The Drosophila melanogaster β2 protein (Dmβ2) has sustained a long evolutionary stasis for the last 60 million years (Nielsen 2006). Even small changes to the protein’s primary amino acid sequence render it non-functional, suggesting its stasis may be due to stringency in the structure/function relationship (Nielsen 2001). This project seeks to understand what has prevented Dmβ2 from evolving, with the two main hypotheses being that Dmβ2 either exists as an ideal protein configuration that competitively bests all alternates or that Dmβ2 is the only possible configuration that will support spermatogenesis in Drosophila melanogaster. In order to test these hypotheses, the ability of other proteins to rescue β2 function must be assessed. Previous work done to test β2 function used the major, non sperm-generator tubulin (β1) as a backbone to test the function of candidate sperm-generating residues. While sperm-generating residues were identified, none were sufficient to rescue fertility in a Dmβ2 null background (Nielsen 2001, Raff 2000). This project represents a different approach to analyzing the evolutionary stasis of Dmβ2 by testing the ability of a known sperm-generating ortholog from Glossina morsitans (commonly known as the tsetse fly) to rescue fertility. This sequence is 96% identical to Dmβ2 and is of particular interest because it is the closest relative to Drosophila melanogaster that possesses a variation in β2 sequence. When expressed in a Dmβ2 null background, the tsetse fly β2 (Gmβ2) generates long-tailed, fertile sperm when examined by light microscopy on testis samples and fertility tests between transgenic males and virgin wild-type females. This evidence supports the first of the two hypotheses outlined above, that β2 alternates exist but Dmβ2 is competitively superior. This shows the potential for β2 to participate in the process of evolution, potentially through allelic effects on sperm-tail length, which plays an important role in the retention of sperm in the female reproductive tract. Comparative analyses of outgroups, such as the human β2 ortholog (Hsβ3), will provide further information necessary to assess the roles of generic aspects of β2 such as motility versus more lineage-specific properties such as sperm tail length in the process of spermatogenesis
