Cell cycle regulators control mesoderm specification in human pluripotent stem cells.

The mesoderm is one of the three germ layers produced during gastrulation from which muscle, bones, kidneys, and the cardiovascular system originate. Understanding the mechanisms that control mesoderm specification could inform many applications, including the development of regenerative medicine therapies to manage diseases affecting these tissues. Here, we used human pluripotent stem cells to investigate the role of cell cycle in mesoderm formation. To this end, using small molecules or conditional gene knockdown, we inhibited proteins controlling G1 and G2/M cell cycle phases during the differentiation of human pluripotent stem cells into lateral plate, cardiac, and presomitic mesoderm. These loss-of-function experiments revealed that regulators of the G1 phase, such as cyclin-dependent kinases and pRb (retinoblastoma protein), are necessary for efficient mesoderm formation in a context-dependent manner. Further investigations disclosed that inhibition of the G2/M regulator cyclin-dependent kinase 1 decreases BMP (bone morphogenetic protein) signaling activity specifically during lateral plate mesoderm formation while reducing fibroblast growth factor/extracellular signaling-regulated kinase 1/2 activity in all mesoderm subtypes. Taken together, our findings reveal that cell cycle regulators direct mesoderm formation by controlling the activity of key developmental pathways.This work was supported by the Wellcome Trust PhD program (PSAG/048 to L.Y.); the European Research Council advanced grant New-Chol (ERC: 741707 to L.V. and R.A.G), a BHF Senior Research Fellowship (FS/13/29/30024 to S.S.), a core support grant from the Wellcome and Medical Research Council to the Wellcome – Medical Research Council Cambridge Stem Cell Institute (PSAG028) and a core support grant from the Wellcome to the Wellcome Sanger Institute (WT206194)

The SMAD2/3 interactome reveals that TGF beta controls m(6)A mRNA methylation in pluripotency

The TGFβ pathway has essential roles in embryonic development, organ homeostasis, tissue repair and disease1,2. These diverse effects are mediated through the intracellular effectors SMAD2 and SMAD3 (hereafter SMAD2/3), whose canonical function is to control the activity of target genes by interacting with transcriptional regulators3. Therefore, a complete description of the factors that interact with SMAD2/3 in a given cell type would have broad implications for many areas of cell biology. Here we describe the interactome of SMAD2/3 in human pluripotent stem cells. This analysis reveals that SMAD2/3 is involved in multiple molecular processes in addition to its role in transcription. In particular, we identify a functional interaction with the METTL3–METTL14–WTAP complex, which mediates the conversion of adenosine to N6-methyladenosine (m6A) on RNA4. We show that SMAD2/3 promotes binding of the m6A methyltransferase complex to a subset of transcripts involved in early cell fate decisions. This mechanism destabilizes specific SMAD2/3 transcriptional targets, including the pluripotency factor gene NANOG, priming them for rapid downregulation upon differentiation to enable timely exit from pluripotency. Collectively, these findings reveal the mechanism by which extracellular signalling can induce rapid cellular responses through regulation of the epitranscriptome. These aspects of TGFβ signalling could have far-reaching implications in many other cell types and in diseases such as cancer

Cholangiocyte organoids can repair bile ducts after transplantation in the human liver.

Organoid technology holds great promise for regenerative medicine but has not yet been applied to humans. We address this challenge using cholangiocyte organoids in the context of cholangiopathies, which represent a key reason for liver transplantation. Using single-cell RNA sequencing, we show that primary human cholangiocytes display transcriptional diversity that is lost in organoid culture. However, cholangiocyte organoids remain plastic and resume their in vivo signatures when transplanted back in the biliary tree. We then utilize a model of cell engraftment in human livers undergoing ex vivo normothermic perfusion to demonstrate that this property allows extrahepatic organoids to repair human intrahepatic ducts after transplantation. Our results provide proof of principle that cholangiocyte organoids can be used to repair human biliary epithelium

Carbon sink removal: Increased photosynthetic production of lactic acid by Synechocystis sp. PCC6803 in a glycogen storage mutant

Deletion of pathways for carbon-storage in the cyanobacterium Synechocystis sp. PCC6803 has been suggested as a strategy to increase the size of the available pyruvate pool for the production of (heterologous) chemical commodities. Here we show that deletion of the pathway for glycogen synthesis leads to a twofold increased lactate production rate, under nitrogen-limited conditions, whereas impairment of polyhydroxybutyrate synthesis does not

Candidate gene and reverse genetics approaches for the analysis of development in barley

Research in our group focuses on the genetic and molecular dissection of barley development as a mean to identify useful genes for the manipulation of plant architecture. Although progress has been made in rice and maize, the molecular bases of development are largely unknown in Triticeae species. Traits under analysis include tillering, spike morphology, and development of leaves and floral bracts. In order to identify and characterize genes involved in these processes, we have been exploiting a wide collection of developmental mutants, combining candidate gene (CG) and reverse genetics approaches. CGs for inflorescence branching and foliar development were identified based on synteny between the barley and rice genomes. CGs for meristem function and floret development were identified through molecular approaches, using as a starting point the Hooded (K) mutant. Support for specific CGs was provided by phenotypic similarities between rice and barley mutants. Validation of barley CGs is based on a range of complementarity between forward and reverse genetics approaches