Ribonukleins�ure-Synthese der Riesenchromosomen

Pelling, Claus

doi:10.1007/bf00326915

Cited by 340 publications

(30 citation statements)

References 58 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…Transcribing bands do, of course, occur and are classified as puffs. The fraction of the total DNA represented by bands that are puffed is not precisely known but should amount to at least 10% (24). The data reported in ref.…”

Section: Resultsmentioning

confidence: 99%

Localization of RNA polymerase in polytene chromosomes of Drosophila melanogaster.

Jamrich¹,

Greenleaf²,

Bautz³

1977

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

160

View full text Add to dashboard Cite

RNA polymerase (RNA nucleotidyltransferase) B (or II) and histone HI of Drosophila melanogaster were localized on salivary gland polytene chromosomes using the indirect immunofluorescence technique. RNA polymerase B is present almost exclusively in puffs and interband regions, whereas histone HI is found primarily in bands. The puff at region 3C, known to be transcriptionally active in larval salivary glands, gives a bright fluorescence with antibodies against RNA polymerase B. This fluorescence disappears after exposure of the larvae to 370 for 45 min. The heat shock treatment results in a general reduction of fluorescence intensity with the appearance of brightly staining heat shock puffs. Heat-induced removal of RNA polymerase molecules from a puff does not immediately alter its morphology. We propose that an interband represents that fraction of the total number of gene copies in a band that are active, the inactive copies being present in a condensed form in the adjacent band. Large puffs would originate through the decondensation and activation of most or all gene copies in a band. Indirect immunofluorescence has become a widely applied tool for localization of not only surface antigens (1), but also intracellular components such as contractile proteins in nonmuscle cells (2). Recently, this technique has been applied to the study of the distribution of chromosomal proteins in polytene chromosomes of Drosophila (3)(4)(5). In these studies, chromatin was fractioned and either histones or nonhistone proteins, the latter being of unknown purities and functions, were used as antigens for their intrachromosomal localization. With the availability of antibodies raised against Drosophila RNA polymerase (RNA nucleotidyltransferase) B (6) we possess a probe to study the localization of a chromosomal protein of known function. Upon treatment of squash preparations from D. melanogaster salivary glands, Plagens et al. observed a distinct fluorescence pattern with the indirect immunofluorescence technique (7). They observed the a-amanitin-sensitive RNA polymerase B (also called polymerase II) not only in positions of visible puffs, but also at numerous other sites along the polytene chromosomes. Exposure of larvae to 37°resulted in a strong fluorescence at sites corresponding to the most prominent "heat shock" puffs (8). Meanwhile, a systematic study on conditions for preparing chromosomal squashes for immunofluorescence led to the observation that the postfixation with formaldehyde used previously caused shrinking of smaller puffs, yielding the impression of bands in the phase contrast pictures. This led to the original interpretation that RNA polymerase B was located also in bands (7). We studied the distribution of RNA polymerase B and of histone HI with the improved technique described here and found that RNA polymerase B occupies exclusively uncondensed regions of the chromosomes, whereas histone Hi is found primarily in condensed regions. Heat shock treatment results in a redistribution of RNA polymerase...

show abstract

Section: Resultsmentioning

confidence: 99%

Localization of RNA polymerase in polytene chromosomes of Drosophila melanogaster.

Jamrich¹,

Greenleaf²,

Bautz³

1977

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

160

View full text Add to dashboard Cite

show abstract

“…How are these observations to be related to the template activities of DNA with respect to replication and transcription in nuclei when these activities depend upon the endogenous polymerases? First, transcription: autoradiographs of RNA synthesis in calf-thymus nuclei (23) and Chironomus polytene chromosomes (24) show that only a very small part of the DNA is transcribed-in the former mainly in the relatively small amount of diffuse chromatin, in the latter mainly in the puffs. It is not generally supposed that the limited transcription in such cases is due to lack of RNA polymerase, but this possibility has not been excluded.…”

Section: Discussionmentioning

confidence: 99%

Accessibility of DNA in Chromatin to DNA Polymerase and RNA Polymerase

Silverman¹,

Mirsky²

1973

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

View full text Add to dashboard Cite

The accessibility of DNA in chromatin to both exogenous DNA polymerase and RNA polymerase is slight when compared to isolated DNA. DNA in extracted chromatin is somewhat more accessible to these enzymes than is DNA in the chromatin of isolated nuclei; and the DNA template of chromatin is more accessible to DNA polymerase than to RNA polymerase. In these experiments we have given much attention to the technique of scintillation counting, since artifacts arising in this procedure can lead to erroneous conclusions.Since DNA in chromatin is complexed with proteins, accessibility to DNA by enzymes and other substances is restricted. The role of histones in restricting access to DNA in chromatin was first observed by staining with basic dyes. Staining of the cell nucleus led Flemming (1) We report in this paper determinations of the accessibility of DNA in chromatin to exogenous DNA polymerase and DNA-dependent RNA polymerase. In our experiments we have studied chromatin in isolated nuclei and also in extracted chromatin (deoxyribonucleoprotein, DNP). Most of the previous work in this field has been on DNP, in which it is possible to replace certain components of chromatin, but the purpose of all these investigations is, of course, to understand chromatin as it is in the nucleus. In the experiments we are now reporting we have compared the activities of both exogenous DNA and RNA polymerases on free DNA and on DNA in both thymus nuclei and extracted chromatin. The comparison shows clearly that only a very small fraction of the DNA in a nucleus is accessible to either DNA or RNA polymerase; DNA in DNP is slightly more accessible. We have used exogenous polymerase in these experiments so that availability of the DNA template should not be obscured by enzymatic variability. It will be seen that we have given much attention to the technique of scintillation counting, for a mistake in this procedure can lead to erroneous conclusions. MATERIALS AND METHODS MaterialsThe preparation of nuclei isolated from calf thymus (6) and liver (7) in 0.25 M sucrose-3 mM CaC12 has been described. DNP (extracted chromatin) was prepared from thymus nuclei by the method of Paul and Gilmour (5, 8

show abstract

“…One-tenth ml antibody solution quantitatively precipitated [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20] ,ug of iodinated human albumin in a microprecipitin test. '4 Radioautographs were prepared from 10-,u sections of salivary glands from animals injected with labeled blood proteins according to the recommendations of Kopriwa and Leblond.18 Exposures were for 2-, 3-, and 4-week intervals.…”

mentioning

confidence: 99%