Roger Chalkley

Vanderbilt University | Vander Bilt · Department of Molecular Physiology and Biophysics

This study examines the intersectional role of citizenship and gender with career self-efficacy amongst 10,803 doctoral and postdoctoral trainees in US universities. These biomedical trainees completed surveys administered by 17 US institutions that participated in the National Institutes of Health Broadening Experiences in Scientific Training (NIH...

Biomedical sciences PhDs pursue a wide range of careers inside and outside academia. However, there is little data regarding how career interests of PhD students relate to the decision to pursue postdoctoral training or to their eventual career outcomes. Here, we present the career goals and career outcomes of 1452 biomedical sciences PhDs who grad...

Biomedical sciences PhDs pursue a wide range of careers inside and outside academia. However, there is little data regarding how career interests of PhD students relate to the decision to pursue postdoctoral training or to their eventual career outcomes. Here, we present the career goals and career outcomes of 1,452 biomedical sciences PhDs who gra...

The present study examines racial, ethnic, and gender disparities in career self-efficacy amongst 6077 US citizens and US naturalized graduate and postdoctoral trainees. Respondents from biomedical fields completed surveys administered by the National Institutes of Health Broadening Experiences in Scientific Training (NIH BEST) programs across 17 U...

This study examines the intersectional role of citizenship and gender with career self-efficacy amongst 10,803 doctoral and postdoctoral trainees in US universities. These biomedical trainees completed surveys administered by 17 US institutions that participated in the National Institutes of Health Broadening Experiences in Scientific Training (NIH...

The present study examines racial, ethnic, and gender disparities in career self-efficacy amongst 6077 US citizens and US naturalized graduate and postdoctoral trainees. Respondents from biomedical fields completed surveys administered by the National Institutes of Health Broadening Experiences in Scientific Training (NIH BEST) programs across 17 U...

COVID-19-associated university closures moved classes online and interrupted ongoing research in universities throughout the US. In Vanderbilt University, first year biomedical sciences PhD students were in the middle of their spring semester coursework and in the process of identifying a thesis research lab, while senior students who had already c...

PhD-trained scientists are essential contributors to the workforce in diverse employment sectors that include academia, industry, government, and nonprofit organizations. Hence, best practices for training the future biomedical workforce are of national concern. Complementing coursework and laboratory research training, many institutions now offer...

PhD-trained scientists are essential contributors to the workforce in diverse employment sectors that include academia, industry, government, and non-profit organizations. Hence, best practices for training the future biomedical workforce are of national concern. Complementing coursework and laboratory research training, many institutions now offer...

The association between GRE scores and academic success in graduate programs is currently of national interest. GRE scores are often assumed to be predictive of student success in graduate school. However, we found no such association in admission data from Vanderbilt’s Initiative for Maximizing Student Diversity (IMSD), which recruited historicall...

The Broadening Experiences in Scientific Training (BEST) program is an NIH-funded effort testing the impact of career development interventions (e.g. internships, workshops, classes) on biomedical trainees (graduate students and postdoctoral fellows). BEST Programs seek to increase trainees’ knowledge, skills and confidence to explore and pursue ex...

MSU Survey. Survey items were gather into factors in which each of the questions were weighted equally to provide a final outcome for that factor. Factors/scales are in BOLD. Other prefatory questions concerning faculty rank, gender, years spent in graduate training, years spent in postdoctoral training (including 0 as a possible answer), employmen...

Vanderbilt University survey. * Questions were answered but are not included in the results of this present study. 1. Prior to taking this survey, were you aware of your institution's BEST program? Potential Answers: Yes, I was aware of my institution's BEST program and what it does. Yes, I was aware but I didn't know much about my institution's BE...

The association between GRE scores and academic success in graduate programs is currently of national interest. GRE scores are often assumed to be predictive of student success in graduate school. However, we found no such association in admission data from Vanderbilt's Initiative for Maximizing Student Diversity (IMSD), which recruited historicall...

