, Lefty-2 and Pitx2c in the left lateral plate mesoderm, while morpholino knockdown of Claudin-10 inhibited expression of Snail1 in the right lateral plate mesoderm. We also determined that amino acids that are predicted to affect ion selectivity and protein interactions that bridge Claudin-10 to the actin cytoskeleton were essential for its left–right patterning function. Collectively, our data demonstrate a novel role for Claudin-10 during the transmission of laterality information from Hensen’s node to both the left and right sides of the embryo and demonstrate that tight junctions have a critical role during the relay of left–right patterning cues from Hensen’s node to the lateral plate mesoderm.

The myogenic repressor gene Holes in muscles is a direct transcriptional target of Twist and Tinman in the Drosophila embryonic mesoderm

Understanding the regulatory circuitry controlling myogenesis is critical to understanding developmental mechanisms and developmentally-derived diseases. We analyzed the transcriptional regulation of a Drosophila myogenic repressor gene, Holes in muscles (Him). Previously, Him was shown to inhibit Myocyte enhancer factor-2 (MEF2) activity, and is expressed in myoblasts but not differentiating myotubes. We demonstrate that different phases of Him embryonic expression arises through the actions of different enhancers, and we characterize the enhancer required for its early mesoderm expression. This Him early mesoderm enhancer contains two conserved binding sites for the basic helix-loop-helix regulator Twist, and one binding site for the NK homeodomain protein Tinman. The sites for both proteins are required for enhancer activity in early embryos. Twist and Tinman activate the enhancer in tissue culture assays, and ectopic expression of either factor is sufficient to direct ectopic expression of a Him–lacZ reporter, or of the endogenous Him gene. Moreover, sustained expression of twist in the mesoderm up-regulates mesodermal Him expression in late embryos. Our findings provide a model to define mechanistically how Twist can both promotes myogenesis through direct activation of Mef2, and can place a brake on myogenesis, through direct activation of Him.

ProNodal acts via FGFR3 to govern duration of Shh expression in the prechordal mesoderm.

The secreted glycoprotein sonic hedgehog (Shh) is expressed in the prechordal mesoderm, where it plays a crucial role in induction and patterning of the ventral forebrain. Currently little is known about how Shh is regulated in prechordal tissue. Here we show that in the embryonic chick, Shh is expressed transiently in prechordal mesoderm, and is governed by unprocessed Nodal. Exposure of prechordal mesoderm microcultures to Nodal-conditioned medium, the Nodal inhibitor CerS, or to an ALK4/5/7 inhibitor reveals that Nodal is required to maintain both Shh and Gsc expression, but whereas Gsc is largely maintained through canonical signalling, Nodal signals through a non-canonical route to maintain Shh. Further, Shh expression can be maintained by a recombinant Nodal cleavage mutant, proNodal, but not by purified mature Nodal. A number of lines of evidence suggest that proNodal acts via FGFR3. ProNodal and FGFR3 co-immunoprecipitate and proNodal increases FGFR3 tyrosine phosphorylation. In microcultures, soluble FGFR3 abolishes Shh without affecting Gsc expression. Further, prechordal mesoderm cells in which Fgfr3 expression is reduced by Fgfr3 siRNA fail to bind to proNodal. Finally, targeted electroporation of Fgfr3 siRNA to prechordal mesoderm in vivo results in premature Shh downregulation without affecting Gsc. We report an inverse correlation between proNodal-FGFR3 signalling and pSmad1/5/8, and show that proNodal-FGFR3 signalling antagonises BMP-mediated pSmad1/5/8 signalling, which is poised to downregulate Shh. Our studies suggest that proNodal/FGFR3 signalling governs Shh duration by repressing canonical BMP signalling, and that local BMPs rapidly silence Shh once endogenous Nodal-FGFR3 signalling is downregulated.

