Essential role of Drosophila black-pearl is mediated by its effects on mitochondrial respiration.
Black-pearl (Blp) is a highly conserved, essential inner-mitochondrial membrane protein. The yeast Blp homologue, Magmas/Pam16, is required for mitochondrial protein transport, growth, and survival. Our purpose was to determine the role of Drosophila Blp in mitochondrial function, cell survival, and proliferation. To this end, we performed mitotic recombination in Drosophila melanogaster, RNAi-mediated knockdown, MitoTracker staining, measurement of reactive oxygen species (ROS), flow cytometry, electron transport chain complex assays, and hemocyte isolation from Drosophila larvae. Proliferation-defective, Blp-deficient Drosophila Schneider cells exhibited mitochondrial membrane depolarization, a 60% decrease in ATP levels, increased amounts of ROS (3.5-fold), cell cycle arrest, and activation of autophagy that were associated with a selective 65% reduction of cytochrome c oxidase activity. N-acetyl cysteine (NAC) rescued Blp-RNAi-treated cells from cell cycle arrest, indicating that increased production of ROS is the primary cause of the proliferation and survival defects in Blp-depleted cells. blp hypomorph larvae had a 35% decreased number of plasmatocytes with a 45% reduced active mitochondrial staining and their viability was increased 2-fold by administration of NAC, which blocked melanotic lesions. Loss of Blp decreases cytochrome c oxidase activity and uncouples oxidative phosphorylation, causing ROS production, which selectively affects mitochondria-rich plasmatocyte survival and function, leading to melanotic lesions in Blp-deficient flies.
Department of Developmental and Molecular Biology, Albert Einstein College of Medicine, Bronx, NY 10461, USA.
Soumit
S
Roy
Mary K
MK
Short
E Richard
ER
Stanley
Paul T
PT
Jubinsky
D000111:Acetylcysteine / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001343:Autophagy / Q000502:physiology; D001483:Base Sequence; D015153:Blotting, Western; D017931:DNA Primers; D029721:Drosophila Proteins / Q000502:physiology*; D004331:Drosophila melanogaster / Q000502:physiology*; D004579:Electron Transport; D003576:Electron Transport Complex IV / Q000378:metabolism; D005434:Flow Cytometry; D016193:G1 Phase; D008855:Microscopy, Electron, Scanning; D008928:Mitochondria / Q000378:metabolism / Q000502:physiology*; D034622:RNA Interference; D017382:Reactive Oxygen Species / Q000378:metabolism; D016196:S Phase
C429744:BLP protein, Drosophila; D017931:DNA Primers; D029721:Drosophila Proteins; D017382:Reactive Oxygen Species; D000255:Adenosine Triphosphate; D003576:Electron Transport Complex IV; D000111:Acetylcysteine