View Program - 63rd Annual Drosophila Research Conference

Proliferation is a key cellular process that is tightly regulated in cells and essential for the proper growth of multicellular organisms. Many studies have identified genes and proteins that are essential for regulated proliferation, but much less is understood about environmental factors that control proliferation, such as intracellular pH (pHi). pHi is tightly regulated by cells, and emerging evidence suggests regulated pHi dynamics modulate regulated cell proliferation. To regulate pHi, cells use a wide variety of ion exchangers and acid loaders/extruders to maintain pH near physiological levels. NHE1 in mammals (DNhe2 in Drosophila) is a ubiquitously expressed sodium proton exchanger that acts as a rheostat to maintain physiological pH. In diseases such as cancer, cells have constitutively increased pHi which in turn alters functions of pH-sensitive proteins leading to altered cell behaviors, like increased proliferation. However, it is unknown which specific pH-sensitive proteins are dysregulated at an increased pHi. To study the role of dysregulated pHi in cancer, our lab generated transgenic flies that inducibly express DNhe2, the homolog of NHE1, in the Drosophila eye. In previous work, our lab demonstrated that overexpression of DNhe2 in developing Drosophila tissues is sufficient to increase pHi and increase cell proliferation in vivo, and results in a rough eye phenotype in adult flies. Here we describe a reverse genetic screen to identify candidate pH-sensitive proteins that promote cell proliferation. We screened a collection of 193 Drosophila lines covering 94% of the second chromosome. We visually inspected flies for enhancement or suppression of the GMR>DNhe2 rough eye phenotype. We identified 35 regions of the second chromosome that show an interaction with GMR>DNhe2. We are focusing on two overlapping deficiencies that both enhanced the GMR>DNhe2 rough eye phenotype, Df (2L)ED1203 and Df(2L)ED1315 spanning 38B4-38C6. We obtained smaller deficiencies and single-gene mutations within this region to map the interaction to a single gene, and are currently testing for genetic interactions with GMR>DNhe2. Next, we will quantify the effects of candidate genes by quantifying proliferating cells in the third larval instar eye and wing imaginal discs. Understanding the effects of increased pHi and which possible pH-sensitive proteins induce hyperproliferation can help us understand and possibly uncover therapeutic targets.

Identifying Proteins that Mediate Increased Proliferation at Higher Intracellular pH