Recent findings suggest that membrane vesicle transport during mitosis may be generally inhibited. To test this, we examined the kinetics of uptake and exocytosis of RITC-transferrin in mitotic and interphase HeLa cells. We used quantitative image-intensification fluorescence microscopy to analyze the content of ligands in single cells. This technique was validated by comparison of 3H or RITC-transferrin release from interphase cells determined by microscopy or radiometry. Both methods gave a t1/2 of release of 5-6 min. The uptake of RITC-transferrin was depressed in mitotics. More importantly, we monitored the exocytosis of label during mitosis. Labeled mitotics were obtained by the progression of interphase cells into mitosis during a 50 min incubation with RITC-transferrin. After 30 min chase with unlabeled transferrin, the intensities of interphase cells approached background, whereas those of mitotic cells remained nearly constant. Thus both exocytosis and endocytosis of transferrin were exocytosis and endocytosis of transferrin were blocked during mitosis.
Occupational exposure to the human immunodeficiency virus (HIV) has led to a low but finite incidence of infection among health care providers. In such circumstances, postexposure administration of 3'-azido-3'-deoxythymidine (zidovudine; AZT) might be beneficial. To test this possibility, the SCID-hu mouse (the immunodeficient C.B-17 scid/scid mouse engrafted with human hematolymphoid organs) was treated with AZT at different times after intravenous infection with a standard dose of HIV (known to infect 100% of animals). If given within 2 h, AZT suppressed infection in all animals; if given after 2 days, no suppression was observed. At least in some animals, an AZT-sensitive phase lasted for as long as 36 h. These data support the hypothesis that prompt administration of AZT might be efficacious in suppressing acute HIV infection in humans. Further studies in the SCID-hu mouse might provide insight into treatment protocols of even greater efficacy.
Abstract. Several unique aspects of mitotic spindle formation have been revealed by investigation of an autoantibody present in the serum of a patient with the CREST (calcinosis, Raynaud's phenomenon, esophageal dysmotility, schlerodacytyly, and telangiectasias) syndrome. This antibody was previously shown to label at the spindle poles of metaphase and anaphase cells and to be absent from interphase cells. We show here that the serum stained discrete cytoplasmic foci in early prophase cells and only later localized to the spindle poles. The cytoplasmic distribution of the antigen was also seen in nocodazole-arrested cells and prophase cells in populations treated with taxol. In normal and taxol-treated cells, the microtubules appeared to emanate from the cytoplasmic foci and polar stain, and in cells released from nocodazole block, microtubules regrew from antigen-containing centers. This characteristic distribution suggests that the antigen is part of a microtubule organizing center. Thus, we propose that a prophase originating polar antigen functions in spindle pole organization as a coalescing microtubule organizing center that is present only during mitosis. Characterization of the serum showed reactions with multiple proteins at 115, 110, 50, 36, 30, and 28 kD. However, affinity-eluted antibody from the ll5/ll0-kD bands was shown to specifically label the spindle pole and cytosolic foci in prophase cells.
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