Several reports have shown that doctoral and postdoctoral trainees in biomedical research pursue diverse careers that advance science meaningful to society. Several groups have proposed 3-tier career taxonomy to showcase these outcomes. This 3-tier taxonomy will be a valuable resource for institutions committed to greater transparency in reporting...

There is a persistent shortage of underrepresented minority (URM) faculty who are involved in basic biomedical research at medical schools. We examined the entire training pathway of potential candidates to identify the points of greatest loss. Using a range of recent national data sources, including the National Science Foundation’s Survey of Earn...

Percentage of students who complete the bachelor’s degree in biological sciences after expressing an interest in the area at matriculation. As seen in the synthetic cohort, Whites who express an interest in biological sciences are nearly twice as likely to complete a bachelor’s degree in biological sciences four years later than URM students. (TIFF...

Summary of data sources used for tracking educational pathways. (PDF)

Percent of first-year college students who express an interest in biological or agricultural sciences. For all ethnic groups, there is an increasing amount of interest in the biological sciences upon matriculation into college. Black, Latino, and White students express similar interest in this area. (TIFF)

Historically, admissions committees for biomedical Ph.D. programs have heavily weighed GRE scores when considering applications for admission. The predictive validity of GRE scores on graduate student success is unclear, and there have been no recent investigations specifically on the relationship between general GRE scores and graduate student suc...

Details regarding the sample and additional results tables. (DOCX)

Recent national reports and commentaries on the current status and needs of the U.S. biomedical research workforce have highlighted the limited career development opportunities for predoctoral and postdoctoral trainees in academia, yet little attention is paid to preparation for career pathways outside of the traditional faculty path. Recognizing t...

The Medical College Admission Test (MCAT) is a quantitative metric used by MD and MD-PhD programs to evaluate applicants for admission. This study assessed the validity of the MCAT in predicting training performance measures and career outcomes for MD-PhD students at a single institution. The study population consisted of 153 graduates of the Vande...

The past few years have placed scientific workforce pressures in the limelight: National committees, academic thought-leaders, trainees, politicians, and the popular press have highlighted, in reports and essays, issues such as the increasing numbers of Ph.D.-trained scientists, the shrinking supply

Hypoxia causes up-regulation and activation of xanthine dehydrogenase/xanthine oxidase (XDH/XO) in vitro and in the lungs in vivo. This up-regulation, and the likely corresponding production of reactive oxygen species, may underlie the pathogenesis of an array of disorders. Thus, compounds that prevent hypoxia-induced increase in XDH/XO activity ma...

Tobacco smoking has been causally linked to the development of chronic obstructive pulmonary disease. It has been reported that the reactive oxygen species (ROS)- generating enzyme xanthine dehydrogenase/oxidase (XO) is increased in smoking-related stomach ulcers and that gastric mucosal damage caused by tobacco smoke can be blocked by the XO inhib...

The purpose of this study is to design and develop educational technology tools, primarily using the World Wide Web, for effectively recruiting young scientists to Vanderbilt University School of Medicine. We have taken the position that technology would offer major advantages in administration, teaching and learning, and have embraced this approac...

We used an allelogenic Cre/loxP gene targeting strategy in mice to determine the role of cytosolic phosphoenolpyruvate carboxykinase (PEPCK) in hepatic energy metabolism. Mice that lack this enzyme die within 3 days of birth, while mice with at least a 90% global reduction of PEPCK, or a liver-specific knockout of PEPCK, are viable. Surprisingly, i...

The kidney-specific chromatin structure of the phosphoenolpyruvate carboxykinase (PEPCK) gene was examined and compared to that of the liver. Kidney nuclear extracts were found to lack a liver-enriched factor, pepA, that binds to HSS A, a distal enhancer of the PEPCK gene that may be involved in opening the chromatin domain of the PEPCK gene in the...

A 72 kilobase pair DNA fragment that contains the mouse phosphoenolpyruvate carboxykinase (PEPCK) gene locus, pck1, was isolated from a genomic bacterial artificial chromosome library. The region from approximately -5.5 to +6.6 kilobase pairs relative to the pck1 transcription start site was sequenced and exhibits a high degree of homology to the r...

