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Biomedical subjects

A Cavanaugh

Publications and source records attributed to A Cavanaugh.

8 recordsLinked to original sources
PubMed2001

The role of acetylation in rDNA transcription.

Treatment of NIH 3T3 cells with trichostatin A (TSA), an inhibitor of histone deacetylase (HDAC), resulted in a dose-dependent increase in transcription from a rDNA reporter and from endogenous rRNA genes. Chromatin immunoprecipitation using anti-acetyl-histone H4 antibodies demonstrated a direct effect of TSA on the acetylation state of the ribosomal chromatin. TSA did not reverse inhibition of transcription from the rDNA reporter by retinoblastoma (Rb) protein, suggesting that the main mechanism by which Rb blocks rDNA transcription may not involve recruitment of deacetylases to rDNA chromatin. Overexpression of histone transacetylases p300, CBP and PCAF stimulated transcription in transfected NIH 3T3 cells. Recombinant p300, but not PCAF, stimulated rDNA transcription in vitro in the absence of nucleosomes, suggesting that the stimulation of rDNA transcription by TSA might have a chromatin-independent component. We found that the rDNA transcription factor UBF was acetylated in vivo. Finally, we also demonstrated the nucleolar localization of CBP. Our results suggest that the organization of ribosomal chromatin of higher eukaryotes is not static and that acetylation may be involved in affecting these dynamic changes directly through histone acetylation and/or through acetylation of UBF or one of the other components of rDNA transcription.

3T3 Cells↗
PubMed1999

Identification of a mammalian RNA polymerase I holoenzyme containing components of the DNA repair/replication system.

Traditional models for transcription initiation by RNA polymerase I include a stepwise assembly of basic transcription factors/regulatory proteins on the core promoter to form a preinitiation complex. In contrast, we have identified a preassembled RNA polymerase I (RPI) complex that contains all the factors necessary and sufficient to initiate transcription from the rDNA promoter in vitro. The purified RPI holoenzyme contains the RPI homolog of TFIID, SL-1 and the rDNA transcription terminator factor (TTF-1), but lacks UBF, an activator of rDNA transcription. Certain components of the DNA repair/replication system, including Ku70/80, DNA topoisomerase I and PCNA, are also associated with the RPI complex. We have found that the holo-enzyme supported specific transcription and that specific transcription was stimulated by the RPI transcription activator UBF. These results support the hypothesis that a fraction of the RPI exists as a preassembled, transcriptionally competent complex that is readily recruited to the rDNA promoter, i.e. as a holoenzyme, and provide important new insights into the mechanisms governing initiation by RPI.

Animals↗
PubMed1998

Affinity purification of mammalian RNA polymerase I. Identification of an associated kinase.

Overlapping cDNA clones encoding the two largest subunits of rat RNA polymerase I, designated A194 and A127, were isolated from a Reuber hepatoma cDNA library. Analyses of the deduced amino acid sequences revealed that A194 and A127 are the homologues of yeast A190 and A135 and have homology to the beta' and beta subunits of Escherichia coli RNA polymerase I. Antibodies raised against the recombinant A194 and A127 proteins recognized single proteins of approximately 190 and 120 kDa on Western blots of total cellular proteins of mammalian origin. N1S1 cell lines expressing recombinant His-tagged A194 and FLAG-tagged A127 proteins were isolated. These proteins were incorporated into functional RNA polymerase I complexes, and active enzyme, containing FLAG-tagged A127, could be immunopurified to approximately 80% homogeneity in a single chromatographic step over an anti-FLAG affinity column. Immunoprecipitation of A194 from 32P metabolically labeled cells with anti-A194 antiserum demonstrated that this subunit is a phosphoprotein. Incubation of the FLAG affinity-purified RNA polymerase I complex with [gamma-32P]ATP resulted in autophosphorylation of the A194 subunit of RPI, indicating the presence of associated kinase(s). One of these kinases was demonstrated to be CK2, a serine/threonine protein kinase implicated in the regulation of cell growth and proliferation.

Amino Acid Sequence↗
PubMed1993

Lipoprotein subclasses in genetic studies: the Berkeley data set.

In conjunction with a study examining the inheritance of LDL subclass patterns in a healthy population, measurements of lipids, lipoproteins, and lipoprotein subclasses were performed in 301 individuals in 27 kindreds. Questionnaires were used to obtain information on use of medications, hormones, cigarettes, and alcohol. Laboratory data from this study (the Berkeley data set) include measurements of LDL and HDL size subclasses by nondenaturing gradient gel electrophoresis, and measurement of apolipoprotein A-I by radial immunodiffusion.

Adult↗