Zebrafish Wnt9b Patterns the First Pharyngeal Arch into D-I-V Domains and Promotes Anterior-Medial Outgrowth

Disrupted morphogenesis and growth of the embryonic maxillary jaw lead to oral facial clefting in humans (OFC) and result in an incompletely formed secondary mouth and face. A requirement for Wnt signaling and Wnt9b in particular are postulated in the etiology of OFC from association studies in humans and from animal models. Loss of murine Wnt9b leads to reduced upper jaw (maxillary) outgrowth and OFC, though the signaling architecture leading to this phenotype is poorly understood. Previous murine Wnt9b studies largely overlooked cranial neural crest cell (CNCC) patterning events and instead focused on later events during fusion of facial prominences. Using zebrafish and a morpholino-mediated knockdown approach, we demonstrate functional requirements for Wnt9b signaling during two crucial stages of facial development: 1) CNCC patterning into Dorsal-Intermediate-Ventral (D-I-V) domains; and 2) facial outgrowth during the primary to secondary mouth transition (PM to SM). Zebrafish embryos deficient for Wnt9b (Wnt9b morphants) exhibit an open bite with fused jaw joints as well as a flat face. Open bite and jaw joint fusion in Wnt9b morphants phenocopies characteristics of edn1 pathway is mutant zebrafish with disrupted D-I-V patterning of CNCC. Expression studies show Wnt9b morphants exhibit perturbed expression of edn1 signaling targets including dlx2a, jag1b, and msxe, consistent with disrupted CNCC patterning. Wnt9b morphant upper jaws have stunted outgrowth reminiscent of murine Wnt9b mutants and Wnt9b morphant skulls phenocopy the broad class of foreshortened skull zebrafish mutants known as hammerheads. Wnt9b morphants show upregulated expression of pitx2a after the opening of the primary mouth and disrupted expression of Wnt5b which is consistent with disrupted chondrocyte stacking. Strong upregulation of dorsal mesodermal frzb expression in the prechordal plate of Wnt9b morphants suggests a role for Wnt9b in primary mouth induction or maintenance. Collectively these results argue that Wnt9b has a much earlier developmental requirement. This work draws attention to potential vertebrate homologies that pattern CNCC and facial outgrowth and therefore calls for a reexamination of Wnt9b’s role during mammalian craniofacial development.

Dachsous1b cadherin regulates actin and microtubule cytoskeleton during early zebrafish embryogenesis.

Dachsous (Dchs), an atypical cadherin, is an evolutionarily conserved regulator of planar cell polarity, tissue size and cell adhesion. In humans, DCHS1 mutations cause pleiotropic Van Maldergem syndrome. Here, we report that mutations in zebrafish dchs1b and dchs2 disrupt several aspects of embryogenesis, including gastrulation. Unexpectedly, maternal zygotic (MZ) dchs1b mutants show defects in the earliest developmental stage, egg activation, including abnormal cortical granule exocytosis (CGE), cytoplasmic segregation, cleavages and maternal mRNA translocation, in transcriptionally quiescent embryos. Later, MZdchs1b mutants exhibit altered dorsal organizer and mesendodermal gene expression, due to impaired dorsal determinant transport and Nodal signaling. Mechanistically, MZdchs1b phenotypes can be explained in part by defective actin or microtubule networks, which appear bundled in mutants. Accordingly, disruption of actin cytoskeleton in wild-type embryos phenocopied MZdchs1b mutant defects in cytoplasmic segregation and CGE, whereas interfering with microtubules in wild-type embryos impaired dorsal organizer and mesodermal gene expression without perceptible earlier phenotypes. Moreover, the bundled microtubule phenotype was partially rescued by expressing either full-length Dchs1b or its intracellular domain, suggesting that Dchs1b affects microtubules and some developmental processes independent of its known ligand Fat. Our results indicate novel roles for vertebrate Dchs in actin and microtubule cytoskeleton regulation in the unanticipated context of the single-celled embryo.

Abstract 20699: GMP-Grade Cardiomyocyte Differentiation Media System for Pluripotent Stem Cells in Basic and Translational Research

