“Survival” of Mitochondria <i>in Vitro</i>

Romani, Roger J.; Ozelkok, S.

doi:10.1104/pp.51.4.702

Cited by 17 publications

(10 citation statements)

References 12 publications

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“…ATP will support restoration, metabolically derived energy appears to be more effective. The latter observation is consistent with earlier studies on the requisites for long-term maintenance ofenergylinked functions under energized conditions (32), and in the restoration of state 3 rates reported by Raison et al (27). It is also in accord with Luzikov's (17) contention that various mitochondrial systems are kept functional more by underlying metabolic activity than by available energy per se.…”

Section: Energy Requirementsupporting

confidence: 82%

“…The latter, which relies largely on the removal ofharmful substances and the use ofprotective agents, has passive connotations. Maintenance, on the other hand, is dynamic and energy-dependent (32). Nonetheless, confirmation of mitochondrial self-restoration calls for direct evidence along the lines first explored in a preliminary study by Romani and Howard (30).…”

mentioning

confidence: 91%

“…The pioneering study of Millerd, Bonner, and Biale published in 1953 (20) gave impetus to much research on the role dria were isolated as previously described (32) with the exception that PVP was increased from 0.2% to as high as 0.8% for late season fruit high in lipids. Where indicated, further purification of the organelles was achieved via the self-forming Percoll gradient centrifugation procedure of Moreau and Romani (22).…”

mentioning

confidence: 99%

“…Where indicated, further purification of the organelles was achieved via the self-forming Percoll gradient centrifugation procedure of Moreau and Romani (22). Mitochondrial protein was initially determined by a modified Lowry procedure (19) (31,32). Chloramphenicol, added to suppress bacterial contamination, had been shown to be effective for at least 24 h (33).…”

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confidence: 99%

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Metabolically Driven Self-Restoration of Energy-Linked Functions by Avocado Mitochondria

Huang¹,

Romani²

1991

Plant Physiol.

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To assess the restorative capacity of isolated avocado (Persea americana) fruit mitochondria, the organelles were first aged in the absence of an energy source at 250C for several hours until respiratory control and oxidative phosphorylation were greatly diminished or totally lost. Energy-linked functions were then gradually restored over a period of several hours after the addition of substrate. Restoration of respiratory control resulted from both an increase in state 3 and a decrease in state 4 respiratory rates. Either a-ketoglutarate or succinate served as restorants, each with distinctive rates of recovery in state 3 and state 4 respiration. ATP also served as a restorative agent but not as effectively as metabolizable substrate. ATP synthase activity was modulated by stress and restoration but neither the extent nor the rate of change was sufficient to constrain state 3 rates. Orthophosphate was released from the mitochondria during substrate-deprived stress. Restoration of phosphorylation preceded that of RC with phosphate uptake and phosphorylation being evident immediately upon the addition of substrate. During restoration [32P]orthophosphate was incorporated into several organic fractions: phospholipid, ATP, a trichloroacetic acid-precipitable mitochondrial fraction, and an organophosphate that accumulated in the medium in relatively large amounts. The organophosphate was tentatively identified as a hexosephosphate. Incorporation into ATP and the putative hexosephosphate continued unabated beyond the point of maximum restoration. Phosphate metabolism thus appears to be a necessary but not sufficient precondition for mitochondrial restoration and maintenance. Based on the recovery kinetics of the various phosphorylated components, the mitochondrial-bound fraction appears to be most directly linked with restoration. Results are discussed with reference to specific characteristics and components of selfrestoration and to possible underlying mechanisms. We suggest that a degree of self-restoration is consistent with the quasiautonomous nature of mitochondria and that this intrinsic capacity may be pivotal to the respiratory climacteric in senescent fruit cells and to cellular homeostasis in general.of mitochondria in the climacteric respiratory rise that accompanies the senescence of many fruit and other plant organs. Decades later it is still generally held that the respiratory climacteric has mitochondrial origins but causal relationships remain unresolved. Early postulates were premised on the notion that mitochondrial respiration increased during the climacteric in compensation for declining mitochondrial efficiency, with the latter attributed to either phosphorylative uncoupling (20) or the interposition of the energy-inefficient alternative electron transport pathway (37). These postulates gave way to subsequent evidence that mitochondria remain energy-efficient throughout the respiratory climacteric (16), and that the alternative pathway, though present, remains unengaged (39).Based on the ...

