Identification of the extracellular metallo-endopeptidases ADAM and ADAMTS in the yellow fever mosquito Aedes aegypti

Abstract

The mosquito Aedes aegypti is a major vector for dengue, Zika, yellow fever, and chikungunya (CHIKV) viruses, which cause significant morbidity and mortality among human populations in the tropical regions of the world. Following ingestion of a viremic bloodmeal from a vertebrate host, an arbovirus needs to productively infect the midgut epithelium of the mosquito. De novo synthesized virions then exit the midgut by traversing the surrounding basal lamina (BL) in order to disseminate to secondary tissues and infect those. Once the salivary glands are infected, the virus is transmitted to a vertebrate host along with saliva released during probing of the mosquito. Midgut tissue distention due to bloodmeal ingestion leads to remodeling of the midgut structure and facilitates virus dissemination from the organ. Previously, we described the matrix-metalloproteinases (MMP) of Ae. aegypti as zinc ion dependent endopeptidases (Metzincins) and showed MMP activity during midgut BL rearrangement as a consequence of bloodmeal ingestion and subsequent digestion thereby affecting arbovirus dissemination from the midgut. Here we investigate the ADAM/ADAMTS of Ae. aegypti, which form another major group of multi-domain proteinases within the Metzincin superfamily and are active during extra-cellular matrix (ECM) remodeling. Seven different ADAM and five ADAMTS were identified in Ae. aegypti. The functional protein domain structures of the identified mosquito ADAM resembled those of human ADAM10, ADAM12, and ADAM17, while two of the five mosquito ADAMTS had human orthologs. Expression profiling of Ae. aegypti ADAM/ADAMTS in immature forms, whole body-females, midguts, and ovarian tissues showed transcriptional activity of the proteinases during metamorphosis, bloodmeal ingestion/digestion, and female reproduction. Custom-made antibodies to ADAM10a and ADAM12c showed that both were strongly expressed in midgut and ovarian tissues. Furthermore, transient silencing of ADAM12c significantly reduced the carcass infection rate with CHIKV at 24 h post-infection, while silencing of ADAM12a significantly increased viral titers in secondary tissues at the same time point. Our results indicate a functional specificity for several ADAM/ADAMTS in those selected mosquito tissues.

Keywords: ADAM; ADAMTS; Aedes aegypti; Chikungunya virus; Functional domains; Gene expression; Gene-silencing; In situ detection; Midgut; Ovary; Phylogenetic analysis; Western blot.