Development of a Course-Based Undergraduate Research Experience to Introduce Drug-Receptor Concepts.

Course-based research experiences (CUREs) are currently of high interest due to their potential for engaging undergraduate students in authentic research and maintaining their interest in science, technology, engineering, and mathematics (STEM) majors. As part of a campus-wide initiative called , which is a living learning program offered to freshman STEM majors at the University of Kentucky funded by a grant from Howard Hughes Medical Institute, we have developed a CURE for freshmen interested in pursuing health care careers. Our course, entitled "Drug-Drug Interactions in Breast Cancer," utilized a semester-long, in-class authentic research project and instructor-led discussions to engage students in a full spectrum of research activities, ranging from developing hypotheses and experimental design to generating original data, collaboratively interpreting results and presenting a poster at a campus-wide symposium.

WITHDRAWN: Protein sumoylation and human diseases.

This review has been withdrawn at the request of the author(s) and/or editor. The Publisher apologizes for any inconvenience this may cause. The full Elsevier Policy on Article Withdrawal can be found at http://www.

VDR and CYP27B1 are expressed in C2C12 cells and regenerating skeletal muscle: potential role in suppression of myoblast proliferation.

1α,25(OH)(2)D(3), the active form of vitamin D(3), has been reported to regulate the cell biology of skeletal muscle. However, there has been some controversy about the expression of the vitamin D receptor (VDR) and thus the potential role of vitamin D(3) in skeletal muscle. In this study, we isolated and sequenced the full-length Vdr and Cyp27b1 transcripts in C2C12 myoblasts and myotubes.

SUMO and its role in human diseases.

The covalent attachment of small ubiquition-like modifier (SUMO) polypeptides, or sumoylation, is an important regulator of the functional properties of many proteins. Among these are many proteins implicated in human diseases including cancer and Huntington's, Alzheimer's, and Parkinson's diseases, as well as spinocerebellar ataxia 1 and amyotrophic lateral sclerosis. The results of two more recent studies identify two additional human disease-associated proteins that are sumoylated, amyloid precursor protein (APP), and lamin A.

Detection of proteins sumoylated in vivo and in vitro.

Small ubiquitin-related modifier (SUMO) is an ubiquitin-like protein that is covalently attached to a variety of target proteins. Unlike ubiquitination, sumoylation does not target proteins for proteolytic breakdown, but is instead involved in regulating multiple protein functional properties including protein-protein interactions and subcellular targeting, to name a few. Protein sumoylation has been particularly well characterized as a regulator of many nuclear processes as well as nuclear structure, making the characterization of this modification vital for understanding nuclear structure and function.

Chromosome-wide analysis of protein binding and modifications.

In order to fully understand the functions of a DNA-binding protein it is necessary to identify all of its binding sites in chromosomes and assess the role of each site in the overall biological function of the factor. An approach ChIP-on-Chip which combines the chromatin immunoprecipitation technique with chromosomal DNA microarray analysis, has proven to be a powerful means for the chromosome-wide identification of protein binding sites. This approach can also be used to characterize chromosome-wide variations in patterns of post-translational protein modifications, for example histone modifications.

Sumoylation and human disease pathogenesis.

Covalent modification by SUMO polypeptides, or sumoylation, is an important regulator of the functional properties of many proteins. Among these are several proteins implicated in human diseases including cancer, Huntington's, Alzheimer's, and Parkinson's diseases, as well as spinocerebellar ataxia 1 and amyotrophic lateral sclerosis. Recent reports reveal two new examples of human disease-associated proteins that are SUMO modified: amyloid precursor protein and lamin A.

Mitotic bookmarking of formerly active genes: keeping epigenetic memories from fading.

In order for cell lineages to be maintained, daughter cells must have the same patterns of gene expression as the cells from which they were divided so that they can have the same phenotypes. However, during mitosis transcription ceases, chromosomal DNA is compacted, and most sequence-specific binding factors dissociate from DNA, making it difficult to understand how the "memory" of gene expression patterns is remembered and propagated to daughter cells. The process of remembering patterns of active gene expression during mitosis for transmission to daughter cells is called gene bookmarking.

Detection of sumoylated proteins.

Small ubiquitin-related modifier (SUMO) is an ubiquitin-like protein that is covalently attached to a variety of target proteins. Unlike ubiquitination, sumoylation does not target proteins for proteolytic breakdown, but is instead involved in regulating a variety of different protein functional properties, including protein-protein interactions and subcellular targeting, to name a few. Protein sumoylation has been particularly well characterized as a regulator of many nuclear processes as well as of nuclear structure, making the characterization of this modification vital for understanding nuclear structure and function.

