The effect of estrogen on the concentration of ovalbumin gene sequence in the 2 M NaCl residual fraction of oviduct chromatin

Mirell, Carol J.; Bekhor, Isaac

doi:10.1016/0014-5793(82)81316-1

Cited by 5 publications

(4 citation statements)

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“…Our previous observations that active tissue-specific genes (18)(19)(20) are concentrated in the DNA-P fraction along with RNA (19) also suggest that the preferential precipitation of these active genes could be due to the fact that these genes contain high concentrations of nascent RNA complexed with RNA polymerase (29). However, in the case of erythrocytes in which transcription of the f3-globin gene is not occurring (23), the observation that this gene is concentrated in the nuclear matrix DNA to about the same extent as in reticulocyte DNA-P (18) Fig.…”

Section: Resultsmentioning

confidence: 99%

“…It is also known that RNA polymerase-DNA complexes are not dissociated by high salt concentration, and therefore factors for transcription and subunits of RNA polymerase associated with DNA may become matrix components (28). Our previous studies on chromatin isolated from tissues active in ,3-globin (18), human placental lactogen (19), and ovalbumin (20) gene expression suggested that the 2 M NaCl-insoluble DNA-protein fraction contained high concentrations of their respective specialized gene sequences. Therefore, nonrandom sequences of DNA are found both in nuclear matrix DNA and in DNA-P. Robinson et al (21) suggested that the observed preferential association of the ovalbumin gene with nuclear matrix DNA isolated from chicken oviduct may have some implication for gene expression and the organization of nuclear DNA into supercoiled loop domains.…”

Section: Resultsmentioning

confidence: 99%

“…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.…”

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confidence: 91%

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

Hentzen¹,

Rho²,

Bekhor³

1984

Proc. Natl. Acad. Sci. U.S.A.

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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...

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 91%

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

Hentzen¹,

Rho²,

Bekhor³

1984

Proc. Natl. Acad. Sci. U.S.A.

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“…Thus, proteins in the CP-3 fraction appear to contain "acceptor activity" which endow DNA with the capacity for nativelike binding of PR. Similar fractions of tightly bound, non-histone proteins are associated with the nuclear matrix (Berezney & Coffey, 1977; Barrack & Coffey, 1980), with active-diffuse chromatin fractions (Wang et al, 1976) and with active structural genes (Bekhor & Mirell, 1979;Gates & Bekhor, 1980; Norman & Bekhor, 1981; Mirell & Bekhor, 1982). This same class of proteins has been reported to enhance transcription of DNA by increasing the binding of RNA polymerase to DNA (Bekhor & Samal, 1977).…”

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Reconstitution of nativelike nuclear acceptor sites of the avian oviduct progesterone receptor: evidence for involvement of specific chromatin proteins and specific DNA sequences

Spelsberg¹,

Gosse

Littlefield

et al. 1984

Biochemistry

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A specific fraction of avian oviduct chromosomal proteins can be reannealed to pure avian DNA to reconstitute nativelike specific nuclear binding sites (acceptor sites) for the oviduct progesterone receptor (PR). These specific nuclear binding sites represent the difference between the binding to the reconstituted NAP and that to pure DNA. The specific fraction of chromatin protein which contains the acceptor activity, fraction CP-3, is very tightly bound to hen DNA in a complex termed nucleoacidic protein (NAP). Removal of the CP-3 fraction from NAP results in a loss of specific PR binding sites. Resins containing chromatin adsorbed to hydroxylapatite are used as a rapid method to isolate the CP-3 fraction. Reconstitution of the CP-3 fraction to DNA by the described method involving a regressing gradient of 6-0 M guanidine hydrochloride (Gdn-HCl) results in a reconstituted NAP which displays specific PR binding sites identical with those in native (undissociated) NAP and whole chromatin. Optimal conditions and potential problems for reconstituting these nucleoproteins are described. Only partially purified receptor preparations were used in these cell-free binding analyses since they have been shown to bind with similar properties and patterns as the nuclear binding in vivo. Therefore, the binding of PR to the reconstituted NAPs was demonstrated to be receptor dependent, saturable, and of high affinity. Further, the pattern of binding to the reconstituted sites mimics those which are observed in vivo. Thus, nonfunctional receptors that cannot translocate and bind to the nuclear acceptor sites in vivo also failed to bind to the acceptor sites on the reconstituted NAPs generated by the acceptor proteins. In contrast, the binding to pure DNA does not reflect these receptor differences in receptor bindings. Specific binding of PR to reconstituted NAP can be reversed by again removing the protein fraction. Moreover, the specific binding can be destroyed by proteases and protected by protease inhibitors, indicating that acceptor activity is proteinaceous in nature. The reconstitution of the activity is both a concentration-dependent and time-dependent process. During the reconstitution, acceptor activity appears to reconstitute on the DNA when the Gdn-HCl concentration reaches 2.0 M. By use of the reconstitution method as an assay for acceptor activity, the activity in the CP-3 fraction was shown by molecular sieve chromatography to elute in a relatively broad molecular weight range between 13 000 and 25 000. The activity also focuses in isoelectric focusing resins with apparent pI's of 5.2 and 6.4.(ABSTRACT TRUNCATED AT 400 WORDS)

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Activated murine α-globin gene is not preferentially associated with the nuclear matrix

Kirov

Tsanev

1986

International Journal of Biochemistry

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The effect of estrogen on the concentration of ovalbumin gene sequence in the 2 M NaCl residual fraction of oviduct chromatin

Cited by 5 publications

References 35 publications

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

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

Reconstitution of nativelike nuclear acceptor sites of the avian oviduct progesterone receptor: evidence for involvement of specific chromatin proteins and specific DNA sequences

Activated murine α-globin gene is not preferentially associated with the nuclear matrix

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