Uncoupling Factor Research Articles

A tissue-specific uncoupler of mitochondrial oxidative phosphorylation

The influence of endogenous uncoupling agent from the rat liver on oxidative phosphorylation of the liver, kidney, heart, lungs, and brain mitochondria was studied; the tissue-specificity of its action was demonstrated. The selective action of the uncoupler on the mitochondrial membrane is more obviously expressed within the range of pH 6.9--6.3 approaching such of the hepatocyte cytoplasm. The tissue-specificity was independent of the original state of mitochondria and could be eliminated only by heat injury of the oxidative phosphorylation. The activity of the tissue-specific uncoupling factor failed to diminish when the ribosomes and microsomes were removed by centrifugation. Addition of bovine albumin to the incubation medium had no influence of the value of the tissue-specific uncoupling. Participation of the uncoupler in the intratissue proliferation control is discussed.

Photodynamic action of phloxine on mitochondrial respiration.

Photodynamic action of phloxine on mitochondrial respiration was examined, using rat liver mitochondria. Rat liver mitochondria were suspended in a medium containing phloxine, and irradiated with visible light in the presence of oxygen. The respiratory activities of the photooxidized mitochondria were assayed. The results are summarized as follows. (1) Phloxine-sensitized photooxidation of mitochondria inhibited 2, 4-dinitrophenol (2, 4-DNP)-released respiration. It can be deduced, from an approximate calculation, that the released respiration was inhibited by 10% as every 3 nmoles of oxygen per mg of mitochondrial protein was consumed by photooxidation. (2) Phloxine-sensitized photooxidation of mitochondria released the suppressed respiration. The supernatant obtained by centrifugation of photooxidized mitochondria also released the suppressed respiration of intact mitochondria. This suggests that certain uncoupling factors were released from photooxidized mitochondria. Uncoupling factor in question may be unsaturated fatty acids or U-factor, because bovine serum albumin abolished the respiration-releasing effect of the supernatant of photooxidized mitochondria.

Effect of insulin on mitochondrial respiratory control and oxidative phosphorylation of alloxan diabetic rats. A possible new action of insulin on liver mitochondria.

Respiratory contro l i n dex and ADP/0 ra tio of the liver m itochondria from a gr oup of a lloxan diabetic rats were s ignificantly decreased compared to a normal group. Insulin treatment on alloxan diabetic rats improved both i ndexes up to the normal levels. However, insulin addition in vitro into mitochondria s uspension had no effect on the both indexes. It was f ound that insulin addition in vitro blocks the uncoupled respirat ion induced by 2, 4·dinit rophenol. One of insulin actions, a disulfide hormone, in vi vo was proposed to protec t mitochondri a from uncoupling factor in conn ection with thioldisu l fide interchange to produce the conformationa l in mitochodri a membrane. The reason of reduc ed efficiency of oxidative phosphorylat ion in alloxan dia· be tic rats might be partl y due to la ck of protecti ve action of insulin .

Circadian organization of chitin in some insect skeletons

ABSTRACT A circadian clock is shown to be involved in the control of macromolecular orientation of chitin by cells secreting and organizing insect endocuticle. Daily organization of locust endocuticle into alternating lamellate and non-lamellate layers persists in constant temperature (36 ° C) and constant darkness for at least 2 weeks; the freerunning period is then about 23 h, so that after a number of days the circadian clock is 180° out of phase with the astronomical clock, with which it is normally phased. The rhythm is almost independent of temperature, with a Q10 of 1·04, as contrasted with a Q10 of 2·0 for the actual rate of increase of endocuticular thickness. Locust epidermal cells differ in response to specific imposed environmental conditions according to their location in the integument. In some cells, constant low temperature uncouples chitin lamellogenesis from the circadian clock, provided that illumination (light or dark) is constant also: the result is continuously lamellate endocuticle. In other cells constant light acts as an uncoupling factor, provided that temperature (high or low) is constant also: the result in this case is continuously non-lamellate endocuticle. The circadian rhythm of chitin lamellogenesis persists in a cave cricket (Dolichopoda linderi). A similar circadian lamellogenesis rhythm occurs in the endocuticle of nymphs and adults of the cockroach Periplaneta americana. A crossed-fibre multiple-ply endocuticle in the legs and wings of giant toe-biter water bugs (Belostomatidae) also displays circadian organization, the chitin macromolecules in any one layer lying in parallel fibres, at an angle of approximately 60° to those in the next layer. It is suggested that daily organization of the skeleton may be a general feature of arthropods. Examples include the phenomena of timing of chitin lamellogenesis; chitin crossed-fibrillar organization; degree of fluorescence of the rubber-like protein resilin; and mineralization of crayfish gastroliths.

