A high-throughput screening for mammalian cell death effectors identifies the mitochondrial phosphate carrier as a regulator of cytochrome c release

Oncogene. 2008 Jan 3;27(1):44-54. doi: 10.1038/sj.onc.1210600. Epub 2007 Jul 9.

Abstract

Functional annotation of complex genomes requires the development of novel experimental platforms with increased capacity. Here, we describe a high-throughput system designed to identify cDNAs whose overexpression induces morphologically distinct cell death modalities. The methodology incorporates two robotized steps, and relies on coexpression of library clones with GFP to reveal the morphological features presented by the dying cells. By using this system we screened 135 000 cDNA clones and obtained 90 independent molecules. Interestingly, three death categories were identified, namely; apoptotic, vacuolated and autophagic. Among the pro-apoptotic clones, we found four members of the mitochondrial carrier family: the phosphate and adenine nucleotide (type 3) transporters, and the mitochondrial carrier homologs (MTCHs) 1 and 2. Expression of these molecules induced cytochrome c release and caspase-9-dependent death. One of them, the phosphate carrier, was able to interact with members of the permeability transition pore complex ANT1 and VDAC1, and its binding to ANT1 was stabilized in the presence of apoptotic activators. Depletion of this carrier by siRNA delayed cytochrome c mobilization and apoptosis. These results attribute a previously undescribed apoptotic function to the phosphate carrier and, more generally, suggest that a common property of various mitochondrial transporters was exploited during evolution to regulate apoptosis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Apoptosis / physiology*
  • Apoptosis Regulatory Proteins / physiology*
  • Cloning, Molecular / methods
  • Cytochromes c / metabolism*
  • HCT116 Cells
  • HeLa Cells
  • Humans
  • Mitochondria / enzymology*
  • Mitochondria / metabolism
  • Mitochondrial Membrane Transport Proteins / physiology*
  • Mitochondrial Proteins / physiology*
  • Oligonucleotide Array Sequence Analysis
  • Phosphate Transport Proteins / physiology*

Substances

  • Apoptosis Regulatory Proteins
  • Mitochondrial Membrane Transport Proteins
  • Mitochondrial Proteins
  • Phosphate Transport Proteins
  • SLC25A3 protein, human
  • Cytochromes c