A major 62-kD intranuclear matrix polypeptide is a component of metaphase chromosomes.
We have isolated and partially characterized a major intranuclear matrix polypeptide from rat liver. This polypeptide, which is reversibly stabilized into the intranuclear matrix under conditions which promote intermolecular disulfide bond formation, has a Mr of 62,000 and pI of 6.8-7.2 as determined by two-dimensional IEF/SDS-PAGE. A chicken polyclonal antiserum was raised against the polypeptide purified from two-dimensional polyacrylamide gels. Affinity-purified anti-62-kD IgG was prepared and used to immunolocalize this polypeptide in rat liver tissue hepatocytes. In interphase hepatocytes the 62-kD antigen is localized in small, discrete patches within the nucleus consistent with the distribution of chromatin. The staining is most prominent at the nuclear periphery and somewhat less dense in the nuclear interior. Nucleoli and cytoplasm are devoid of staining. During mitosis the 62-kD antigen localizes to the condensed chromosomes with no apparent staining of cytoplasmic areas. The chromosomal staining during mitosis is uniform with no suggestion of the patching seen in interphase nuclei. Fractionation and immunoblotting studies using rat hepatoma tissue culture cells blocked in metaphase with colcemid confirm the chromosomal localization of this 62-kD intranuclear protein during mitosis. The 62-kD polypeptide fractionates completely with metaphase chromosome scaffolds generated by sequential treatment of isolated chromosomes with DNAse I and 1.6 M NaCl, suggesting that this major 62-kD intranuclear protein may be involved in maintaining metaphase chromosomal architecture.
Fields
A P
AP
Department of Pharmacology and Molecular Sciences, Johns Hopkins University, School of Medicine, Baltimore, Maryland 21205.
Shaper
J H
JH
D000818:Animals; D034961:Antigens, Nuclear; D002467:Cell Nucleus / Q000032:analysis; D002875:Chromosomes / Q000032:analysis*; D004591:Electrophoresis, Polyacrylamide Gel; D007118:Immunoassay; D007150:Immunohistochemistry; D007399:Interphase; D007525:Isoelectric Focusing; D008099:Liver / Q000032:analysis* / Q000648:ultrastructure; D008297:Male; D008677:Metaphase; D008938:Mitosis; D009687:Nuclear Proteins / Q000032:analysis / Q000302:isolation & purification*; D010449:Peptide Mapping; D010455:Peptides / Q000032:analysis / Q000302:isolation & purification; D051381:Rats; D011919:Rats, Inbred Strains
D034961:Antigens, Nuclear; D009687:Nuclear Proteins; D010455:Peptides