We previously reported that the TATA-less rat xanthine dehydrogenase/oxidase (XDH/XO) promoter is organized with multiple initiator elements (Inr 1, 2, 3 and 4). Additionally, we identified six factor binding footprints in the upstream region of this promoter (FP 1-FP 6), two of which (FP 2 and FP 4) we showed to be C/EBP binding sites. In this rep...

In the present study, we have shown that a downstream element located in the coding region of the TATA-less rat xanthine dehydrogenase/oxidase (XDH/XO) gene (-7 to +42) plays an important role in transcription initiation and C/EBP transcriptional activation. Previous work from our laboratory has shown that the promoter is organized with multiple in...

Studies of the phosphoenolpyruvate carboxykinase (PEPCK) gene in transgenic mice have shown that distinct regulatory elements control transcription of this gene in the liver and kidney. While proximal promoter and upstream sequences are sufficient in the liver, expression in the kidney requires unidentified elements located in the coding region or...

In the present study, we have explored an unexpected observation in transcription initiation that is mediated by single-stranded oligonucleotides. Initially, our goal was to understand the function of different upstream regulatory elements/initiation sites in the rat xanthine dehydrogenase/oxidase (XDH/XO) promoter. We performed in vitro transcript...

A bacterial artificial chromosome librarv, derived from mouse genornic DNA (strain 129 SV). was screened with a mouse PEPCK cDNA fragment. One positive clone. >25 kbp, was isolated. Sequence from this clone was identical in the. mouse cDNA, confirming thai the locus isolated contains the mouse PKPCK gene. Comparison of muPEPCK to the rat gene shows...

Two regions of D Nase 1 hypersensitivity have been mapped in the PEPCK gene in the rat kidney. One spans a series of protein binding elements in the proximal promoter. The other, named HSS E. is located 3" of the polyadenylation site and has been observed in both PK PC K-expressing and non- expressing cells. We are studying the role of HSS Ein PEPC...

Glucokinase (GK) gene transcription occurs in the liver and the beta cell of the endocrine pancreas where it is subject to different modes of regulation. This is accomplished largely through the use of two linked, cell-specific promoters separated by at least 12 kbp. We have used DNase I hypersensitivity to explore the chromatin structure surroundi...

In the present study, we have explored further the organization of the TATA-less rat xanthine dehydrogenase/oxidase gene (XDH/XO). A DNase I hypersensitive site has been identified which it colocalizes with the basal promoter reported previously [Chow et al. (1994) Nucleic Acids Res., 22, 1846-1854]. Gel mobility shift assays indicate the presence...

We have analyzed site occupancy in vivo with constructs containing one or more copies of the binding site for the yeast trans-activator, yIBF. The data indicate only a modest difference in site occupancy (at most 2-fold) even though multiple copies activate transcription several hundredfold more than one copy. Using studies in which we have mutated...

Inflammation and ischemia--reperfusion tissue injury are important pathophysiologic processes with a wide spectrum of clinical presentations; the enzyme xanthine dehydrogenase/oxidase (XDH/XO) is thought to play a key role in ischemia--reperfusion injury. Recent studies have shown the transcriptional regulation of XDH/XO by cytokines (Dupont et al....

We have studied the competitive binding of histones and the Rous sarcoma virus internal enhancer binding factor (IBF) factor (which recent studies indicate is almost certainly cEBP beta). We find that histones and IBF are incapable of forming a ternary complex with a 159-base pair (bp) fragment of DNA containing a single IBF binding site and that h...

Transcription initiation of the gene encoding phosphoenolpyruvate carboxykinase (PEPCK) is stimulated by glucocorticoids and glucagon, via cAMP, and dominantly inhibited by insulin in rat liver and H4IIE cells. Lysolecithin-permeabilized H4IIE cells recover completely and continue to multiply, yet are transiently penetrable by macromolecules. These...

Genomic Southern blot analysis of rat EFIA (gene encoding enhancer factor I subunit A) reveals a complex band pattern when cDNA subfragment probes are used. Screening a rat genomic library with a rat EFIA cDNA probe yields two different processed EFIA pseudogenes, designated rat psi EFIA#(2/3) and #(4/7), in addition to two other different, but les...

