SETDB1 modulates the differentiation of both the crystal cells and the lamellocytes in Drosophila.

Proper genetic and epigenetic regulation is necessary to maintain the identity and integrity of cells. Enzymes involved in post-transcriptional modifications of histones are key factors in epigenetic mechanisms. Such modifications are also gaining importance for their role in growth and development of cancer. SETDB1 catalyzes the epigenetic mark of lysine-9 methylation of histone-3. In this study, we explored the role of SETDB1 in Drosophila hematopoiesis. We show that SETDB1 controls the differentiation of matured blood cells in wandering third instar larvae. There are three matured blood cells in wild type Drosophila melanogaster : plasmatocytes, crystal cells and lamellocytes. We found that loss-of-function mutants of SETDB1 show hematopoietic defects; increased blood cell proliferation, decreased number of crystal cells, greater differentiation of blood cells into lamellocytes, dysplasia of the anterior lobes of lymph gland and presence of hematopoietic tumors. Cell type specific knockdown of SETDB1 provided similar phenotype i.e., decreased number of crystal cells and an increase in lamellocyte differentiation. In animals with loss of function of SETDB1, Notch pathway was downregulated. Further, over-expression of SETDB1 in blood cells resulted in an increase in the number of crystal cells. This increase is accompanied with an increase in the number of NotchICD expressing cells. We therefore performed genetic rescue using UAS-GAL4 system to rescue loss of function SETDB1 mutants. Our data show that the rescued larvae carrying a wild type copy of SETDB1 in mutant background are devoid of blood tumors. We have identified a novel dual function of SETDB1 methylatransferase as a critical regulator of two of the matured hemocytes, crystal cells and lamellocytes. We propose a novel role of SETDB1 in modulating the differentiation of crystal cells and lamellocytes from a common progenitor and underscore the importance of SETDB1 in Drosophila blood tumor suppression. • SETDB1 regulates crystal cell differentiation: Over-expression of SETDB1 increases the crystal cell number (≤5% in the wild type) i.e. when SETDB1 levels are increased in the blood cells we detected an increase in the number of crystal cells. Our results demonstrate that SETDB1 has a hold on maintaining the number of crystal number in wild type flies. • SETDB1 controls lamellocyte differentiation: Loss-of-function of SETDB1 leads to increased differentiation of blood cells into lamellocytes (<1% in the wild type) i.e. regulates progenitor differentiation into lamellocytes. • SETDB1 is a blood tumor suppressor: We observed blood tumors both, in the circulating hemolymph and in the developing hematopoietic organ, the lymph gland. These tumors are exclusively composed of blood cells i.e. abnormal plasmatocytes and also lamellocytes. Genetic rescue with the wild type SETDB1 expression in the loss-of-function mutants of SETDB1 relieves it from both lamellocyte and blood tumor formation. Therefore, our results clearly establish the novel function of SETDB1 as a blood tumor suppressor. [ABSTRACT FROM AUTHOR]