UNCOUPLING OF OXIDATIVE PHOSPHORYLATION BY IONIZING RADIATION: II. THE STABILITY OF MITOCHONDRIAL LIPIDS AND CYTOCHROMEc

Mitochondria isolated from the thymus of rats or rabbits 4 hours after 800 rads of ionizing radiation show uncoupling of their oxidative phosphorylation. There are no significant differences in the contents of phospholipids, neutral lipids, or total fatty acids between normal and uncoupled mitochondria. There was no evidence of the formation of any lipid uncoupling factor in thymus mitochondria from irradiated animals.Heart or liver mitochondria isolated from animals irradiated up to 12 hours previously showed no uncoupling of oxidative phosphorylation although preparations in various media could be made with reduced cytochrome c contents. No effect of irradiation upon endogenous cytochrome c or its binding in these mitochondria could be detected.Thymus mitochondria would appear to contain very low quantities of cytochrome c (less than 0.2 μg per mg protein), yet still be tightly coupled. It is suggested that this low level may indicate a critical condition at which radiation damage could become manifest.

UNCOUPLING OF OXIDATIVE PHOSPHORYLATION BY IONIZING RADIATION: II. THE STABILITY OF MITOCHONDRIAL LIPIDS AND CYTOCHROMEc

Mitochondria isolated from the thymus of rats or rabbits 4 hours after 800 rads of ionizing radiation show uncoupling of their oxidative phosphorylation. There are no significant differences in the contents of phospholipids, neutral lipids, or total fatty acids between normal and uncoupled mitochondria. There was no evidence of the formation of any lipid uncoupling factor in thymus mitochondria from irradiated animals.Heart or liver mitochondria isolated from animals irradiated up to 12 hours previously showed no uncoupling of oxidative phosphorylation although preparations in various media could be made with reduced cytochrome c contents. No effect of irradiation upon endogenous cytochrome c or its binding in these mitochondria could be detected.Thymus mitochondria would appear to contain very low quantities of cytochrome c (less than 0.2 μg per mg protein), yet still be tightly coupled. It is suggested that this low level may indicate a critical condition at which radiation damage could become manifest.

Oxidative phosphorylation in guinea pig mammary gland mitochondria during various functional states

Oxidative phosphorylation was measured in the mammary gland mitochondria of guinea pigs during pregnancy, at parturition, and during lactation and retrogression. The phosphorylation concomitant with oxidation of succinate, α-ketoglutarate, and β-hydroxybutyrate was shown to be uncoupled in mitochondria from glands at all functional states except during lactation. The phosphorylative capacity could be restored by the in vitro addition of bovine serum albumin to the reaction system. Mitochondrial and microsomal fractions prepared from retrogressing guinea pig mammary glands completely uncoupled phosphorylation concomitant with oxidation of succinate in a rat liver mitochondrial preparation. The supernatant fraction from these glands had no effect on either the oxidative or phosphorylative capacity of the liver mitochondria. Preliminary evidence is presented to suggest that a specific uncoupling factor for oxidative phosphorylation is present in mammary tissue preparations during specific functional states.

Formation and disappearance of an endogenous uncoupling factor during swelling and contraction of mitochondria

During swelling of rat-liver mitochondria which occurs spontaneously or is induced by Ca 2+ or thyroxine, direct analytical measurements show a parallel intramitochondrial formation of U factor, a heat-stable, isooctane-soluble uncoupling and swelling agent of fatty acid nature. On the other hand, mitochondrial swelling induced by glutathione or by phosphate is not accompanied by increased formation of U factor. Serum albumin, which is a “trap” for U factor by virtue of its ability to bind fatty acids, prevents swelling induced by Ca 2+ and thyroxine, but not produced by GSH and phosphate. These findings therefore indicated that Ca 2+ and thyroxine accelerate mitochondrial swelling indirectly, by accelerating the enzymic formation of U factor from a precursor lipid. Stimulation of U factor formation is given by Ca 2+ in sonicated mitochondria, but not by l-thyroxine. When swollen mitochondria are contracted again by ATP, U factor rapidly disappears at a rate which parallels the rate of contraction and ceases when contraction ceases. While there is an enormous increase in the level of U factor during swelling, experiments with [ 14C]oleate as an indicator of U factor show that oxidative removal of oleate occurs simultaneously. On addition of ATP, which produces contraction of mitochondria and disappearance of U factor, [ 14C]oleate rapidly disappears. Most of th [ 14C]oleate is rapidly incorporated into the phosphatidic acid and cephalin fractions of mitochondrial lipids. These findings thus show that enzymic formation and removal of U factor are closely geared to mictochondrial swelling and contraction and that accumulation of U factor may be cause of swelling in the presence of Ca 2+ and thyroxine.

Binding of modified serum albumins with the uncoupling factor (fatty acids) from insect mitochondria.

Binding of modified serum albumins with the uncoupling factor (fatty acids) from insect mitochondria.