Energy-dependent flip of fluorescence-labeled phospholipids is regulated by nutrient starvation and transcription factors, PDR1 and PDR3.
The yeast Saccharomyces cerevisiae readily accumulates short-chain, fluorescent 7-nitrobenz-2-oxa-1,3-diazol-4-yl (NBD)-labeled phosphatidylcholine and phosphatidylethanolamine at the nuclear envelope/endoplasmic reticulum and mitochondria. The net intracellular accumulation reflects the sum of their inwardly and outwardly directed transbilayer translocation across the plasma membrane (flip and flop, respectively). The rate of flop is negligible in energy-depleted cells as well as at low temperature (2 degrees C). Although flip is reduced at 2 degrees C, it can still be measured by flow cytometry, allowing the rate of flip, independent of flop, to be characterized at this temperature. Flip requires the energy of the plasma membrane proton electrochemical gradient and is down-regulated as cells pass through the diauxic shift and enter stationary phase. Furthermore, drug-resistant, gain-of-function mutations in the transcription factors, PDR1 and PDR3, result in a dramatic down-regulation of flip in addition to their already established up-regulation of flop. These results imply that down-regulation of the NBD-phospholipid flip pathway is a physiological response to environmental stress.
Hanson
P K
PK
Department of Physiology, Emory University School of Medicine, Atlanta, Georgia 30322, USA.
Nichols
J W
JW
D001386:Azides / Q000494:pharmacology; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000494:pharmacology; D002455:Cell Division; D002462:Cell Membrane / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D015536:Down-Regulation; D004721:Endoplasmic Reticulum / Q000378:metabolism; D005434:Flow Cytometry; D008564:Membrane Potentials; D008856:Microscopy, Fluorescence; D008928:Mitochondria / Q000378:metabolism; D009154:Mutation; D010713:Phosphatidylcholines / Q000378:metabolism; D010714:Phosphatidylethanolamines / Q000378:metabolism; D010743:Phospholipids / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D013696:Temperature; D013997:Time Factors; D015534:Trans-Activators / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*; D015854:Up-Regulation
D001386:Azides; D004268:DNA-Binding Proteins; C085480:PDR1 protein, S cerevisiae; C089261:PDR3 protein, S cerevisiae; D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D010743:Phospholipids; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators; D014157:Transcription Factors; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone