Genetic and chemical inhibition of autophagy in zebrafish induced myeloproliferation

Autophagy is an evolutionary conserved and dynamic lysosomal degradation process for cellular homeostasis and remodelling, which is essential for the development and maintenance of different hematopoietic fates. However, the roles of autophagy in definitive hematopoiesis remain elusive. Here, we exploited zebrafish (Danio rerio) to investigate the effect of knocking-out unc-51 like autophagy activating kinase 1b and 2 (ulk1b and ulk2), homologous of human ULK1 and ULK2, respectively, on definitive hematopoiesis. Upon ulk1b or ulk2 mutation, autophagosome formation was blocked in zebrafish embryos. More importantly, pan-leukocytes (lcp1), common myeloid progenitors (spi1b), neutrophils (mpx), and macrophages (mpeg1.1) significantly elevated, while the hematopoietic stem and progenitor cell (HSPC) (myb), erythroid progenitors (gata1), and embryonic hemoglobin (hbae1.1) significantly reduced in the caudal hematopoietic tissue (CHT) of ulk1b or ulk2 mutant zebrafish embryos. On the other hand, chemically modulated autophagy induction by calpeptin, a downstream autophagy inducer for ulk complex, was insufficient to ameliorate dysregulated hematopoiesis in both ulk1b and ulk2 mutants. Conversely, autophagy inhibitor 3-Methyladenine functioned parallelly with the ulk mutants to maintain defective hematopoiesis. These observations raised a link between autophagy and definitive hematopoiesis and potentiates the fact that autophagy deficiency incorporates with myeloproliferation and anemia, which warrants the significance of autophagy in regulating definitive hematopoiesis.