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.

24Anthocyanins are the key factors controlling the coloration of plant 25 tissues. However, the molecular mechanism underlying the effects of 26 environmental pH on the synthesis of apple anthocyanins is unclear. 27 In this study, we analyzed the anthocyanin contents of apple calli 28 cultured in media at different pHs (5.5, 6.0, and 6.5). The highest 29 anthocyanin content was observed at pH 6.0. Additionally, the 30 moderately acidic conditions up-regulated the expression of 31 MdMYB3 as well as specific anthocyanin biosynthesis structural 32 genes (MdDFR and MdUFGT). Moreover, the anthocyanin content 33 was higher in calli overexpressing MdMYB3 than in the wild-type 34 controls at different pHs. Yeast one-hybrid assay results indicated 35 that MdMYB3 binds to the MdDFR and MdUFGT promoters in 36 vivo. An analysis of the MdDFR and MdUFGT promoters revealed 37 multiple MYB-binding sites. Meanwhile, electrophoretic mobility 38 shift assays confirmed that MdMYB3 binds to the MdDFR and 39 MdUFGT promoters in vitro. Furthermore, GUS promoter activity 40 assays suggested that the MdDFR and MdUFGT promoter activities 41 are enhanced by acidic conditions, and the binding of MdMYB3 42 may further enhance activity. These results implied that an 43 acid-induced apple MYB transcription factor (MdMYB3) promotes 44 anthocyanin accumulation by up-regulating the expression of 45 MdDFR and MdUFGT under moderately acidic conditions. 46 47 65 temperatures, hormones, high irradiance, sugar, and pH [10-14]. 66There is clear evidence that high pH levels enhance the degradation 67 of anthocyanins in carrot [12], while exposure to low pH induces the 68 expression of anthocyanin biosynthesis genes in crabapple leaves, 69 ultimately resulting in increased anthocyanin levels [15]. 70 Previous studies revealed that the accumulation of anthocyanins in 71 the vacuoles of plant tissue cells depends on the pH of the vacuoles 72 in which anthocyanins localize [16, 17]. In morning glory (Ipomoea 73 tricolor) petals, the pH of vacuoles is relatively low when the flower 74 bud opens, causing the petals to be red, but the vacuolar pH 75 increases over time and the petals eventually turn blue [18]. This 76 color change is due to the Na + /H + exchanger encoded by the 77 PURPLE gene, which transports sodium ions into the vacuole and 78 moves protons out of the vacuole to increase the pH [19]. Petunia 79 hybrida flowers normally have a lower pH than I. tricolor flowers, 80 and the wild-type (WT) flowers remain on the reddish (low pH) side 81 of the color spectrum. The PH4 gene encodes a MYB domain and is 82 expressed in the epidermis of petals [20]. A mutation to PH4 results 83 in the lightening of petal colors as well as increases in the pH of 84 petal extracts and, in some genetic backgrounds, the elimination of 85 anthocyanins in flowers [20]. Additionally, mutations to 86 ANTHOCYANIN1 (AN1), AN2, and AN11 lead to a loss of 87 anthocyanin pigments and an increase in the pH of petal extracts 88 [21]. In apple, some MYB TFs control cell pH, an...