When rat-liver rough microsomes were treated with 1.0% Triton X-100, the resulting polyribosomes were found to be still in association with membrane-like fragments or vesicles. Freeze-fracture studies of these membranous structures revealed no broad fracture faces of the lipid bilayer, but showed only a few 8 to 10nm-diameter particles similar to intramembrane particles, arranged in rows, clusters, or circles on the surface of the replica. A protein fraction solubilized from the membranous structures by sodium deoxycholate was reincorporated into artificial lipid vesicles by the detergentdialysis method. While protein-free liposomes did not bind ribosomes, the protein-containing ones bound ribosomes at 0°C. The cross-section of a vesicle recombined with ribosomes resembled that of a native hepatic rough microsome in thin section. The presence of the intramembrane particles in the protein-containing liposomes was shown by freeze-fracture techniques.

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The effects of cytochalasin B (CB) and lead acetate, inhibitors of "primary streaming" in Characean cells, were examined on "secondary streaming" in Vallisneria mesophyll cells with reference to the cessation of streaming and the reversal of its direction.
Cytoplasmic streaming was reversibly inhibited by CB in the concentration range of 10-100 μg/ml. The ratio of cells showing newly established streaming in the reverse direction after removal of CB to the total number of cells observed before treatment increased depending on the concentration and the duration of treatment, reaching 50 % after 24 h for CB treatment at 10-100 μg/ml. Lead acetate at 50-100 mM caused the complete cessation of streaming, but the streaming gradually recovered after its removal; however, lead acetate did not induce a change in the streaming direction. Properties of the motile apparatus in secondary streaming are discussed on this basis.

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The transport of phosphoenolpyruvate into red cells suspended in media with impermeable solutes was studied at different osmolalities and in the presence of echinocytogenic drugs. The transport rate decreased when cells were incubated in a hyper-osmotic media. The echinocytogenic reagents dipyridamol, salicylate, 2, 4-dinitrophenol and deoxycholate inhibited the transport rate, whereas the spherocytogenic reagents chlorpromazine, procaine, colchicine, vitamin E and Triton X-100 did not.

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The effects of salt treatment of isolated nuclei on the nature of DNA synthesis and on the solubilization of DNA polymerase a have been investigated. Treatment with 0.2 M KCl resulted in a deficiency in the function to join Okazaki fragments, whereas the synthesis of these fragments was little affected. No DNA synthesis was observed in 0.3 M KO-treated nuclei. DNA synthesis in 0.2 M KCl-treated nuclei required ATP, Mg²⁺ and four deoxy- ribonucleoside triphosphates, and was markedly inhibited by arabinofuranosyl-cytosine triphosphate and aphidicolin, but not by 2', 3'-dideoxythymidine triphosphate. This suggests that the DNA synthesis is the replicative-type, catalyzed by DNA polymerase a. DNA polymerase a in 0.3 M Kcl-nuclear extract was detected as two active forms distinct in their binding affinities for DNA. In 0.2 M KCl-treated nuclei, only one active form with high binding affinity was retained, indicating that this form is responsible for the synthesis of Okazaki fragments.

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Axonemal microtubules in a large heliozoan, Echinosphaerium nucleofilum strain EC, were examined especially for morphological changes under a highly sensitive polarizing microscope and with an electron microscope. Cold treatment (2°C) caused axopodial degradation to about 10 % of the initial length after 3 h, accompanied by a considerable decrease in the positive birefringence of the intra-axopodial components. Release from the cold treatment resulted in an increment in birefringence and in appearance of 40 nm macrotubules; as the axopodial re-extension proceeded, the diameter of the macrotubules decreased gradually, being replaced by 24 nm micro-tubules in fully-extended axopodia. Colchicine treatment (20 mM, 10 min) caused complete degradation of the axonemal microtubules; no fibrous structures could be detected within the cell body under a polarizing microscope. During the recovery process from colchicine, the cells showed increments in birefringence and decreases in the diameters of tubular structures (macro-tubule-to-microtubule transition). Furthermore, axopodial re-extension was not inhibited by protein synthesis inhibitors (100 μg cycloheximide/ml or 100μg puromycin/ml), but it was inhibited by energy inhibitors (50 μM KCN or 500μM NaN3). The possible mechanisms for macrotubule-to-microtubule transition are discussed mainly on the basis of morphological changes in the tubular structures.

