title: COPI-mediated blood meal digestion in vector mosquitoes is independent of midgut ARF-GEF and ARF-GAP regulatory activities.
abstract: We have previously shown that defects in COPI coatomer proteins cause 80% mortality in blood fed Aedes aegypti mosquitoes by 96 h post-feeding. In this study we show that similar deficiencies in COPII and clathrin mediated vesicle transport do not disrupt blood meal digestion and are not lethal, even though both COPII and clathrin functions are required for ovarian development. Since COPI vesicle transport is controlled in mammalian cells by upstream G proteins and associated regulatory factors, we investigated the function of the orthologous ADP-ribosylation factor 1 (ARF1) and ARF4 proteins in mosquito tissues. We found that both ARF1 and ARF4 function upstream of COPI vesicle transport in blood fed mosquitoes given that an ARF1/ARF4 double deficiency is required to phenocopy the feeding-induced mortality observed in COPI coatomer deficient mosquitoes. Small molecule inhibitors of guanine nucleotide exchange factors (GEFs) and GTPase-activating proteins (GAPs) are often transitory, and therefore, we investigated the role of five Ae. aegypti ARF-GEF and ARF-GAP proteins in blood meal digestion using RNA interference. Surprisingly, we found that ARF-GEF and ARF-GAP functions are not required for blood meal digestion, even though both vitellogenesis and ovarian development in Ae. aegypti are dependent on GBF1 (ARF-GEF) and GAP1/GAP2 (ARF-GAPs) proteins. Moreover, deficiencies in orthologous COPI regulating genes in Anopheles stephensi mosquitoes had similar phenotypes, indicating conserved functions in these two mosquito species. We propose that based on the need for rapid initiation of protein digestion and peritrophic membrane formation, COPI vesicle transport in midgut epithelial cells is not dependent on ARF-GEF and ARF-GAP regulatory proteins to mediate vesicle recycling within the first 48 h post-feeding.
authors.affiliation: Department of Chemistry & Biochemistry, Center for Insect Science, BioSciences West, University of Arizona, Tucson, AZ 85721, USA.
authors.forename: Jun
authors.initials: J
authors.lastname: Isoe
authors.forename: Weston
authors.initials: W
authors.lastname: Stover
authors.forename: R Barrett
authors.initials: RB
authors.lastname: Miesfeld
authors.forename: Roger L
authors.initials: RL
authors.lastname: Miesfeld
mesh_terms: D020727:ADP-Ribosylation Factors / Q000378:metabolism*; D000330:Aedes / Q000254:growth & development / Q000378:metabolism*; D000818:Animals; D000852:Anopheles / Q000254:growth & development / Q000378:metabolism*; D005075:Biological Evolution; D022181:COP-Coated Vesicles / Q000502:physiology; D020755:Coat Protein Complex I / Q000378:metabolism*; D004063:Digestion; D004847:Epithelial Cells / Q000502:physiology; D005247:Feeding Behavior; D005260:Female; D020690:GTPase-Activating Proteins / Q000378:metabolism*; D041981:Gastrointestinal Tract / Q000378:metabolism; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism*; D006801:Humans; D007303:Insect Vectors / Q000254:growth & development / Q000378:metabolism; D010053:Ovary / Q000254:growth & development; D010447:Peptide Hydrolases / Q000378:metabolism
chemical_list: D020755:Coat Protein Complex I; D020690:GTPase-Activating Proteins; D020662:Guanine Nucleotide Exchange Factors; D010447:Peptide Hydrolases; D020727:ADP-Ribosylation Factors
keywords: Mortality; Secretion; Vesicle transport; Vitellogenesis