Project description:Morphogen signalling forms an activity gradient and instructs cell identities in a signalling strength-dependent manner to pattern developing tissues. However, developing tissues also undergo dynamic morphogenesis, which may produce cells with unfit morphogen signalling and consequent noisy morphogen gradient. Here we show that a cell competition-related system corrects such noisy morphogen gradients. Zebrafish imaging analyses of the Wnt/β-catenin signalling gradient, which acts as a morphogen to establish embryonic anterior-posterior patterning, revealed that unfit cells with abnormal Wnt/β-catenin activity spontaneously appear and produce noise in the gradient. Communication between unfit and neighbouring fit cells via cadherin proteins stimulates apoptosis of the unfit cells by activating Smad signalling and reactive oxygen species production. This unfit cell elimination is required for proper Wnt/β-catenin gradient formation and consequent anterior-posterior patterning. Because this gradient controls patterning not only in the embryo but also in adult tissues, this system may support tissue robustness and disease prevention.
Project description:This randomized phase I/II clinical trial is studying the side effects and best dose of gamma-secretase/notch signalling pathway inhibitor RO4929097 when given together with vismodegib and to see how well they work in treating patients with advanced or metastatic sarcoma. Vismodegib may slow the growth of tumor cells. Gamma-secretase/notch signalling pathway inhibitor RO4929097 may stop the growth of tumor cells by blocking some of the enzymes needed for cell growth. Giving vismodegib together with gamma-secretase/notch signalling pathway inhibitor RO4929097 may be an effective treatment for sarcoma.
Project description:Activin/Nodal signalling is necessary to maintain pluripotency of human Embryonic Stem Cells (hESCs) and to induce their differentiation towards endoderm. However, the mechanisms by which Activin/Nodal signalling achieves these opposite functions remain unclear. To unravel these mechanisms, we examined the transcriptional network controlled in hESCs by Smad2 and Smad3 which represent the direct effectors of Activin/Nodal signalling. These analyses reveal that Smad2/3 participate in the control of the core transcriptional network characterising pluripotency which includes Oct-4, Nanog, FoxD3, Dppa4, Tert, Myc and UTF-1. In addition, similar experiments performed on endoderm cells confirm that a broad part of the transcriptional network directing differentiation is downstream of Smad2/3. Therefore, Activin/Nodal signalling appears to control divergent transcriptional networks in hESCs and in endoderm. Importantly, we observed an overlap between the transcriptional network downstream of Nanog and Smad2/3 in hESCs while functional studies showed that both factors cooperate to control the expression of pluripotency genes. Therefore, the effect of Activin/Nodal signalling on pluripotency and differentiation could be dictated by tissue specific Smad2/3 partners such as Nanog, explaining the mechanisms by which signalling pathways can orchestrate divergent cell fate decisions. Identification of Smad2/3 binding sites in pluripotent hESCs. 5 ChIP-Seq samples including 1 input control sample and 4 ChIP samples (two conditions x two replicates).
Project description:Drugs that target pre-mRNA splicing hold great therapeutic potential, but the mechanistic understanding of how these drugs function is limited. Here we introduce a biophysical modeling framework that can quantitatively describe the sequence-specific and concentration-dependent behavior of splice-modifying drugs. Using massively parallel splicing assays, RNA-seq experiments, and precision dose-response curves, we apply this framework to two drugs, risdiplam and branaplam, developed for treating spinal muscular atrophy. The results quantitatively define the specificities of risdiplam and branaplam for 5’ splice site sequences, suggest that branaplam recognizes 5’ splice sites via two distinct interaction modes, and disprove the prevailing two-site hypothesis for risdiplam activity at SMN2 exon 7. The results also show, more generally, that single-drug cooperativity and multi-drug synergy are widespread among splice-modifying drugs. Our biophysical modeling approach thus clarifies the mechanisms of existing splice-modifying treatments and provides a quantitative basis for the rational development of new therapies.
Project description:Hypothesis and proof of concept:
Our hypothesis is that increase in cellular iron import proteins (TfR1, DMT1) occur early in the adenoma-carcinoma sequence through mutations in APC and lead to cellular iron loading. As demonstrated in our previous work the effects of this iron loading is to mediate increased Wnt signalling resulting in c-myc induction. This in turn serves to increase the expression of iron import proteins (TfR1, DMT1) and decrease the expression of iron export (ferroportin [FPN]) and storage (ferritin) proteins. Such a hypothesis explains how Wnt signalling controls iron metabolism and ensures that there is adequate cellular iron for ATP generation and cellular proliferation.
Experimental design:
To test such a hypothesis we aim to prospectively collect the following colorectal tissue from patients attending for colonoscopy:
1. Normal colonic mucosa in patients with no colorectal pathology (n = 30)
2. Polyps and matched normal colon (n = 30)
3. Colorectal cancers and matched normal colon (n = 30)
We also intend to collect serum and urine from the following patient groups:
4. Normal colonoscopy (n = 30)
5. Colorectal adenomas (n = 30)
6. Colorectal cancers (n = 30)
Primary outcome(s): Measured at baseline, using the expression of proteins in the tissue and serum to detect the cellular and systemic iron transport proteins. The techniques used will include mass spectrometry, western blotting, real time PCR and immunohistochemistry.
Project description:Understanding the molecular mechanisms underlying frontotemporal dementia (FTD) is essential for the development of successful therapies. Here, we present Phase 1 of a multi-omics, multi-model data resource for FTD research which will allows in-depth molecular research into these mechanisms. We have integrated and analysed data from the frontal lobe of FTD patients with mutations in MAPT, GRN and C9orf72 and detected common and distinct dysregulated cellular pathways. Our results highlight that excitatory neurons are the most vulnerable neuronal cell type and that vascular aberrations are a common hallmark in FTD. Via integration of multi-omics data, we detected several transcription factors and pathways which regulate the strong neuroinflammation observed in FTD-GRN. Finally, using small RNA-seq data and verification experiments in cellular models, we identified several up-regulated miRNAs that inhibit cellular trafficking pathways in FTD and lead to microglial activation. In this work we shed light on novel mechanistic and pathophysiological hallmarks of FTD. In addition, we believe that this comprehensive, multi-omics data resource will further mechanistic FTD research by the community.
Project description:We report the transcritional targets of transcription factor downstream of insulin signalling. We knocked down transcription factors under reduced insulin signalling in Caenorhabditis elegans and did transcriptom analysis. This study identified target genes interaction complexity in analogous genetic and experimental conditions. It aid in gaining insight into the new molecular mechanisms downstream of rIIS
Project description:In this study, we have investigated the molecular basis of Shh signalling during development of the secondary palate and how CNCC patterning and fate is influenced by the Shh signalling network. Using a gain-of-function mouse model to activate Smoothened (R26SmoM2) signalling in the palatal mesenchyme (Osr2-IresCre), we demonstrate ectopic Hh-Smo signalling results in fully penetrant cleft palate, disrupted oral-nasal patterning and defective palatine bone formation. We show that a series of Fox transcription factors, including the novel direct target Foxl1, function downstream of Hh signalling in the secondary palate. Furthermore, we demonstrate that Wnt/BMP antagonists, in particular Sostdc1, are positively regulated by Hh signalling, concomitant with down-regulation of key regulators of osteogenesis and BMP signalling effectors. Microarray analysis was performed on excised palatal shelves from Osr2-IresCre+/- (wild-type) and Osr2-IresCre;Smo+/M2 (mutant) embryos at embryonic day (E)13.5. Osr2-IresCre (PMID:17941042) and R26SmoM2 (PMID:15107405) mice have been described previously.