Appl Regulates The Function Of Endocrine Cells And The Fate Of Stem Cells In Drosophila

Appl is a transmembrane protein encoded by Drosophila genomes.It is highly conservative both structure and function,so it has important physiological functions.Appl is not only expressed in all neurons in the adult Drosophila brain,but also highly expressed in enteroendocrine cells(EEs)in the intestinal tissue.However,the physiological function of Appl in intestinal tissue remains unclear.In this study,genetic methods were used to explore the role of Drosophila Appl in regulating intestinal lipid metabolism and determining the differentiation of intestinal stem cells(ISCs)through immunofluorescence staining.At first,we found that knocking down the expression of Appl in EEs would lead to the accumulation of intestinal lipids,and the same result was obtained in the Appl^dmutant.The increased expressions of Fatty acid synthase(FAS)gene and Acetyl CoA Carboxylase(ACC)gene in the Appl^dmutants were detected by q PCR technique,which was similar to the intestinal lipid accumulation phenotype caused by the decrease of intestinal Tachykinin(TK)secretion.In order to investigate whether the intestinal lipid metabolism regulated by Appl is related to TK,we introduced TKg-Gal4 which specifically expressed in the intestine into the Appl^dmutant by using GAL4/UAS system,and obtained a stable genetic recombinant Drosophila strain.The results of the overexpression of TRAP1 showed that promoting the secretion of TK EEs could reverse the accumulation of lipid in the intestine of the Appl^dmutant,suggesting that Appl affects the secretion function of TK EEs and thus regulates lipid metabolism in the intestine.By expressing exocrine Appl.s and secretion-blocking Appl.sd in the TK EEs,we found that the intracellular AICD fragment of Appl is the key to regulate intestinal lipid metabolism.By controlling the expression level of early endosome regulator gene Rab5 in TK EEs,we found that Appl plays a role through Rab5 to regulate lipid metabolism in the intestine.Due to the homology between human App695 and drosophilia Appl and the conserved evolutionary function,we found that overexpression of human App695 in TK EEs can reverse the accumulation of intestinal lipid in Appl^dmutants.The EEs were stained by cell immunofluorescence technology.Through the statistics of the number of EEs,it was found that the number of EEs in the intestine of the Appl^dmutant was more than that of the iso-w¹¹¹⁸wild-type;knocking down Appl during the ISCs period resulted in an increase in the number of EEs.The above results all suggests that Appl may be involved in the differentiation of ISCs to EEs.At the same time,the results of expression of exocrine Appl.s and secretion-blocking Appl.sd during the ISCs period suggests that the intracellular AICD region of Appl may be the key to regulate the differentiation direction of ISCs.In summary,Drosophila Appl regulates the secretory function of EEs and the fate of ISCs through its intracellular AICD region.