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J Newport

Publications and source records attributed to J Newport.

27 records · Page 2Linked to original sources
PubMed1987

Disassembly of the nucleus in mitotic extracts: membrane vesicularization, lamin disassembly, and chromosome condensation are independent processes.

We describe a stable cell-free mitotic extract derived from Xenopus eggs that contains activities necessary for nuclear envelope breakdown and chromosome condensation during mitosis. Using these cell-free extracts, we have demonstrated that nuclear envelope vesicularization, lamina solubilization, and chromosome condensation are independent and separable biochemical processes. We present evidence indicating that during mitosis nuclear membrane breakdown may involve the binding of a coating protein, lamin solubilization is enzymatically driven, and chromosome condensation involves both binding proteins and enzymatic activities including topoisomerase II. These results provide a coherent framework for investigating structural modification of the nucleus during mitosis at the biochemical level.

Animals↗
PubMed1984

Interconversion of metaphase and interphase microtubule arrays, as studied by the injection of centrosomes and nuclei into Xenopus eggs.

We have designed experiments that distinguish centrosomal , nuclear, and cytoplasmic contributions to the assembly of the mitotic spindle. Mammalian centrosomes acting as microtubule-organizing centers were assayed by injection into Xenopus eggs either in a metaphase or an interphase state. Injection of partially purified centrosomes into interphase eggs induced the formation of extensive asters. Although centrosomes injected into unactivated eggs (metaphase) did not form asters, inhibition of centrosomes is not irreversible in metaphase cytoplasm: subsequent activation caused aster formation. When cytoskeletons containing nuclei and centrosomes were injected into the metaphase cytoplasm, they produced spindle-like structures with clearly defined poles. Electron microscopy revealed centrioles with nucleated microtubules. However, injection of nuclei prepared from karyoplasts that were devoid of centrosomes produced anastral microtubule arrays around condensing chromatin. Co-injection of karyoplast nuclei with centrosomes reconstituted the formation of spindle-like structures with well-defined poles. We conclude from these experiments that in mitosis, the centrosome acts as a microtubule-organizing center only in the proximity of the nucleus or chromatin, whereas in interphase it functions independently. The general implications of these results for the interconversion of metaphase and interphase microtubule arrays in all cells are discussed.

Animals↗
PubMed1984

Respective roles of centrosomes and chromatin in the conversion of microtubule arrays from interphase to metaphase.

We report the results of studies in which partially purified centrosomes, nuclei, and DNA were injected into frog's eggs, which are naturally arrested in metaphase or interphase. These results have led to an independent assessment of the contributions of the centrosome and the chromatin to the formation of the mitotic spindle and suggest a simple explanation for the transition from interphase to metaphase microtubule arrays.

Animals↗
PubMed1983

Maturation-promoting factor induces nuclear envelope breakdown in cycloheximide-arrested embryos of Xenopus laevis.

We have studied the effect of maturation-promoting factor (MPF) on embryonic nuclei during the early cleavage stage of Xenopus laevis development. When protein synthesis is inhibited by cycloheximide during this stage, the embryonic cell cycle arrests in an artificially produced G2 phase-like state, after completion of one additional round of DNA synthesis. Approximately 100 nuclei can be arrested in a common cytoplasm if cytokinesis is first inhibited by cytochalasin B. Within 5 min after injection of MPF into such embryos, the nuclear envelope surrounding each nucleus disperses, as determined histologically or by immunofluorescent staining of the nuclear lamina with antilamin antiserum. The breakdown of the nuclear envelope occurs at levels of MPF comparable to or slightly lower than those required for oocyte maturation. Amplification of MPF activity, however, does not occur in the arrested egg as it does in the oocyte. These results suggest that MPF can act to advance interphase nuclei into the first events of mitosis and show that the nuclear lamina responds rapidly to MPF.

Animals↗
PubMed1982

A major developmental transition in early Xenopus embryos: I. characterization and timing of cellular changes at the midblastula stage.

The Xenopus embryo undergoes 12 rapid synchronous cleavages followed by a period of slower asynchronous divisions more typical of somatic cells. This change in cell cleavage has been termed the midblastula transition (MBT). We show that at the MBT the blastomeres become motile and transcriptionally active for the first time. We have investigated the timing of the MBT and found that it does not depend on cell division, on time since fertilization or on a counting mechanism involving the sequential modification of DNA. Rather, the timing of the MBT depends on reaching a critical ratio of nucleus to cytoplasm. We view the MBT as a consequence of the titration of some substance, originally present in the egg, by the exponentially increasing nuclear material. When this substance is exhausted a new cell program is engaged, leading to the acquisition of several new cell properties.

Age Factors↗
PubMed1982

A major developmental transition in early Xenopus embryos: II. Control of the onset of transcription.

We have shown in the accompanying paper that a developmental transition occurs at the midblastula stage (cleavage 12) in Xenopus embryos, and that this midblastula transition (MBT) is apparently initiated when the ratio of nucleus to cytoplasm reaches a critical value. One manifestation of this transition is the onset of transcription. We show here that a plasmid containing a cloned gene coding for a yeast leucine tRNA comes under developmental control when injected into cleaving eggs. In pre-MBT eggs this plasmid is transiently transcribed and then becomes inactive; however, it becomes transcriptionally active again at the MBT. This pre-MBT suppression of transcription can be reversed by addition of competing DNA. The amount of DNA needed to induce premature transcription is equal to the amount of nuclear DNA present after 12 cleavages (24 ng), suggesting that the MBT is triggered by the DNA through titration of suppressor components present in the egg.

Animals↗