The logistics of Wnt production and delivery.

Wnts are secreted proteins that control stem cell maintenance, cell fate decisions, and growth during development and adult homeostasis. Wnts carry a post-translational modification not seen in any other secreted protein: during biosynthesis, they are appended with a palmitoleoyl moiety that is required for signaling but also impairs solubility and hence diffusion in the extracellular space. In some contexts, Wnts act only in a juxtacrine manner but there are also instances of long range action. Several proteins and processes ensure that active Wnts reach the appropriate target cells. Some, like Porcupine, Wntless, and Notum are dedicated to Wnt function; we describe their activities in molecular detail. We also outline how the cell infrastructure (secretory, endocytic, and retromer pathways) contribute to the progression of Wnts from production to delivery. We then address how Wnts spread in the extracellular space and form a signaling gradient despite carrying a hydrophobic moiety. We highlight particularly the role of lipid-binding Wnt interactors and heparan sulfate proteoglycans. Finally, we briefly discuss how evolution might have led to the emergence of this unusual signaling pathway.

Development of a 3D atlas of the embryonic pancreas for topological and quantitative analysis of heterologous cell interactions.

Generating comprehensive image maps, while preserving spatial three-dimensional (3D) context, is essential in order to locate and assess quantitatively specific cellular features and cell-cell interactions during organ development. Despite recent advances in 3D imaging approaches, our current knowledge of the spatial organization of distinct cell types in the embryonic pancreatic tissue is still largely based on two-dimensional histological sections. Here, we present a light-sheet fluorescence microscopy approach to image the pancreas in three dimensions and map tissue interactions at key time points in the mouse embryo. We demonstrate the utility of the approach by providing volumetric data, 3D distribution of three main cellular components (epithelial, mesenchymal and endothelial cells) within the developing pancreas, and quantification of their relative cellular abundance within the tissue. Interestingly, our 3D images show that endocrine cells are constantly and increasingly in contact with endothelial cells forming small vessels, whereas the interactions with mesenchymal cells decrease over time. These findings suggest distinct cell-cell interaction requirements for early endocrine cell specification and late differentiation. Lastly, we combine our image data in an open-source online repository (referred to as the Pancreas Embryonic Cell Atlas).

Quantitative lineage analysis identifies a hepato-pancreato-biliary progenitor niche.

Studies based on single cells have revealed vast cellular heterogeneity in stem cell and progenitor compartments, suggesting continuous differentiation trajectories with intermixing of cells at various states of lineage commitment and notable degrees of plasticity during organogenesis1-5. The hepato-pancreato-biliary organ system relies on a small endoderm progenitor compartment that gives rise to a variety of different adult tissues, including the liver, pancreas, gall bladder and extra-hepatic bile ducts6,7. Experimental manipulation of various developmental signals in the mouse embryo has underscored important cellular plasticity in this embryonic territory6. This is reflected in the existence of human genetic syndromes as well as congenital malformations featuring multi-organ phenotypes in liver, pancreas and gall bladder6. Nevertheless, the precise lineage hierarchy and succession of events leading to the segregation of an endoderm progenitor compartment into hepatic, biliary and pancreatic structures have not yet been established. Here we combine computational modelling approaches with genetic lineage tracing to accurately reconstruct the hepato-pancreato-biliary lineage tree. We show that a multipotent progenitor subpopulation persists in the pancreato-biliary organ rudiment, contributing cells not only to the pancreas and gall bladder but also to the liver. Moreover, using single-cell RNA sequencing and functional experiments we define a specialized niche that supports this subpopulation in a multipotent state for an extended time during development. Together these findings indicate sustained plasticity underlying hepato-pancreato-biliary development that might also explain the rapid expansion of the liver while attenuating pancreato-biliary growth.

A Specialized Niche in the Pancreatic Microenvironment Promotes Endocrine Differentiation.

The interplay between pancreatic epithelium and the surrounding microenvironment is pivotal for pancreas formation and differentiation as well as adult organ homeostasis. The mesenchyme is the main component of the embryonic pancreatic microenvironment, yet its cellular identity is broadly defined, and whether it comprises functionally distinct cell subsets is not known. Using genetic lineage tracing, transcriptome, and functional studies, we identified mesenchymal populations with different roles during pancreatic development. Moreover, we showed that Pbx transcription factors act within the mouse pancreatic mesenchyme to define a pro-endocrine specialized niche. Pbx directs differentiation of endocrine progenitors into insulin- and glucagon-positive cells through non-cell-autonomous regulation of ECM-integrin interactions and soluble molecules. Next, we measured functional conservation between mouse and human pancreatic mesenchyme by testing identified mesenchymal factors in an iPSC-based differentiation model. Our findings provide insights into how lineage-specific crosstalk between epithelium and neighboring mesenchymal cells underpin the generation of different pancreatic cell types.

Direct Lineage Reprogramming: Harnessing Cell Plasticity between Liver and Pancreas.

Direct lineage reprogramming of abundant and accessible cells into therapeutically useful cell types holds tremendous potential in regenerative medicine. To date, a number of different cell types have been generated by lineage reprogramming methods, including cells from the neural, cardiac, hepatic, and pancreatic lineages. The success of this strategy relies on developmental biology and the knowledge of cell-fate-defining transcriptional networks. Hepatocytes represent a prime target for ß cell conversion for numerous reasons, including close developmental origin, accessibility, and regenerative potential. We present here an overview of pancreatic and hepatic development, with a particular focus on the mechanisms underlying the divergence between the two cell lineages. Additionally, we discuss to what extent this lineage relationship can be exploited in efforts to reprogram one cell type into the other and whether such an approach may provide a suitable strategy for regenerative therapies of diabetes.

Robo signalling controls pancreatic progenitor identity by regulating Tead transcription factors.

A complex interplay of intrinsic factors and extrinsic signalling pathways controls both cell lineage commitment and maintenance of cell identity. Loss of defined cellular states is the cause of many different cancers, including pancreatic cancer. Recent findings suggest a clinical role for the conserved SLIT/ROBO signalling pathway in pancreatic cancer. However, whilst this pathway has been extensively studied in many processes, a role for Slit and Robo genes in pancreas cell identity and plasticity has not been established yet. Here, we identify Slit/Robo signalling as a key regulator of pancreatic progenitor identity. We find that Robo1 and Robo2 are required for preserving pancreatic cell identity shortly after fate induction and, subsequently, for expansion of the pancreatic progenitor pool in the mouse. Furthermore, we show that Robo receptors control the expression of Tead transcription factors as well as its downstream transcriptional activity. Our work identifies an interplay between Slit/Robo pathway and Tead intrinsic regulators, functioning as gatekeeper of pancreatic cell identity.