Identification of AaAtg8 as a marker of autophagy and a functional autophagy-related protein in Aedes albopictus

Aedes albopictus is a primary vector of hundreds of pathogens. Strong environmental adaptability and extensive global distribution of Aedes albopictus make it a severe threat to human health. Autophagy is a cellular process involved in maintenance of cellular homeostasis and recirculation of cytoplasm to generate macromolecule constituents and energy under different stress conditions. Many autophagy-related (Atg) proteins have been identified in yeast and were found in various organisms subsequently, indicating that the basic mechanism of autophagy is well conserved in eukaryotes. Among all Atg proteins, Atg8 plays important roles in autophagy and is widely used as a marker to monitor autophagic activity in yeast, Drosophila, nematodes, zebrafish and mammals. By now, Atg proteins in Aedes albopictus have not been reported yet and the autophagy pathway in Aedes albopictus remains unclear. This study identified a homolog of Atg8 from Aedes albopictus and named it AaAtg8. Sequence analysis revealed that AaAtg8 was highly conserved in the Atg8 family. This work proved that AaAtg8 was a functional Atg protein of Aedes albopictus and expressed during developmental and adult stages of Aedes albopictus. Moreover, the study also established the basic methods for autophagy study in C6/36 cells. First, it was proved that both rapamycin and starvation were applicable ways to induce autophagy in C6/36 cells, and that 3-methyladenine and chloroquine could be used to inhibit early and late stages of autophagy in C6/36 cells, respectively. Second, the results in this study showed that monodansylcadaverine staining could be used to detect autophagy in C6/36 cells. Additionally, the study revealed that the level of autophagy in C6/36 cells could be monitored by the turnover assay of AaAtg8 or fluorescent AaAtg8. Taken together, this study identified AaAtg8, the first reported Atg protein in Aedes albopictus. It also provided useful methods for studying autophagy in Aedes albopictus. To our knowledge, this is the first work about autophagy in Aedes albopictus.