Tissue-specific gene expression defines cellular identity and function, but knowledge of early human development is limited, hampering application of cell-based therapies. Here we profiled 5 distinct cell types at a single fetal stage, as well as chondrocytes at 4 stages in vivo and 2 stages during in vitro differentiation. Network analysis delineated five tissue-specific gene modules; these modules and chromatin state analysis defined broad similarities in gene expression during cartilage specification and maturation in vitro and in vivo, including early expression and progressive silencing of muscle- and bone-specific genes. Finally, ontogenetic analysis of freshly isolated and pluripotent stem cell-derived articular chondrocytes identified that integrin alpha 4 defines 2 subsets of functionally and molecularly distinct chondrocytes characterized by their gene expression, osteochondral potential in vitro and proliferative signature in vivo. These analyses provide new insight into human musculoskeletal development and provide an essential comparative resource for disease modeling and regenerative medicine.

These results identify a novel strategy for AC remediation via small molecule-mediated modulation of gp130 signalling.

Summary Cellular microenvironments established by the spatial and temporal expression of specific signaling molecules are critical for both the maintenance and lineage-specific differentiation of progenitor cells. In Drosophila, a population of hematopoietic progenitors, or prohemocytes, within the larval lymph gland [1] gives rise to three mature cell types: plasmatocytes, lamellocytes, and crystal cells. Removal of the secreted signaling molecules Hedgehog [2] and PVF1 [3] from the Posterior Signaling Center (PSC)[2, 4, 5] which acts as a niche, leads to a loss of progenitors and complete differentiation of the lymph gland. Here, we characterize a novel population of signaling cells within the lymph gland, distinct from the PSC, that are required for lineage specific differentiation of crystal cells. We provide evidence that Yorkie[6] and Scalloped[7], the Drosophila homologues of YAP and TEAD, are required in Lineage Specifying Cells to regulate expression of Serrate, the Notch ligand responsible for the initiation of the crystal cell differentiation program[8] [5]. Genetic manipulation of yorkie and scalloped in the lymph gland specifically alters Serrate expression and crystal cell differentiation. Furthermore, Serrate expression in Lineage Specifying Cells is eliminated in the Lymph Gland upon the immune response induced by wasp parasitization to ensure the proper differentiation of lamellocytes at the expense of crystal cells. These findings expand the roles for Yorkie/Scalloped beyond growth to encompass specific cell fate determination in the context of blood development. Similar regulatory functions may extend to their homologues in vertebrate progenitor cell niches that are required for specifying cell fate.

The Drosophila lymph gland is a well-characterized hematopoietic organ in which a population of multipotent stem-like progenitors is maintained by a combination of signals from different cellular populations within the organ. The lymph gland serves as an ideal model both for the interrogation of signaling mechanisms involved in progenitor maintenance as well as a tool for the identification of novel regulatory mechanisms in the highly conserved process of hematopoiesis. Here, we demonstrate a requirement for the TEAD transcription factor Scalloped in the maintenance and proliferation of hematopoietic progenitors. We have characterized a novel population of hemocytes in the early lymph gland identified by the expression of Hand, Scalloped, and the PVR ligand PVF2. In this unique population, we show that Scalloped maintains PVF2 expression, which is required for hemocyte proliferation and achievement of normal lymph gland size. We further demonstrate that STAT signaling marks actively proliferating hemocytes in the early lymph gland, and inhibition of this pathway causes decreased lymph gland growth similar to loss of Scalloped and PVF2, demonstrating a requirement for PVR/STAT signaling in the regulation of lymph gland size. Finally, we demonstrate that Scalloped regulates PVR expression and the maintenance of progenitors downstream of PVR/STAT/ADGF signaling. These findings further establish the role of the TEAD family transcription factors in the regulation of important signaling molecules, and expand our mechanistic insight into the balance between progenitor maintenance and proliferation required for the regulation of lymph gland homeostasis.

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