Genetic Determinants of Phosphate Response in Drosophila

Clemens Bergwitz,Adam Friedman,Charles Derobertis,Norbert Perrimon,Jonathan Cohen,Stephanie E Mohr,Mark J Wee,Michael Schnall-Levin,Lawrence B Mensah,Sumi Sinha,Yanhui Hu,Joanne Huang,Meghana Kulkarni,Arunachalam Vinayagam,Lizabeth A Perkins,Bonnie Berger

doi:10.1371/journal.pone.0056753

Abstract

Phosphate is required for many important cellular processes and having too little phosphate or too much can cause disease and reduce life span in humans. However, the mechanisms underlying homeostatic control of extracellular phosphate levels and cellular effects of phosphate are poorly understood. Here, we establish Drosophila melanogaster as a model system for the study of phosphate effects. We found that Drosophila larval development depends on the availability of phosphate in the medium. Conversely, life span is reduced when adult flies are cultured on high phosphate medium or when hemolymph phosphate is increased in flies with impaired Malpighian tubules. In addition, RNAi-mediated inhibition of MAPK-signaling by knockdown of Ras85D, phl/D-Raf or Dsor1/MEK affects larval development, adult life span and hemolymph phosphate, suggesting that some in vivo effects involve activation of this signaling pathway by phosphate. To identify novel genetic determinants of phosphate responses, we used Drosophila hemocyte-like cultured cells (S2R+) to perform a genome-wide RNAi screen using MAPK activation as the readout. We identified a number of candidate genes potentially important for the cellular response to phosphate. Evaluation of 51 genes in live flies revealed some that affect larval development, adult life span and hemolymph phosphate levels.

Highlights

Inorganic phosphate, the mono- or divalent anion of phosphoric acid [HPO432, H2PO422], is required for cellular functions such as DNA and membrane lipid synthesis, generation of high-energy phosphate esters, and intracellular signaling
Culture of three commonly used laboratory fly strains, y w, Canton S (CS) and Oregon R (OR), on standard medium (SM) or SM supplemented with 30 mM sodium phosphate (P30) supported larval development and eclosion of adult flies after 10 days at 25uC
Similar to what we found for phosphonoformic acid (PFA), the effect of sevelamer could be rescued by supplementation with 30 mM sodium phosphate

Summary

Introduction

The mono- or divalent anion of phosphoric acid [HPO432, H2PO422], is required for cellular functions such as DNA and membrane lipid synthesis, generation of high-energy phosphate esters, and intracellular signaling. Cells and organisms have developed elaborate mechanisms to assure an adequate supply of phosphate. Yeast is the only eukaryote, in which the genetics of cellular phosphate homeostasis is understood [1]. When yeast cells are starved of phosphate, the phosphatedependent cyclin-dependent-kinase (CDK) inhibitor Pho inactivates the Pho80/Pho cyclin/CDK complex. The Phoregulon includes genes coding for high affinity phosphate transporters (Pho, Pho89) and secreted acid phosphatases (Pho, Pho, Pho12) [3]

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