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The piRNA System in Aedes aegypti

  • Author(s): Han, Michael
  • Advisor(s): Atkinson, Peter W
  • et al.
Abstract

The aim of the research presented in this thesis is to examine the piRNA pathway in Aedes aegypti, with an emphasis on understanding the role of the pathway in the soma. Chapter one reviews the piRNA pathway’s role in transposon regulation as well as transposon-independent roles, such as sex-determination in Bombyx mori. In addition, preliminary research from the Atkinson laboratory showed an expansion in the number and expression domain of the PIWI family in Aedes aegypti compared to a model Dipteran organism, Drosophila melanogaster. Chapter two introduces research I performed that showed the somatic expression of an important PIWI gene, Ago 3, in somatic ovarian follicular cells and larval gastric caecum. Piwi 2 was found to have a germline localization. In addition, an Ago 3 RNAi knockdown line (M14) exhibited a phenotype of larval mortality. Chapter three focuses on a new, more stringent method of annotating piRNA clusters in Ae. aegypti from different types of mosquito sRNA libraries, including both somatic and germline tissue. Two fairly distinct sets of piRNA clusters were discovered, one in the soma and one in the germline. Somatic clusters produced piRNA against predominately gypsy elements; somatic piRNA bore strong U1 signatures but weaker A10 signatures, and also bore less hallmarks of the piRNA ping-pong amplification loop. In contrast, germline clusters produced piRNA against a more varied set of transposons, and germline piRNA had both strong U1 and A10 signatures. Germline libraries also had larger quantities of transposon-derived piRNA. Chapter four examines the effect of Ago 3 knockdown in mosquito larvae. Modest decreases in U1 and A10 signatures were seen in piRNA sequenced from Ago 3 knockdown mosquitoes; in addition, the relative percent of piRNA mapping against transposons declined from wild-type and control conditions. A global decrease in mRNA mapping to transposons was also detected. Together, these data show that somatic piRNAs exist in Ae. aegypti. These piRNA play a role in transposon defense, but based on comparison with germline piRNA, somatic piRNA may also play a role in different pathways, such as gene regulation or viral defense.

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