Objective: Current protocols for differentiating pluripotent stem cells (PSCs) have led to heterogeneous results, varying purity levels, and long lead times for generation of cardiomyocytes. We hypothesized that a simplified and rapid cardiomyocyte differentiation media system can be developed in a scalable workflow to enable generation of large numbers of consistent, spontaneously active cardiomyocytes that could be used in basic and translational research. Methods: High quality PSCs were maintained under xenofree, feeder-free culture conditions. At time of passaging, PSC were dissociated with 0.5 mM EDTA, seeded on 1:100 Geltrex © -coated surface as small clusters at ~0.5 to 1 x 10 5 /well of a 12-well plate and maintained for four days under serum-free condition. After reaching target confluence of ~60 to 80%, an induction media was added for two days followed by addition of a second induction media for two days. After the induction step, the media was replaced with maintenance media and re-fed every other day for up to five weeks. PSC-derived cardiomyocytes were analyzed by morphology, gene expression, flow cytometry, immunocytochemistry and multi-electrode array (MEA). Results: We observed individual beating cells by Day 7 and contracting syncytia by Day 10. An over 100 fold increase in cell number was noted from the time of plating to generation of contracting syncytia of cardiomyocytes. Quantitative flow cytometry detected populations of troponin T type 2 (TNNT2)-immunoreactive cells that reached as high as 96.6%. Number of TNNT2-positive cells dropped by 20% when induced at 90% versus 60% confluency. PCR studies confirmed expression of mesoderm (T, MIXL1, MESP1), cardiac mesoderm (ISL1, GATA4, MEF2C) and mature cardiomyocyte genes (NKX2.5, TNNT2, MYH6). Immunocytochemistry studies verified expression of cardiac markers NKX2.5,GATA4, MEF2C, TNNT2 and MYH6. Initial MEA studies corroborated the presence of electrically active cells. Conclusions: We conclude that a simplified complete differentiation media system could serve as a standardized culture system for generating large numbers of consistent, spontaneously active cardiomyocytes for basic and translational research studies.

Identification of novel retinoic acid target genes.

Retinoic acid is required for diverse ontogenic processes and as such identification of the genes and pathways affected by retinoic acid is critical to understanding these pleiotropic effects. The presomitic mesoderm of the E8.5 mouse embryo is composed of undifferentiated cells that are depleted of retinoic acid, yet are competent to respond to the retinoid signal. We have exploited these properties to use this tissue to identify novel retinoic acid-responsive genes, including candidate target genes, by treating E8.5 embryos with retinoic acid and assessing changes in gene expression in the presomitic mesoderm by microarray analysis. This exercise yielded a cohort of genes that were differentially expressed in response to exogenous retinoic acid exposure. Among these were a number of previously characterized retinoic acid targets, validating this approach. In addition, we recovered a number of novel candidate target genes which were confirmed as retinoic acid-responsive by independent analysis. Chromatin immunoprecipitation assays revealed retinoic acid receptor occupancy of the promoters of certain of these genes. We further confirmed direct retinoic acid regulation of the F11r gene, a new RA target, using tissue culture models. Our results reveal a significant number of potential RA targets implicated in embryonic development and offer a novel in vivo system for better understanding of retinoid-dependent transcription.

Mesodermal expression of integrin α5β1 regulates neural crest development and cardiovascular morphogenesis.

Integrin α5-null embryos die in mid-gestation from severe defects in cardiovascular morphogenesis, which stem from defective development of the neural crest, heart and vasculature. To investigate the role of integrin α5β1 in cardiovascular development, we used the Mesp1(Cre) knock-in strain of mice to ablate integrin α5 in the anterior mesoderm, which gives rise to all of the cardiac and many of the vascular and muscle lineages in the anterior portion of the embryo. Surprisingly, we found that mutant embryos displayed numerous defects related to the abnormal development of the neural crest such as cleft palate, ventricular septal defect, abnormal development of hypoglossal nerves, and defective remodeling of the aortic arch arteries. We found that defects in arch artery remodeling stem from the role of mesodermal integrin α5β1 in neural crest proliferation and differentiation into vascular smooth muscle cells, while proliferation of pharyngeal mesoderm and differentiation of mesodermal derivatives into vascular smooth muscle cells was not defective. Taken together our studies demonstrate a requisite role for mesodermal integrin α5β1 in signaling between the mesoderm and the neural crest, thereby regulating neural crest-dependent morphogenesis of essential embryonic structures.