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Section: Energy Requirementsupporting

confidence: 82%

mentioning

confidence: 91%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Metabolically Driven Self-Restoration of Energy-Linked Functions by Avocado Mitochondria

Huang¹,

Romani²

1991

Plant Physiol.

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“…Nonetheless, Romani et al (26) discerned a ripening, age-related change in the mitochondrial system based on the observation that mitochondria in climacteric pears can neither withstand nor recover from doses of ionizing radiation that are tolerated by mitochondria in preclimacteric fruit.It has since been shown that isolated pear and avocado mitochondria can maintain RC' at 25 C for several hr (23,24). In analogy to experiments with long lived chloroplasts (6, 18), the maintenance of RC at 25 C has been referred to (24) as "survival."…”

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confidence: 99%

Ripening and in Vitro Retention of Respiratory Control by Avocado and Pear Mitochondria

Ozelkok¹,

Romani²

1975

Plant Physiol.

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The retention of respiratory control ("surv-ival") The absence, or near absence, of ultrastructural change in the mitochondria of ripening pear (1), apple (17), and tomato (9) fruits plus the persistent capacity of mitochondria to oxidize substrates and demonstrate respiratory control throughout the climacteric of many fruits, including avocado (10, 14), tomato (8), apple (11-13), and banana (7) attest to the fact that mitochondria in senescing fruit cells remain functionally sound. Nonetheless, Romani et al. (26) discerned a ripening, age-related change in the mitochondrial system based on the observation that mitochondria in climacteric pears can neither withstand nor recover from doses of ionizing radiation that are tolerated by mitochondria in preclimacteric fruit.It has since been shown that isolated pear and avocado mitochondria can maintain RC' at 25 C for several hr (23,24). In analogy to experiments with long lived chloroplasts (6, 18), the maintenance of RC at 25 C has been referred to (24) as "survival." It seems reasonable to assume that survival is a more critical index of organelle integrity than a single point-in-time assessment of RC and should be helpful in understanding the status of mitochondria in ripening fruit. In the present study, the possible relationship between the capacity of mitochondria to survive in vitro and fruit ripening and senescence was investigated. 5 mM ,B-mercaptoethanol; 0.2%c PVP). The hard tissues of preclimacteric avocados were macerated by using a porcelain hand grater submerged in the isolation medium. A ratio of 1:3 (w/v), tissue to isolation medium, was used. After sedimentation at 14,000g the mitochondria were dispersed in wash medium (0.25 M sucrose; 50 mM K phosphate, pH 7.2; 0.1C% BSA; 5 mM 3-mercaptoethanol), centrifuged for 5 min at 1,200g to remove clumped particles and then at 8,000g for 10 min. The final pellets were dispersed in a few tenths ml of wash medium resulting in mitochondrial suspensions containing 20 to 30 mg protein /ml. Similar procedures were followed in the isolation of mitochondria from Bartlett pears with the exceptions, based on prior experiments (25), that the PVP concentration was increased from 0.2 to 0.5% (w,/v) and the pH during maceration was maintained within 6.7 to 7 by dropwise addition of I M KOH.Assay and Incubation. Oxidative activity of mitochondria at 25 C was measured with a Clark-type polarographic electrode. The sequence of additions into the cuvette was as follows: 3 ml of reaction medium, mitochondrial suspension, and repeated additions of ADP until anaerobiosis. The reaction mixture consisted of 0.25 M sucrose, 66 mm potassium phosphate buffer (pH 7.2), 1 mM MgCl,, 3.3 J.M CoA, 33 J.M thiamine pyrophosphate, 0.1 mM NAD, 1 mg/ml BSA, 100 j.g'ml chloramphenicol, and 10 mM substrate. Unless otherwise indicated, a-KG was supplied as substrate for both incubation and assay.For incubation at 25 C, done according to Romani and Ozelkok (24), 3 ml of reaction medium were placed in a 20-ml beaker 239 www.plantphysiol....

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