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A new type of temperature-sensitive mutant, 11C3 has been isolated from CHO-K₁ cell and partially characterized. At a nonpermissive temperature the number of mutant cells hardly increases. Incubating mutant cells first at a nonpermissive temperature (39°C) for 24 h reduces the plating efficiency to one third that found without prior incubation. DNA synthesis slows down gradually for 30 h at 39°C, but protein synthesis does not decrease until 48 h after the shift up. Analysis of the DNA content of the individual cells shows that the proportion of cells containing the 4C amount of DNA markedly increases in 28 h after the temperature is shifted up. Light microscopy shows that mitotic figures decrease during this period and that the cells become large and flat with a single large nucleus. Experiments on synchronized cell populations suggest that although the temperature sensitive period exists in the S-phase, cells are arrested near the end of the S-phase or in the G₂-phase.
The colchicine binding assay revealed that the colchicine binding protein in mutant cells increased during incubation at a nonpermissive temperature. This supports the results described above.

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Changes in the content and the size of poly(A) sequences during progesterone-induced maturation of Xenopus laevis oocytes were determined by [³H]poly(U) hybridization and by labeling with exogenously supplied radioactive adenosine. The poly(A) content increased significantly (by about 20 %) by the time of germinal vesicle breakdown (GVBD), then decreased continuously to one half of the initial content by the end of maturation. Gel electrophoresis revealed that poly(A) sequences were being elongated by 20-30 nucleotides before GVBD except for the smallest class of poly-(A) (ca. 20 nucleotides long), but after GVBD they were extensively degraded which gave rise to the egg poly(A) sequences of quite different sizes. All these changes in poly(A) content and size also occurred in oocytes which either were treated with actinomycin D or enucleated before exposure to progesterone; this signifies that they were not dependent on the nucleus. Labeling experi-ments on maturing oocytes showed that poly(A) sequences were synthesized actively independent of the concomitant synthesis of RNA even when a substantial amount of the poly(A) sequences was being degraded (after GVBD). The size distribution of these labeled poly(A) sequences was the same as that of steady-state poly(A) measured by [³H]poly(U) binding, and they were associated with non-radioactive heterogeneous high molecular weight RNA. Thus, the present findings indicate that the poly(A) sequences associated with cytoplasmic maternal mRNA were being turned over extensively and that a portion of them was completely degraded during oocyte maturation. This poly(A) turnover also was indicated in full-grown oocytes which were not treated with progesterone and which showed no changes in the content and size of poly(A) throughout the experimental period.

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During in vitro incubation of thymus cells, proteins which have affinity for single stranded DNA (ss-DNA) are released into the medium. When the thymus cells were labeled with [³H]leucine for 5 h, about 3.5 % of the labeled leucine that was taken up by the cells was released into the medium in trichloroacetic acid-insoluble form, and about 2 % of the total radioactivity in the released proteins bound to the ss-DNA-cellulose. When the [³H]leucine labeled ss-DNA-binding proteins were incubated with spleen cells for 30 min at 37°C, about 2.6 % of the added [³H]labeled ss-DNA-binding proteins re-mained in the cells. In experiments using electron microscope autoradiography, some of the ss-DNA-binding proteins were found to enter the nuclei of spleen cells. Nuclei were isolated from these treated cells and radioactive proteins were recovered. Analysis of these proteins with sodium dodecyl sulfate gel electrophoresis gave five or six components whose molecular weights ranged from 1.4 to 7.2 × 10⁴.

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Rounded mitotic cells of a mouse melanoma cell line B16-C2W were harvested by gentle pipetting at different times after x-irradiation, and their mitotic spindles were visualized by indirect immunofluorescence using antibody against tubulin. At the first mitosis after irradiation with 1, 000R, 91 % of total rounded cells displayed mitotic spindles and 36 % of them showed gross abnormalities. The prominent abnormal features were multiple fluores-cent spots with or without association with spindle microtubules, and partial absence of fluorescence of the microtubules at the site of the lagging chromo-somes. As the irradiated cells progressed into the second or third mitosis, fractions of cells with clumped microtubules or those lacking spindles markedly increased. About 80 % reduction in the figures of anaphase and telophase, and scattered chromosomes on Giemsa staining suggested the prolonged arrest of cells at abnormal metaphase. Dead cells without dye-exclusion ability were selectively found among rounded cells after the second division rather than spread interphase cells. A good correlation was obtained between the fraction of the dead cells and the fractions of cells having spindle abnormality for 4 time-points after irradiation with 1, 000R. These results suggest that the mitotic cells with extensive spindle abnormalities die at that rounded state without the completion of the mitosis.

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Developmental changes in the fine structure of the posterior silk gland cells were studied with a recessive mutant of Bombyx mori named flimsy cocoon (flc) whose cocoon-shell weight is much less than that of the normal strain of B. mori. The flc posterior silk gland develops normally until 60 h after the fourth ecdysis. By the 72nd hour, unusual numbers of globules, which we have called as fibroin globules, begin to accumulate in the apical cytoplasm of the mutant cell. These globules increase in number until the 96th hour, when they fill most of the apical cytoplasm and extend into the basal region. In the normal cell at the corresponding stages, massive amounts of fibroin are synthesized and secreted rapidly with no storage of fibroin globules.

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