Derivation and Expansion of PAX7-Positive Muscle Progenitors from Human and Mouse Embryonic Stem Cells

SummaryCell therapies treating pathological muscle atrophy or damage requires an adequate quantity of muscle progenitor cells (MPCs) not currently attainable from adult donors. Here, we generate cultures of approximately 90% skeletal myogenic cells by treating human embryonic stem cells (ESCs) with the GSK3 inhibitor CHIR99021 followed by FGF2 and N2 supplements. Gene expression analysis identified progressive expression of mesoderm, somite, dermomyotome, and myotome markers, following patterns of embryonic myogenesis. CHIR99021 enhanced transcript levels of the pan-mesoderm gene T and paraxial-mesoderm genes MSGN1 and TBX6; immunofluorescence confirmed that 91% ± 6% of cells expressed T immediately following treatment. By 7 weeks, 47% ± 3% of cells were MYH+ve myocytes/myotubes surrounded by a 43% ± 4% population of PAX7+ve MPCs, indicating 90% of cells had achieved myogenic identity without any cell sorting. Treatment of mouse ESCs with these factors resulted in similar enhancements of myogenesis. These studies establish a foundation for serum-free and chemically defined monolayer skeletal myogenesis of ESCs.

Distinct regulation of the anterior and posterior myeloperoxidase expression by Etv2 and Gata1 during primitive Granulopoiesis in zebrafish

Neutrophilic granulocytes are the most abundant type of myeloid cells and form an essential part of the innate immune system. In vertebrates the first neutrophils are thought to originate during primitive hematopoiesis, which precedes hematopoietic stem cell formation. In zebrafish embryos, it has been suggested that primitive neutrophils may originate in two distinct sites, the anterior (ALPM) and posterior lateral plate mesoderm (PLPM). An ETS-family transcription factor Etsrp/Etv2/ER71 has been implicated in vasculogenesis and hematopoiesis in multiple vertebrates. However, its role during neutrophil development is not well understood. Here we demonstrate using zebrafish embryos that Etv2 has a specific cell-autonomous function during primitive neutropoiesis in the anterior lateral plate mesoderm (ALPM) but has little effect on erythropoiesis or the posterior lateral plate mesoderm (PLPM) expression of neutrophil marker myeloperoxidase mpo/mpx. Our results argue that ALPM-derived neutrophils originate from etv2-expressing cells which downregulate etv2 during neutropoiesis. We further show that Scl functions downstream of Etv2 in anterior neutropoiesis. Additionally, we demonstrate that mpx expression within the PLPM overlaps with gata1 expression, potentially marking the cells with a dual myelo-erythroid potential. Intriguingly, initiation of mpx expression in the PLPM is dependent on gata1 but not etv2 function. Our results demonstrate that mpx expression is controlled differently in the ALPM and PLPM regions and describe novel roles for etv2 and gata1 during primitive neutropoiesis.

Hox expression in the direct‐type developing sand dollar Peronella japonica

Echinoderms are a curious group of deuterostomes that forms a clade with hemichordates but has a pentameral body plan. Hox complex plays a pivotal role in axial patterning in bilaterians and often occurs in a cluster on the chromosome. In contrast to hemichordates with an organized Hox cluster, the sea urchin Strongylocentrotus purpuratus has a Hox cluster with an atypical organization. However, the current data on hox expression in sea urchin rudiments are fragmentary. We report a comprehensive examination of hox expression in a sand dollar echinoid. Nine hox genes are expressed in the adult rudiment, which are classified into two groups, but hox11/13b belongs to both: one with linear expression in the coelomic mesoderm and another with radial expression around the adult mouth. The linear genes may endow the coelom/mesentery with axial information to direct postmetamorphic transformation of the digestive tract, whereas the radial genes developmentally correlate with the morphological novelties of echinoderms and/or sea urchins. Recruitment of the radial genes except hox11/13b appears to be accompanied by the loss of ancestral/axial roles. This in toto co-option of the hox genes provides insight into the molecular mechanisms underlying the evolution of echinoderms from a bilateral ancestor.

