TOMOGRAPHY OF YEAST MITOCHONDRIA THAT HAVE UNDERGONE PERMEABILITY TRANSITION

点击翻译按钮获取中文摘要

(Supported by NIH HL49182 to D..R. Pfeiffer) Yeast mitochondria (Saccharomyces cerevisiae) contain a permeability transition pore which is regulated differently than the pore in mammalian mitochondria. In a mannitol medium containing 10 mM Pi and ethanol (oxidizable substrate), yeast mitochondria accumulate large amounts of Ca2+ upon the addition of an electrophoretic Ca2+ ionophore. Pore opening does not occur following Ca2+ uptake even though ruthenium red-inhibited rat liver mitochondria undergo rapid pore opening under analogous conditions. However, a pore does arise in yeast mitochondria when Ca2+ and Pi are not present, as monitored by swelling, ultrastructure, and matrix solute release. Pore opening is slow unless a respiratory substrate is provided (ethanol or NADH), but also occurs rapidly in response to ATP when oligomycin is present. Iso-osmotic polyethylene glycol-induced contraction of yeast mitochondria swollen during respiration, or in the pres ence of AT P, is 50% effective at a solute size of 1.0-1.1 kDa. In this particular study, we wish to see the ultrastructural changes of yeast mitochondria in a contracted state, an induced swollen state (by addition of ETOH) and then in a recontracted state induced by the addition of PEG. Tomographic reconstructions from thick sections are being used to help us to analyze the changes in structure of the mitochondrial cristae in all three phases. Seven tilt series were recorded using the HVEM, on 0.5(m-thick sections of plastic-embedded isolated yeast mitochondria. Three reconstructions and models were made of control (contracted) mitochondria and one of a recontracted mitochondrion. A high school student, Alice Yip, is assisting on this project for the Westinghouse Science Competition. Thin EM sections of control, swollen, and recontracted yeast mitochondria were photographed to document the variability within each population. The models from the reconstructions showed that in both contracted and recontracted mitochondria, the flow between the intracristal space and the space between the inner and outer membranes is constricted, although the constrictions do not have the same long, tubular appearance as those in other mitochondria (rat liver, neurons, muscle, and Neurospora crassa) we have studied. Preliminary results were shown at the Albany Conference on Frontiers of Mitochondrial Research in September. Mannella, C.A., Buttle, K., Pfeiffer, D.R., Rath, B.K., Marko, M. (1998) Mitochondrial design: implications for the structural basis of the permeability transition. Proc. Albany Conference on Frontiers of Mitochondrial Research.