The Drosophila melanogaster lymph gland is a haematopoietic organ1 -3 in which pluripotent blood cell progenitors proliferate and mature into differentiated haemocytes. Previous work 4 has defined three domains, the medullary zone, the cortical zone and the posterior signalling centre (PSC), within the developing third-instar lymph gland. The medullary zone is populated by a core of undifferentiated, slowly cycling progenitor cells, whereas mature haemocytes comprising plasmatocytes, crystal cells and lamellocytes are peripherally located in the cortical zone. The PSC comprises a third region that was first defined as a small group of cells expressing the Notch ligand Serrate 5 . Here we show that the PSC is specified early in the embryo by the homeotic gene Antennapedia (Antp) and expresses the signalling molecule Hedgehog. In the absence of the PSC or the Hedgehog signal, the precursor population of the medullary zone is lost because cells differentiate prematurely. We conclude that the PSC functions as a haematopoietic niche that is essential for the maintenance of blood cell precursors in Drosophila. Identification of this system allows the opportunity for genetic manipulation and direct in vivo imaging of a haematopoietic niche interacting with blood precursors.The Drosophila lymph gland primordium is formed by the coalescence of three paired clusters of cells that express Odd-skipped (Odd) and arise within segments T1-T3 (Fig. 1a) of the embryonic cardiogenic mesoderm 6 . At developmental stages 11-12, mesodermal expression of Antp is restricted to the T3 segment (Fig. 1b, c). A fraction of these Antp-expressing cells will contribute to the formation of the dorsal vessel 7,8 , whereas the remainder, which also express Odd, give rise to the PSC (Fig. 1d, e). By stages 13-16, the clusters coalesce and Antp is observed in 5-6 cells at the posterior boundary of the lymph gland (Fig. 1d, e). The expression of Antp is subsequently maintained in the PSC through the third larval instar (see Fig. 2a). The embryonic stage 16 PSC can also be distinguished by Fasciclin III expression (Fig. 1f, g) and at stage 17 these are the only cells in the lymph gland that incorporate BrdU (Fig. 1h).Previous studies have identified the transcription factor Collier (Col) as an essential component regulating PSC function 9 . The gene for this protein is initially expressed in the entire embryonicCorrespondence and requests for materials should be addressed to U.B. (banerjee@mbi.ucla.edu). Author Information Reprints and permissions information is available at www.nature.com/reprints. The authors declare no competing financial interests. NIH Public Access Author ManuscriptNature. Author manuscript; available in PMC 2010 January 17. (Fig. 1i). In contrast, col expression is absent in the PSC of Antp mutant embryos (Fig. 1j, k), establishing that Antp functions genetically upstream of Col in the PSC.In imaginal discs, the expression of Antp is related to that of the homeodomain cofactor Homothorax (Hth) 10 . In the embryonic l...

Summary Maintenance of a hematopoietic progenitor population requires extensive interaction with cells within a microenvironment or niche. In the Drosophila hematopoietic organ, niche-derived Hedgehog signaling maintains the progenitor population. Here we show that the hematopoietic progenitors also require a signal mediated by Adenosine deaminase growth factor A (Adgf-A) arising from differentiating cells that regulates extracellular levels of adenosine. The adenosine signal opposes the effects of Hedgehog signaling within the hematopoietic progenitor cells and the magnitude of the adenosine signal is kept in check by the level of Adgf-A secreted from differentiating cells. Our findings reveal signals arising from differentiating cells that are required for maintaining progenitor cell quiescence, and that function with the niche-derived signal in maintaining the progenitor state. Similar homeostatic mechanisms are likely to be utilized in other systems that maintain relatively large numbers of progenitors that are not all in direct contact with the cells of the niche.

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In Drosophila, blood development occurs in a specialized larval hematopoietic organ, the lymph gland (LG), within which stem-like hemocyte precursors or prohemocytes differentiate to multiple blood cell types. Here we show that components of the Wingless (Wg) signaling pathway are expressed in prohemocytes. Loss and gain of function analysis indicates that canonical Wg signaling is required for maintenance of prohemocytes and negatively regulates their differentiation. Wg signals locally in a short range fashion within different compartments of the LG. In addition, Wg signaling positively regulates the proliferation and maintenance of cells that function as a hematopoietic niche in Drosophila, the Posterior Signaling Center (PSC), and in the proliferation of crystal cells. Our studies reveal a conserved function of Wg signaling in the maintenance of stem-like blood progenitors and reveal an involvement of this pathway in the regulation of hemocyte differentiation through its action in the hematopoietic niche.