The genomic copy multiplicity of the CCAAT transcription complex component enhancer factor I subunit A (EFIA) has been examined. When a mammalian genomic Southern blot was hybridized to a rat EFIA cDNA, a complex pattern consisting of numerous related sequences was found in all the species examined, with Bos taurus being the least complex. An EFIA#...

A previously described upstream hypersensitive site (HS) in the PEPCK gene at -4800 bp, termed HS A (1), has been characterized and determined to bind at least two factors. One of these is a member of the ubiquitous CREB/ATF family, and the second is a novel tissue specific protein, pep A. A construct carrying HS A and the PEPCK proximal promoter w...

We have identified DNA elements In the phosphoenolpyruvate carboxykinase (PEPCK) gene promoter which are bound ‘in vivo’ by proteins under conditions of basal level gene expression and have evaluated several hypothesis to account for the tissue specific expression of the gene. in vitro DNase I footprintlng demonstrated that factors which bind to ba...

Enhancer factor I (EFI) is a trans-acting factor which binds to the Rous sarcoma virus long terminal repeat enhancer and promoter at two inverted CCAAT-box motifs. We demonstrate that two forms of EFI DNA binding activity exist in nuclear extracts of avian cells. One form requires two heterologous components (EFIA)(EFIB) for high affinity, specific...

We have previously identified a series of five DNase-I hypersensitive (HS) sites within and around the rat phosphoenolpyruvate carboxykinase (PEPCK) gene. The far upstream region has now been sequenced, and the tissue-specific HS site has been mapped more precisely at 4,800 base pairs upstream of the transcription start site of the PEPCK gene. DNA...

We have analyzed the chromatin structure of the phosphoenolpyruvate carboxykinase (PEPCK) gene in hepatoma x fibroblast hybrids with different extinction phenotypes. These hybrids included a karyotypically complete hybrid in which all liver gene activity was extinguished, a microcell hybrid that contained a single mouse chromosome 11 and in which P...

We have analyzed the chromatin structure of the phosphoenolpyruvate carboxykinase (PEPCK) gene in hepatoma x fibroblast hybrids with different extinction phenotypes. These hybrids included a karyotypically complete hybrid in which all liver gene activity was extinguished, a microcell hybrid that contained a single mouse chromosome 11 and in which P...

We have previously identified a series of five DNase-I hypersensitive (HS) sites within and around the rat phosphoenolpyruvate carboxykinase (PEPCK) gene. The far upstream region has now been sequenced, and the tissue-specific HS site has been mapped more precisely at 4,800 base pairs upstream of the transcription start site of the PEPCK gene. DNA...

The Saccharomyces cerevisiae 5S rRNA gene was used as a model system to study the requirements for assembling transcriptionally active chromatin in vitro with purified components. When a plasmid containing yeast 5S rDNA was assembled into chromatin with purified core histones, the gene was inaccessible to the yeast class III gene transcription mach...

A cis-acting transcriptional activation sequence (IES2) from the Rous sarcoma virus internal enhancer was found to stimulate transcription of a heterologous gene in Saccharomyces cerevisiae. A hamster protein (termed IBF) which binds to IES2 and stimulates transcription in vitro has previously been purified and was found to have a subunit molecular...

The Saccharomyces cerevisiae 5S rRNA gene was used as a model system to study the requirements for assembling transcriptionally active chromatin in vitro with purified components. When a plasmid containing yeast 5S rDNA was assembled into chromatin with purified core histones, the gene was inaccessible to the yeast class III gene transcription mach...

Much effort has been expended towards understanding the details of how nucleosomes are established on newly replicated DNA. More recently it has begun to be possible to study the binding of both trans-acting factors and histones to DNA. This review is concerned with an assessment of the current status of this work. In addition, we discuss some of t...

The internal enhancer binding factor (IBF) that specifically binds sequences within the gag gene internal enhancer of Rous sarcoma virus Schmidt-Ruppin A was purified to near homogeneity from BHK cells. The polypeptides that constituted IBF DNA-binding activity were identified by sodium dodecyl sulfate-polyacrylamide gel analysis. As isolated from...

