Studies On The Function Of Rab Proteins In Drosophila Phagocytosis And Hematopoiesis

Innate immunity is the first line of defense against invading pathogens and parasites.The innate immune response in Drosophila can be divided into two parts:humoral immune response and cellular immune response.In vivo phagocytosis is an essential part in cellular immune response,which is mediated by plasmatocytes.However,the molecular mechanism of phagocytosis is not very clear.The hemocytes involved in cellular immune response are derived from hematopoiesis.Hematopoiesis is a complex biological process,which is essential for individual growth,development and survive.Blood system in Drosophila is a good model for studying hematopoiesis due to highly conserved signaling pathways and factors.The hematopoiesis occurs in two waves:one at the embryonic stage from head mesoderm;another one in the lymph gland,the main hematopoietic organ during the larval stage.Different signaling pathways regulate the differentiation of lymph glands in a cell-autonomous or a non-cell-autonomous manner,whereas the mechanism of which warrants further study.Rab proteins are members of the larger family of Ras-like GTPases;their typical role is to modulate vesicle trafficking.To date 31 Rab proteins are identified.It was shown that Rab proteins regulate individual development and cytoskeleton reorganization.Rab5,Rab7 and Rab11 are makers for early endosomes,late endosomes and recycling endosomes,respectively.They are also members of the core Rab family.However,the functional role of Rab proteins in phagocytosis and hematopoiesis is not very clear.Analyzing their roles will advance our understanding of Rab GTPase as well as the relationship between vesicle transport and phagocytosis as well as hematopoiesis.In this study,we utilized CZ-and hemocyte-specific Gal4 driver,Rab transgenic flies and latex beads to screen 31 Rab proteins.Then we confirmed our results in the candidate proteins using three different pathogens including Erwinia carotovora carotovora 15(Ecc 15),Staphylococcus aureus(S.aureus)and Beauveria bassiana(B.bassiana).In addition,we focused on Rab5,Rab7 and Rabll,and determined their roles in the proliferation and differentiation of hemocytes.Finally,we showed the regulatory network in lamellocyte differentiation.The main results are shown as follows:1.RabX3,RabX5 and RabX6 promoted the phagocytosis of latex beads,Ecc 15,S.aureus and B.bassiana spores,while Rab8,Rab30 and CG9807 suppressed the phagocytosis of latex beads,Ecc 15 and S.aureus;RabX1 suppressed the phagocytosis of latex beads,Ecc 15 and S.aureus.2.Inhibition of Rab5 and Rabll in CZ and in mature hemocytes induced the overproliferation of hemocytes and the overdifferentiation of plasmatocytes and lamellocytes.3.Rab5 and Rab11 regulated hemocyte proliferation through Ras/EGFR signaling pathway.4.In CZ,Rab5 and Rabll regulated the differentiation of plasmatocytes,which is independent of PVR/STAT92E/Adgf-A signaling.In MZ or PSC,Rab5 and Rab11 did not have a role in controlling homeostasis in lymph glands.5.Rab5 and Rab11 regulated lamellocyte differentiation through JNK,Ras/EGFR and Toll signaling pathways.In addition,Ras and Bsk(JNK)worked together to induce high p-JNK levels in endosomes and promoted lamellocyte formation.In addition,the lamellocyte differentiation was autophagy-dependent.The down-regulation of Atg1 or Atg8 could rescue the aberrant lamellocyte differentiation.Moreover,the lysosomal activity was important in this process.In summary,Rab5 and Rabll played a key role in controlling phagocytosis and the homeostasis of the blood system.The study will advance our understanding of Rab proteins as well as the relationship between vesicle transport and hematopoiesis.Furthermore,our results may provide a fundamental basis for studying vesicle transport in the pathology of leukemia.