Multiple regulatory safeguards confine the expression of the GATA factor Serpent to the hemocyte primordium within the Drosophila mesoderm

serpent (srp) encodes a GATA-factor that controls various aspects of embryogenesis in Drosophila, such as fatbody development, gut differentiation and hematopoiesis. During hematopoiesis, srp expression is required in the embryonic head mesoderm and the larval lymph gland, the two known hematopoietic tissues of Drosophila, to obtain mature hemocytes. srp expression in the hemocyte primordium is known to depend on snail and buttonhead, but the regulatory complexity that defines the primordium has not been addressed yet. Here, we find that srp is sufficient to transform trunk mesoderm into hemocytes. We identify two disjoint cis-regulatory modules that direct the early expression in the hemocyte primordium and the late expression in mature hemocytes and lymph gland, respectively. During embryonic hematopoiesis, a combination of snail, buttonhead, empty spiracles and even-skipped confines the mesodermal srp expression to the head region. This restriction to the head mesoderm is crucial as ectopic srp in mesodermal precursors interferes with the development of mesodermal derivates and promotes hemocytes and fatbody development. Thus, several genes work in a combined fashion to restrain early srp expression to the head mesoderm in order to prevent expansion of the hemocyte primordium.

In This Issue

HomeCirculation ResearchVol. 113, No. 10In This Issue Free AccessResearch ArticlePDF/EPUBAboutView PDFView EPUBSections ToolsAdd to favoritesDownload citationsTrack citationsPermissions ShareShare onFacebookTwitterLinked InMendeleyReddit Jump toFree AccessResearch ArticlePDF/EPUBIn This Issue Originally published25 Oct 2013https://doi.org/10.1161/01.res.0000437011.92592.d9Circulation Research. 2013;113:1097Regulation of VSMC Function by miR-663 (p 1117)Download figureDownload PowerPointLi et al identify a new microRNA that regulates phenotype switching in human vascular smooth muscle cells.Fully differentiated vascular smooth muscle cells (VSMCs) in situ are generally non-proliferative, but after a vessel injury these cells dedifferentiate, proliferate and migrate. It is believed that such phenotype switching is an essential part of the healing process, but it can also lead to vessel narrowing, restenosis and other associated problems. Among the factors controlling this phenotype switch are microRNAs (miRs)—that suppress the expression of target mRNAs. While a number of such miRs have been identified in animal VSMCs, little is known about miRs in human VSMCs. Li and colleagues thus analyzed changes in miR expression in human VSMCs following induction of the phenotype switch. They found that miR-663 in particular, was dramatically downregulated. They also found that overexpression of miR-663 in VSMCs induced the expression of differentiation markers, whereas blocking miR-663 expression increased both proliferation and migration. MiR-663 targeted the mRNA of transcription factor JunB, suppressing its expression in human VSMCs. Although miR-663 is not conserved between humans and mice, its target sequence in the JunB mRNA is, and the team found that transfection of miR-663 into mice could suppress JunB expression and could block vessel narrowing. Boosting miR-663 activity or suppressing JunB might thus be strategies for preventing restenosis.Somatic Mesoderm and the Proepicardium (p 1128)Download figureDownload PowerPointSchlueter and Brand investigate the origins of the heart’s proepicardium and discover that some of the cells derive from somatic mesoderm.During embryogenesis, the proepicardium gives rise to epicardial cells as well as cells of the coronary vasculature and cardiac fibroblasts. But the origin of the proepicardium itself is unclear. In addition, it is also unclear how the proepicardium develops its characteristic left-right asymmetry—though the growth factor FGF8 and transcription factor SNAI1 are both known to be involved. Schlueter and Brand now show that a known target gene of SNAI1, a transcription factor called TWIST1, is asymmetrically expressed in lateral plate mesoderm of the early chick embryo, being highly expressed on the right-hand side. Furthermore, the team showed that these cells give rise to a population of proepicardial cells. They found that loss of TWIST1 expression in the right-hand mesoderm led to malformation of the proepicardium, while overexpression of TWIST in the left-hand mesoderm caused these cells to adopt characteristics of the right-hand proepicardium. The findings could have implications not only for understanding cardiac development, but also for understanding disease processes like myocardial fibrosis and for developing approaches for regenerative therapies.MicroRNAs in Cardiac Fibroblasts (p 1138)Download figureDownload PowerPointAbonnenc et al confirm the role of microRNAs in extracellular matrix secretion by cardiac fibroblasts.Changes to the composition of the extracellular matrix (ECM) contribute to the development of fibrosis, a characteristic feature of myocardial remodeling in a number of cardiac diseases. Previous studies have identified several microRNAs (miRs) that regulate cardiac fibrosis, including miR-30c and miR-29b. But a systematic analysis of the targets of these miRs is lacking. Rather than searching for messenger RNA targets of miR-30c and 29b—via computer prediction software or microarray analyses—Abonnenc and colleagues opted to search for proteins secreted by cardiac fibroblasts in response to the two miRs, thus allowing for the identification of both direct and indirect targets. The team used mass spectrometry to study the secretomes of primary mouse cardiac fibroblasts in which the two miRs were either over-expressed or inhibited. While miR-29b suppressed the expression of ECM and fibrosis proteins, including collagens, matrix metalloproteases, leukemia inhibitory factor (LIF) and insulin-like growth factor-1 (IGF-1), miR-30c had little effect on these proteins. The proteomic approach identified previously predicted and novel targets for the two miRs and, say the authors, offers a valuable resource for future studies into cardiac fibrosis. Previous Back to top Next FiguresReferencesRelatedDetails October 25, 2013Vol 113, Issue 10 Advertisement Article InformationMetrics © 2013 American Heart Association, Inc.https://doi.org/10.1161/01.res.0000437011.92592.d9 Originally publishedOctober 25, 2013 PDF download Advertisement

