Nuclear matrix DNA from chicken erythrocytes contains beta-globin gene sequences.

Abstract

Nuclear matrices containing residual DNA were isolated from chicken erythrocytes after extraction of purified nuclei with buffered 2 M NaCI. After further purification of this residual DNA, it was found to contain high concentrations of -Wglobin gene sequences as assayed by dot hybridization with 32P-labeled nick-translated pHB1001. Electron microscopy of a random sample of this residual DNA fraction shows the DNA to be intimately associated with protein at various intervals. A hypothesis for enrichment of active genes in residual DNA from purified chromatin or in nuclear matrix DNA is also discussed.During the past decade many diverse experimental strategies have been applied to the study of eukaryotic transcriptional regulation by the nonhistone chromosomal proteins (1). In spite of all of these efforts, the nonhistones presumably responsible for tissue-specific transcription remain elusive. Studies on DNA sequences have provided evidence suggesting that the nucleotide sequences offlanking and intervening sequences are essential for correct gene expression (2). Nevertheless, analysis of restriction enzyme digests of chicken DNA for ,-globin (3), ovalbumin (4), conalbumin (5), and lysozyme (6) have shown that these genes appear to be indistinguishable between tissues where the genes are either active or inactive. Furthermore, comparison of the sequences around the putative promoter sites of the ovalbumin gene (7) cloned from the DNA of producer and nonproducer tissues revealed no differences. Thus it is reasonable to suggest that the nucleotide sequence alone is insufficient for genomic activation of tissue-specific transcription. Although gene-specific nonhistone proteins have not yet been found in chromatin obtained from specialized tissues, results supporting that possibility have been reported (8)(9)(10)(11)(12)(13).Of the total nuclear proteins, the insoluble nonhistones are significant in their content of tissue-specific proteins (14), in the structure of the nuclear matrix (15), and in their association with transcriptionally active genes (16). Recently, studies by Pettijohn's group also suggested that an insoluble nonhistone protein of high molecular weight appears to be specific to cells undergoing active proliferation (17). Previous results from our laboratory have shown that it is possible to isolate from purified chromatin a DNA fraction significantly enriched in tissue-specific gene sequences (18)(19)(20). We suspect that this DNA, which we have designated DNA-P, is similar to that DNA found associated with the nuclear matrix. Transcribed sequences are associated with the nuclear matrix (21), and these sequences may form the base of the DNA loops in which they reside (22). We show by dot hybridization that the insoluble DNA fraction isolated from chicken erythrocyte nuclear matrices contains high concentrations of f3-globin gene sequences, although this gene is known to be inactive in mature chicken erythrocytes (23). MATERIALS AND METHODSIsolation of DNA Associated with Nonhistones from th...

Transcription of the histone H5 gene is not S-phase regulated.

“…Recently, specific adenine-plus-thymine-rich nucleotide sequences between the Hi and H3 genes in the repeating histone gene units of Drosophila were found to be associated with the nuclear scaffold (28). This is a particularly interesting observation in view of other results which indicate that transcriptionally active P-globin (20), ovalbumin (6,36), and vitellogenin II (21) genes are always associated with the nuclear matrix. Furthermore, fixed sites of DNA replication have been proposed which involve DNA replication complexes being anchored to the matrix.…”

Isolation and characterization of RAT1: an essential gene of Saccharomyces cerevisiae required for the efficient nucleocytoplasmic trafficking of mRNA.

“…The nuclear matrix, a meshwork of thick polymorphic fibers apparently underlayed by a meshwork of intermediate-sized core filaments (Jackson and Cook 1988;He et al 1990), probably consists of elements of this nucleoskeleton. Actively transcribed chromatin is associated with the matrix (Robinson et al 1982;Ross et al 1982;Ciejek et al 1983;Hentzen et al 1984;Small et al 1985;Thorburn et al 1988), and it appears to be the substratum upon which heterogenous nuclear RNA (hnRNA) processing occurs (Zeitlin et al 1987(Zeitlin et al , 1989. m R N A and pre-mRNA are tightly bound to structures within the eukaryotic nucleus (Zeitlin et al 1987(Zeitlin et al , 1989) and move along "tracks" from sites of synthesis in the nuclear interior to the periphery (Lawrence et al 1989;Huang and Spector 1991;Xing and Lawrence 1991), possibly in association with fibers of the nucleoskeleton.…”

Core filaments of the nuclear matrix.

“…Additionally, the matrix may be the scaffold that determines higher order chromatin architecture (Nickerson et al, 1989). Actively transcribed genes are greatly enriched in nuclear matrix preparations suggesting the association of active chromatin regions with the matrix (Robinson et al, 1982;Ross et al, 1982;Ciejek et al, 1983;Hentzen et al, 1984;Small et al, 1985;Thorburn et al, 1988).…”