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An attempt to isolate intact mitochondria from dry pea seeds (Pisum sativum var. Alaska) ended in failure. Cytochrome oxidase in crude mitochondrial fraction from dry seeds was separated into three fractions by sucrose density gradient centrifugation. Two of the fractions contained malate dehydrogenase, whereas the other did not. Equilibrium centrifugation of mitochondrial membrane on sucrose gradients revealed that the membrane from the fraction without malate dehydrogenase was lighter than that from the others. Differences were observed in relative content of phospholipid to protein and in polypeptide composition analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis among the membranes from three fractions and imbibed cotyledons. Membrane from the fraction without malate dehydrogenase was rich in phospholipid and lacking in polypeptides with relatively high molecular weights as compared with that from others. During imbibition, the fraction without malate dehydrogenase and one of the other two disappeared rapidly after a lag phase lasting for at least 1 hour. Concomitantly, active and stable mitochondria increased in the cotyledons. The results were interpreted to indicate that there were at least three types of mitochondria in dry seeds, the membranes of which differed in their biochemical properties, and that the mitochondria became active and stable through assembly of protein into the membranes during imbibition.

Two RNases in bound forms associated with the microsomal membrane and with the ribosomes or unknown particles in pea root tissue were solubilized by subjecting the membrane to sonic oscillation in the presence of EDTA and KC1 and by treating the particles with EDTA, respectively. The RNases were than purified by DEAE-cellulose and Sephadex G-75 column chromatographies. The elution profiles of RNases from the columns were very similar. No significant differences were observed in their electrophoretic mobilities in polyacrylamide gels, in molecular weight, in activation by inorganic ions, urea or phospholipid micelles or in the dependence of their activities upon pH. The purified RNASES were not different from the bound enzymes as regards activation by inorganic ions and urea and the dependence of the activity upon pH. Triton X-100 stimulated the activity only if RNase was in a bound form associated with the microsomal membrane. We propose that the two RNases may be the same molecular species and differ only in the form of association with intracellular structures.

l-Leucine-U-(14)C was incorporated into mitochondrial protein in pea (Pisum sativum var. Alaska) cotyledons during the imbibing stages. Incorporation was almost completely inhibited by cycloheximide but not by chloramphenicol. Both antibiotics did not affect increases in mitochondrial activities and components of the cotyledons during imbibition. Therefore, mitochondrial development seems to be achieved by a transfer of protein pre-existing in the cytoplasm into the mitochondria rather than by de novo synthesis of mitochondrial protein. Cycloheximide stimulated an increase in bile saltsoluble protein of mitochondria in imbibing pea cotyledons. The recovery of cytochrome oxidase activity after sucrose density gradient centrifugation was enhanced, and the morphological properties of mitochondria were altered by cycloheximide.

The acid-insoluble nitrogen content, lipid content, and cytochrome oxidase activity in the mitochondrial fraction are found to increase during incubation of slices of sweet potato (Ipomoea batatas) root tissue. These increases appear to be related to an increase in the number of the mitochondrial particles. The increase in the mitochondrial fraction is not accompanied by an increase in cell number. The nitrogen content in the mitochondrial fraction increases prior to the changes in the activity of cytochrome oxidase and lipid content. The increase in the numbers of the mitochondrial particles lags behind the increase in the cytochrome oxidase activity. Such findings are also found in the tissue infected by Ceratocystis fimbriata.The respiratory increase in response to wounding and infection appears to be a result of an increase in mitochondrial particles.

Rapid increases in activities and components of mitochondrial particles isolated from cotyledons of Pisum sativum var. Alaska during the early stage of germination are described. Respiratory rate of the cotyledons increased rapidly as hydration proceeded. A similar but slightly delayed increase in respiratory activity of the isolated mitochondrial fraction was observed. The respiratory control ratio and adenosine 5'-pyrophosphate/oxygen ratio rose during imbibition. Cytochrome oxidase and malate dehydrogenase activities in the mitochondrial fraction increased during the initial phase of imbibition. The increase seemed to precede that in respiratory activity. A significant activity of cytochrome oxidase and most of the malate dehydrogenase activity in the cotyledons were present in the postmitochondrial fraction in the case of the dry seeds. Mitochondrial protein and phospholipid also increased during imbibition, and the rise in the components seemed to concur with that in respiratory activity. The mechanism of mitochondrial development during imbibition is discussed.