DNS SEQUENCES IN NUCLEAR BINDING OF PROGESTERONE

  • Spelsberg, Thomas C, (PI)

Project: Research project

Project Details

Description

The nuclear binding sites (acceptor sites) for the progesterone-receptor
complex (PR) in the avian oviduct have been characterized in this
laboratory as involving specific nonhistone proteins, termed acceptor
proteins. Recent studies in this laboratory have revealed that these
proteins must be complexed to specific DNA sequences to generate the
acceptor sites. The reconstitution of hen oviduct acceptor proteins to the
DNAs from E. coli, salmon, plant (corn), and plasmid pBR 322 result in
protein-DNA complexes with no PR binding. In contrast, the reconstitution
of these proteins to hen DNA yields a protein-DNA complex with marked PR
binding. Eighty percent of the DNA from partially deproteinized chromatin
(NAP) can be degraded by DNAse I digestion with no loss of acceptor site
binding activity. Apparently the remaining proteins protect certainregions
of the DNA from nuclease digestion. The DNA in these protected regions
shows a marked enrichment of repetitive DNA sequences. It is planned to
further enrich the nuclease generated acceptor protein-DNA compelxes from
the multitude of other protein-DNA complexes initially on the basis of
differences in physical properties. A steroid receptor affinity resin and
an antiacceptor protein antibody affinity resin will also be applied for
the enrichment. The final enriched deoxyribonucleoprotein with acceptor
activity will de deproteinized, 32P-nick translated, and used to probe a
chicken genomic library for homologous sequences. Such sequences will be
unambiguously characterized by restriction endonuclease and nucleoride
sequence analysis and for acceptor activity by reconstituting with the
acceptor protein and measuring PR binding. The characterization of the
enriched acceptor-DNA sequences will also include the location of such
sequences in and around the ovalbumin gene. It is hoped that these studies
will identify the DNA sequences involved in the steroid regulation of gene
expression, the role these sequences perform in this function, and the
distribution of these sequences in the chick genome. The biological
function of specific nonhistone proteins and DNA sequences and/or
regulatory gene sequences may be achieved from these studies.
StatusFinished
Effective start/end date7/1/8212/31/86

Funding

  • National Institutes of Health

ASJC

  • Medicine(all)

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