The internal enhancer binding factor (IBF) that specifically binds sequences within the gag gene internal enhancer of Rous sarcoma virus Schmidt-Ruppin A was purified to near homogeneity from BHK cells. The polypeptides that constituted IBF DNA-binding activity were identified by sodium dodecyl sulfate-polyacrylamide gel analysis. As isolated from...

We used indirect end labeling to identify a series of five hypersensitive (HS) sites in the phosphoenolpyruvate carboxykinase (PEPCK) gene in H4IIE rat hepatoma cells. These sites were found at -4800 base pairs (bp) (site A), at -1300 bp (site B), over a broad domain between -400 and -30 bp (site C), at +4650 bp (site D), and at +6200 bp (site E)....

We used indirect end labeling to identify a series of five hypersensitive (HS) sites in the phosphoenolpyruvate carboxykinase (PEPCK) gene in H4IIE rat hepatoma cells. These sites were found at -4800 base pairs (bp) (site A), at -1300 bp (site B), over a broad domain between -400 and -30 bp (site C), at +4650 bp (site D), and at +6200 bp (site E)....

The purification of trans-acting regulatory proteins has been studied extensively. These agents modulate the transcriptional activity of genes, though their mechanism of action is not understood. As a consequence, there has been renewed interest in reassembling chromatin containing these factors to assay the interrelationship between structure and...

We have developed a method for the separation of transcriptionally engaged chromatin from inactive genes as well as from active genes which are not being transcribed. This approach is dependent upon the integrity of the growing transcript and is reflected in a significant decrease in the density of the chromatin during transcription. The decrease i...

We have purified a nucleoplasmin-like protein from the nuclei of somatic Xenopus laevis cells. This protein possesses a number of the distinctive features of nucleoplasmin isolated from oocytes or unfertilized eggs. The protein is recognized by both monoclonal and polyclonal antisera raised against egg nucleoplasmin. The protein has an oligomeric s...

We have documented that the Rous sarcoma virus (RSV) internal enhancer functions in the nontransformed Baby Hamster Kidney (BHK) cell line. The sequences within this region were assayed for their ability to bind to specific factors present in BHK nuclear extracts using the gel retardation assay and DNAse I footprinting. At least two sequences withi...

We have purified a nucleoplasmin-like protein from the nuclei of somatic Xenopus laevis cells. This protein possesses a number of the distinctive features of nucleoplasmin isolated from oocytes or unfertilized eggs. The protein is recognized by both monoclonal and polyclonal antisera raised against egg nucleoplasmin. The protein has an oligomeric s...

We have observed that ethanol precipitation and subsequent drying of small (<400 bp) radiolabelled DNA fragments is able to induce a transition to a form that migrates aberrantly on acrylamide gels. This unusual form has increased sensitivity to S1 nuclease, decreased sensitivity to restriction enzymes, and a concentration dependence for the revers...

Bleomycin, an important chemotherapeutic agent useful in the treatment of testicular carcinoma, can cause lung parenchymal injury. Prior studies showing that different cell types have different susceptibilities to bleomycin suggest that intracellular conditions for bleomycin-DNA interactions vary. The purpose of the present studies was to test the...

Novobiocin concentrations normally used to inhibit a putative eukaryotic DNA gyrase have been found to inhibit transcription of a yeast 5S rRNA gene using an in vitro yeast transcription system. Purified RNA polymerase III and three yeast transcription factors (chromatographically separated, partially purified and free of any detectable gyrase acti...

We used the sensitive gel electrophoresis DNA-binding assay and DNase I footprinting to detect at least two protein factors (EFI and EFII) that bound specifically to the Rous sarcoma virus (RSV) enhancer in vitro. These factors were differentially extracted from quail cell nuclei, recognized different nucleotide sequences in the U3 region of the RS...

