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Nucleic Acids Research, 1982, Vol. 10, No. 13 4107-4117
© 1982

MOLECULAR BIOLOGY

Chromosome-bound mitotic factors: release by endonucleases

Ramesh C. Adlakha1, Chintaman G. Sahasrabuddhe2, David A. Wright3, William F. Lindsey3, Marion L. Smith1 and Potu N. Rao1

1Departments of Developmental Therapeutics, The University of Texas, M.D.Anderson Hospital and Tumor Institute at Houston Houston, TX 77030, USA 2Departments of Pathology, The University of Texas, M.D.Anderson Hospital and Tumor Institute at Houston Houston, TX 77030, USA 3Departments of Genetics, The University of Texas, M.D.Anderson Hospital and Tumor Institute at Houston Houston, TX 77030, USA

Received March 19, 1982. Revised June 7, 1982. Accepted June 7, 1982.

Additional evidence is presented to support our recently reported conclusion that the mitotic factors of mammalian cells, which induce germinal vesicle breakdown and chromosome condensation when injected into fully grown Xenopus laevis oocytes, are localized on metaphase chromosomes. Chromosomes isolated from mitotic HeLa cells were further purified on sucrose gradients and digested for varying periods with either the micrococcal nuclease or DNase II. At each time point of digestion the amount of mitotic factors released was determined by injecting a supernatant of these fractions, obtained by high-speed centrifugation, into oocytes. The amount of DNA rendered add soluble under the conditions of digestion used was 3% to 5% of the total chromosomal DNA. The extent of release of mitotic factors with both nucleases was estimated to be about 30% to 40% as evidenced by the reextraction of the undigested chromosomal pellet with 0.2 M NaCl. Similar results were obtained when nuclei from G2 cells were digested under identical conditions. The release of these chromosome-bound mitotic factors by mild digestion with these nucleases though only partial, clearly demonstrates that a significant proportion of these factors are localized on metaphase chromosomes.


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