Actively transcribed genes are associated with the nuclear matrix.

In the chicken oviduct, it has been well documented that steroid hormones stimulate the transcription of specific genes such as the ovalbumin gene. In addition to the presence of specific hormone receptors in the tissue, gene expression seems to require that target genes exist in large DNase I sensitive chromosomal domains. This structure appears necessary but not sufficient for transcriptional activation. In search of still other levels of control, we have investigated the interactions of genes with the nuclear matrix, a structure which has been implicated in DNA synthesis, transcription and RNA processing. Here we have isolated nuclear matrix and used a nondegradative method to fractionate nuclear DNA based on its preferential association with the matrix. The preparation was digested with a restriction enzyme and both matrix-bound and released DNAs were recovered. We found that only actively expressed genes were associated with the matrix. Furthermore, within a 100-kilobase (kb) DNase I sensitive chromosomal domain, only the transcribed regions were associated with the matrix. This association was shown to be reversible when hormone was withdrawn. Our results suggest that the nuclear matrix is the site of nuclear transcription and may represent another potential level of control for regulation of gene expression in the eukaryotic cell.

Ribonucleic acid precursors are associated with the chick oviduct nuclear matrix.

Nuclear matrix was prepared by sequential treatment of oviduct nuclei with Triton X-100, DNase I, and 2 M NaCl. Published procedures were modified such that as many steps as possible were performed at -20 degrees C to minimize endogenous ribonuclease activity. Examination of electron micrographs confirmed the isolation of intact nuclear matrix structures. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the proteins in these structures showed an absence of histones and an enrichment of certain nonhistone proteins. RNA was isolated from the nuclear matrix preparations and subjected to denaturing gel electrophoresis. Gels were analyzed by ethidium bromide staining and by hybridization of Northern blots to cloned DNA probes for ovalbumin, ovomucoid, 5.8S ribosomal RNA, and U1 RNA. All of the precursors to ovalbumin and ovomucoid mRNAs (including various splicing intermediates) and all of the precursors to ribosomal RNA were associated exclusively with the nuclear matrix fraction. By contrast, mature ovalbumin and ovomucoid mRNAs were distributed between matrix and nonmatrix fractions. These observations were further supported by quantitative hybridization analysis of the RNA in nuclear and matrix fractions. It was found that less than 50% of the mature message of intact nuclei was recovered in the matrix, while most significantly, over 95% of the mRNA precursors remained associated with the matrix. Finally, mature ribosomal RNAs and virtually all of the small nuclear RNAs (including U1 RNA) were also distributed between matrix and nonmatrix fractions. Our results suggest that all precursor RNAs (be they precursors to mRNA or rRNA) are exclusively associated with the nuclear matrix and support the notion that the nuclear matrix may be the structural site for RNA processing within the nuclei of eucaryotic cells.