Retrogenes are a group of functional genes produced by gene retroduplication events during evolution. It has been observed that many retrogenes have formed since the evolutionary divergence of Anopheles mosquitoes from the Aedes lineage as a result of developing heteromorphic sex chromosomes. It has been further observed that these retroduplications predominately occur from parent genes on the heteromorphic X chromosome to autosomes and have a predisposition to have enriched expression in testis. In order to investigate the nature of this male-biased expression in testis, we utilized bioinformatic techniques to identify retrotransposition events and assign them relative ages based on evolutionary branches of divergence. This list of parent genes and retrogenes were then analyzed and a total of twenty-five gene pairs were selected for further examination. Available gene expression data in the form of RNA-seq and DNA microarray were used in tandem with gene annotation data to computationally investigate gene pairs in An. coluzzii. These pairs were further investigated experimentally by means of RT-PCR conducted on dissected head, thorax, abdomen, and reproductive organs in both male and female Anopheles coluzzii Mopti strain. Testis and male accessory glands (MAGs) were also investigated by this method in An. coluzzii. Available expression data support previously observed testis enriched expression of retrogenes and provides evidence for the predominate expression of retrogenes occurring in postmeiotic cells suggesting retrogene involvement in sperm development. Experimental evidence revealed a small group of five retrogenes which exhibit the expected male-biased expression in male testis with little to no expression in female ovaries, although a shared expression in the heads of both sexes was observed. Of the five retrogenes, four carry out energy related functions involving mitochondria, suggesting contribution to energy requirements of developing sperm. Testis and MAG experiments in An. coluzzii revealed a predisposition for retrogenes to be expressed in testis while parent genes tended to have higher expression in MAGs, and this phenomenon is partially supported by DNA microarray expression data. Overall, these results suggest further investigation of retrogenes in An. coluzzii may reveal unique functions in male mosquito fertility that are exploitable in genetic approaches to mosquito control. / Master of Science in Life Sciences / Malaria is a potentially deadly disease which effects thousands of people every year. Malaria around the world is spread by multiple species of mosquitoes in a genus called Anopheles. Controlling the populations of these disease spreading mosquitoes is essential to preventing the spread of malaria. Current insecticide-based approaches used to stop mosquitoes are becoming less effective overtime as mosquitoes become resistant. A potential way to develop new techniques for mosquito control is through research involving mosquito reproductive genetics. Understanding the genes involved in how mosquitoes reproduce could improve future techniques designed to reduce or prevent mosquitos from reproducing. This research focuses on a group of genes called retrogenes which have formed over the evolution of these mosquitoes via the duplication from a separate parent gene. In mosquitoes these retrogenes are understood to be involved in male reproduction. The retrogenes involved in male mosquito reproduction could have important functions in male sexual reproduction and sterility. These important genes could be manipulated to interrupt whatever important functions these genes have in reproduction. In this research we first computationally identified retrogenes and their parent genes and categorized them by age. We then utilized available annotation and expression data to analyze the potential significance of retrogenes to male fertility and found that multiple retrogenes tended to be expressed during sperm development. Lastly, we conducted gene expression experiments using dissected head, thorax, abdomen, and reproductive organs in both male and female Anopheles mosquitoes. Results revealed unique patterns of expression that suggest male specific roles of five retrogenes in testes and head expression in both males and females suggesting a possible role in mating behavior. These results provide evidence that retrogenes do have functional roles in male fertility specifically related to the maturation and development of sperm.
| Identifer | oai:union.ndltd.org:VTETD/oai:vtechworks.lib.vt.edu:10919/107304 |
| Date | 09 July 2020 |
| Creators | Miller, Duncan Joseph |
| Contributors | Entomology, Sharakhov, Igor V., Sharakhova, Maria V., Tu, Zhijian |
| Publisher | Virginia Tech |
| Source Sets | Virginia Tech Theses and Dissertation |
| Detected Language | English |
| Type | Thesis |
| Format | ETD, application/pdf |
| Rights | In Copyright, http://rightsstatements.org/vocab/InC/1.0/ |
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