Regulation of mesenchymal‐to‐epithelial transition by PARAXIS during somitogenesis

Dynamic alterations in cell shape, migration, and adhesion play a central role in tissue morphogenesis during embryonic development and congenital disease. The mesenchymal-to-epithelial transition that occurs during vertebrate somitogenesis is required for proper patterning of the axial musculoskeletal system. Somitic MET is initiated in the presomitic mesoderm by PARAXIS-dependent changes in cell adhesion, cell polarity, and the composition of the extracellular matrix. However, the target genes downstream of the transcription factor PARAXIS remain poorly described. A genome-wide comparison of gene expression in the anterior presomitic mesoderm and newly formed somites of Paraxis(-/-) embryos resulted in a set of deregulated genes enriched for factors associated with extracellular matrix and cytoskeletal organization and cell-cell and cell-ECM adhesion. The greatest change in expression was seen in fibroblast activation protein alpha (Fap), encoding a dipeptidyl peptidase capable of increasing fibronectin and collagen fiber organization in extracellular matrix. Further, downstream genes in the Wnt and Notch signaling pathways were downregulated, predicting that PARAXIS participates in positive feedback loops in both pathways. These data demonstrate that PARAXIS initiates and stabilizes somite epithelialization by integrating signals from multiple pathways to control the reorganization of the ECM, cytoskeleton, and adhesion junctions during MET.

Correlation of mesodermal and pluripotency gene expression during in vitro differentiation of human embryonic stem cells

Human embryonic stem cells (hESCs) have capacities to self-renew and differentiate into three germ layers, therefore, genes involved in the fate of stem cells are endogenously regulated to maintain the balance between pluripotency and initiation of differentiation. In this study, we investigated the stage-specific expression of pluripotency- and differentiation-related genes during differentiation into cardiomyocytes from hESCs.

Homothorax plays autonomous and nonautonomous roles in proximodistal axis formation and migration of the Drosophila renal tubules

The Drosophila Malpighian tubules (MpTs) serve as a functional equivalent of the mammalian renal tubules. The MpTs are composed of two pairs of epithelial tubes that bud from the midgut-hindgut boundary during embryogenesis. The MpT primordia grow, elongate and migrate through the body cavity to assume their final position and shape. The stereotypic pattern of MpT migration is regulated by multiple intrinsic and extrinsic signals, many of which are still obscure. In this work, we implicate the TALE-class homeoprotein Homothorax (Hth) in MpT patterning. We show that in the absence of Hth the tubules fail to rearrange and migrate. Hth plays both autonomous and nonautonomous roles in this developmental process. Within the tubules Hth is required for convergent extension and for defining distal versus proximal cell identities. The difference between distal and proximal cell identities seems to be required for proper formation of the leading loop. Outside the tubules, wide-range mesodermal expression of Hth is required for directing anterior migration. The nonautonomous effects of Hth on MpT migration can be partially attributed to its effects on homeotic determination along the anterior posterior axis of the embryo and to its effects on stellate cell (SC) incorporation into the MpT.