We used the sensitive gel electrophoresis DNA-binding assay and DNase I footprinting to detect at least two protein factors (EFI and EFII) that bound specifically to the Rous sarcoma virus (RSV) enhancer in vitro. These factors were differentially extracted from quail cell nuclei, recognized different nucleotide sequences in the U3 region of the RS...

Novobiocin, an inhibitor of prokaryotic DNA gyrase and eukaryotic type II topoisomerase enzymes, interferes with in vitro chromatin assembly using purified histones, DNA and nucleoplasmin. The target of inhibition is not topoisomerase II; this energy-independent assembly system lacks any ATP and Mg2+-dependent type II topoisomerase or gyrase activi...

Nucleoplasmin isolated from unfertilized Xenopus laevis eggs possesses an in vitro chromatin assembly activity which is superior to nucleoplasmin isolated from oocytes. It is demonstrated here that the two forms of the protein differ in the amount of attached phosphate, with the egg protein possessing nearly 20 phosphate groups per protein monomer...

The cell line 4IC6, adapted for growth in 6 mM sodium butyrate from Hepatoma Tissue Culture cells [R. Chalkley, and A. Shires (1985) J. Biol. Chem. 260, 7698-7704], exhibits a fourfold increase in histone acetate turnover. The 4IC6 cells were about 25 times more resistant to butyrate relative to the parental cell line as measured by cloning efficie...

At concentrations normally used to inhibit eukaryotic type II topoisomerase activity (100–1000 μg/ml) novobiocin binds core histones. Approximately 15 moles of novobiocin bind per mole of histone resulting in histone precipitation from solution in either 0.15 M or 2 M NaCl. The interaction between novobiocin and proteins appears to involve arginine...

Nucleoplasmin has been purified from either oocytes or unfertilized eggs of the frog, Xenopus laevis. We find that the pentameric form of egg nucleoplasmin exhibits an apparent molecular mass approximately 15 000 daltons larger than its oocyte counterpart upon sodium dodecyl sulfate (SDS)-acrylamide gel electrophoresis. Egg nucleoplasmin monomers a...

The expression of a vascular smooth muscle specific alpha-actin isoform can be induced in mouse BC3H1 smooth muscle cells by treating confluent monolayers with serum-free medium (Strauch, A. R., and Rubenstein, P. A. (1984) J. Biol. Chem. 259, 3152-3159; 7224-7229). Using blot hybridization techniques, two size classes of actin RNA were identified...

At concentrations normally used to inhibit eukaryotic type II topoisomerase activity (100-1000 micrograms/ml) novobiocin binds core histones. Approximately 15 moles of novobiocin bind per mole of histone resulting in histone precipitation from solution in either 0.15 M or 2 M NaCl. The interaction between novobiocin and proteins appears to involve...

Hepatoma tissue culture cells were synchronized in G1 and in S phase in order to examine the level of synthesis of different histone types and to determine the rate, timing, and location of their deposition onto DNA. We observe a basal level of synthesis in G1 (5% of that seen in S phase) for H2A.1, H2A.2, H3.2, H2B, and H4. The minor histone varia...

We have reinvestigated the mode of segregation of preexisting histones onto replicating chromosomes. Since our previous data have indicated that only histones H3 and H4 do not appear to move from their association with the DNA strand with which they are bound until the next round of replication, we have concentrated our attention on these two histo...

We have asked whether histones synthesized in the absence of DNA synthesis can exchange into nucleosomal structures. DNA synthesis was inhibited by incubating hepatoma tissue culture cells in medium containing 5.0 mM hydroxyurea for 40 min. During the final 20 min, the cells were pulsed with [3H]lysine to radiolabel the histones (all five histones...

We have obtained a number of variant HTC cells which are capable of vigorous replication in the presence of 6 mM sodium butyrate. These cells show characteristic changes in histone acetylation. H2A/H2B are no longer modified and the turnover of histones H3/H4 acetate is about 4-fold greater than in control HTC cells at the same butyrate concentrati...

We have examined the effect of histone acetylation on the in vitro assembly of nucleosomes with DNA and purified histones at physiological ionic strength in the presence of polyglutamic acid. We have found that hyperacetylated histones assemble nucleosomes with greater efficiency, and to a greater extent, than either control or hypoacetylated histo...

