Isolation and characterization of nuclear envelopes from three variant cell lines of the Shionogi mouse mammary carcinoma: Identification of androgen-dependent peptides

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Abstract
We have isolated and purified, with good yields, nuclear envelopes from an androgen-responsive and from two androgen-unresponsive cell lines of the Shionogi mouse mammary carcinoma after subjecting purified nuclei to DNase at high pH and characterized them morphologically, chemically, and enzymatically. Phase-contrast microscopy revealed the nuclei to be free of cytoplasmic tags and that the nuclear envelopes were isolated as membrane "ghosts." Electron micrographs clearly showed the double-membrane system with nuclear pore complexes which illustrates that the nuclear envelopes were ultra-structurally intact. The nuclear envelopes contained little DNA, low levels of arylesterase or acid phosphatase activity, and undetectable levels of succinate dehydrogenase and 5′-nucleotidase activity. Coomassie blue staining of the nuclear envelope fractions on sodium dodecyl sulfate – polyacrylamide gels for all three cell lines revealed that most of the polypeptides were similar. However, we have identified androgen-dependent peptides of molecular weights 29 000, 32 000, and 34 000 in nuclear envelopes of the androgen-responsive cell line peptide profiles by comparing the nuclear envelopes prepared from the androgen-responsive cell line grown in intact mice, in castrated mice, and in mice which had been injected with testosterone after castration. Further investigation of the androgen regulation of these nuclear envelope peptides may help us understand the molecular mechanisms involved during morphological changes of the nucleus which occur in response to different hormonal environments.
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