HTC cells have been labeled by short exposures to [3H]thymidine in order to identify newly synthesized DNA. By either isolating nuclei directly or isolating them after an extensive fixation with formaldehyde, we have been able to identify two phases in the maturation process of newly replicated chromatin. The first phase which is relatively brief (...

We have asked whether exogenous, radiolabeled histones can exchange with nucleosomal histones in an in vitro system. Using two different electrophoretic techniques, we were able to separate the histones contained in nucleosomes from those histones which were simply bound to the surface of the chromatin. Fluorography was used to determine which of t...

We have found that butyrate selectively inhibits hormonal induction of a few specific proteins and messenger RNAs in hepatoma cells. The fatty acid salt reversibly abolishes induction of tyrosine aminotransferase by dexamethasone and dibutyryl cyclic AMP in HTC cells by inhibiting the production of tyrosine aminotransferase messenger RNA. Half-maxi...

We have found that butyrate selectively inhibits hormonal induction of a few specific proteins and messenger RNAs in hepatoma cells. The fatty acid salt reversibly abolishes induction of tyrosine aminotransferase by dexamethasone and dibutyryl cyclic AMP in HTC cells by inhibiting the production of tyrosine aminotransferase messenger RNA. Half-maxi...

We have analyzed the RNA synthetic activity in intact HTC cells which have artificially high or low levels of histone acetylation. Rates of total RNA synthesis and the number of nascent transcription complexes have been measured for 20-h butyrate-treated cells and for butyrate-released cells which contain hypoacetylated histones. Exposure of HTC ce...

The effect of histone acetylation on chromatin solubility has been studied. Nucleosome cores are fairly soluble over a range of Mg2+ and Na+ concentrations. Increasing the amount of Na+ relative to a fixed concentration of Mg2+ leads to increased solubility. Although the effect is not large, acetylated cores are more soluble than control cores, all...

Exposure of HTC cells to 6 mM sodium butyrate for 20 h results in significantly reduced amounts of nuclear histone acetyltransferase. While the direct butyrate inhibition of histone deacetylases is rapidly reversed upon removal of cells from butyrate, histone acetyltransferase activity increases only gradually following release of cells from butyra...

We have studied histone acetylation in chicken erythrocytes. We find that about 30% of the histone in these cells is acetylated, however the majority of these histones are not in a dynamic steady state typical of other chicken cells and of mammalian cells, but rather are frozen in this state of modification. A very small fraction of erythrocyte his...

Formaldehyde fixation of simian virus 40 (SV40)-infected CV-1 cells at appropriate times after infection permits us to isolate crosslinked complexes of SV40 minichromosomes during the time of DNA replication and during packaging with viral proteins. Such crosslinked complexes can be separated on the basis of density on CsCl/guanidine . HCl density...

The chromosomal fragments produced by nuclease digestion of freshly replicated chromatin migrate more rapidly relative to bulk chromatin when analyzed in nucleoprotein gels. The cause of the anomolous migration has been studied and the evidence indicates that rather than reflecting a shorter nucleosomal repeat in vivo that it may be a consequence o...

In this study, we have density-labeled newly replicated DNA in hepatoma tissue culture cells and separated the newly replicated nucleoprotein from bulk material using density gradient centrifugation. These experiments indicate that only newly synthesized histones H3 and H4 deposit specifically on newly replicated DNA. Histones H2A and H2B show a pa...

We have examined the effects of histone hyperacetylation upon nuclease digestion of nuclei and subsequent fractionation of chromosomal material in the presence of MgCl2. DNase I shows a maximum sensitivity towards hyperacetylated nuclei at somewhat elevated ionic strengths (150-200 mM NaCl), whereas micrococcal nuclease exhibits no specificity for...

We have developed a new method for isolating subcellular components after fixation of whole cells with formaldehyde. By a number of criteria we establish that the fixation does not alter or cause rearrangement of nucleosomal structure of either newly replicated or old chromatin. Using this approach we can almost completely resolve newly replicated...