Gene bookmarking: keeping the pages open.

'Gene bookmarking' is a mechanism of epigenetic memory that functions to transmit through mitosis the pattern of active genes and/or genes that can be activated to daughter cells. It is thought that, at a point before mitosis, genes that exist in an open, transcriptionally competent state are bound by proteins or marked by some kind of modification event. This is thought to facilitate the assembly of transcription complexes on the promoters in early G1, thereby ensuring that daughter cells have the same pattern of gene expression as the cell from which they derived.

Transactivation of the parathyroid hormone promoter by specificity proteins and the nuclear factor Y complex.

We previously identified a highly conserved specificity protein 1 (Sp1) DNA element in mammalian PTH promoters that acted as an enhancer of gene transcription and bound Sp1 and Sp3 proteins present in parathyroid gland nuclear extracts. More recently, a nuclear factor (NF)-Y element (NF-Y(prox)) was also described by our group, which was located approximately 30 bp downstream from the Sp1 site in the human PTH (hPTH) promoter and by itself acted as a weak enhancer of gene transcription. We now report that Sp proteins and NF-Y can synergistically enhance transcription of a minimal hPTH promoter construct.

Mechanism of hsp70i gene bookmarking.

In contrast to most genomic DNA in mitotic cells, the promoter regions of some genes, such as the stress-inducible hsp70i gene that codes for a heat shock protein, remain uncompacted, a phenomenon called bookmarking. Here we show that hsp70i bookmarking is mediated by a transcription factor called HSF2, which binds this promoter in mitotic cells, recruits protein phosphatase 2A, and interacts with the CAP-G subunit of the condensin enzyme to promote efficient dephosphorylation and inactivation of condensin complexes in the vicinity, thereby preventing compaction at this site. Blocking HSF2-mediated bookmarking by HSF2 RNA interference decreases hsp70i induction and survival of stressed cells in the G1 phase, which demonstrates the biological importance of gene bookmarking.

Identification of Xenopus heat shock transcription factor-2: conserved role of sumoylation in regulating deoxyribonucleic acid-binding activity of heat shock transcription factor-2 proteins.

Heat shock transcription factor (Hsf)-1 and Hsf2 are members of the heat shock factor (HSF) protein family involved in heat shock protein (hsp) gene regulation, a regulation that is critical for the ability of cells to survive exposure to stress conditions. Although the role of Hsf1 in binding and activating transcription of hsp gene promoters in response to cell stress is well established, how Hsf2 enhances stress-induced hsp expression is not understood. To gain an insight into the critical conserved features of the regulation and function of Hsf2, we have identified and characterized the Hsf2 protein from Xenopus laevis.

Suppression of the human parathyroid hormone promoter by vitamin D involves displacement of NF-Y binding to the vitamin D response element.

An earlier report in the literature indicated the vitamin D response element (VDRE) in the human parathyroid hormone (hPTH) promoter could be specifically bound by an unidentified transcription factor in addition to the vitamin D receptor (VDR) complex. We confirmed that OK and HeLa cell nuclear extracts formed a specific complex with the hPTH VDRE that was insensitive to competition with other VDRE sequences. However, this factor could be competed for by a consensus NF-Y DNA-binding site, and an anti-NF-Y antibody was able to supershift the bound band.

Contrasting mammalian parathyroid hormone (PTH) promoters: nuclear factor-Y binds to a deoxyribonucleic acid element unique to the human PTH promoter and acts as a transcriptional enhancer.

The identification of a highly conserved specificity protein 1 (Sp1) DNA element in mammalian PTH promoters was recently reported. However, the presence of a novel DNA-binding complex was subsequently observed exclusively with the human PTH (hPTH) Sp1 element in mobility shift studies. Point mutations in the hPTH Sp1 element revealed the factor recognized a CAAT-like sequence resulting from a single nucleotide difference unique to the human sequence relative to other mammalian promoters.

HSF1 modulation of Hsp70 mRNA polyadenylation via interaction with symplekin.

Induction of heat shock protein (HSP) gene expression by stress is initiated by binding of HSF1 to HSP gene promoters to increase their transcription. The cytoprotective functions of these HSPs are essential for cell survival, and thus it is critical that inducible HSP gene expression be executed rapidly and efficiently. Here we report an interaction between heat shock factor 1 (HSF1) and symplekin, a protein known to form a complex with the polyadenylation factors CstF and CPSF.

Estradiol represses prolactin-induced expression of Na+/taurocholate cotransporting polypeptide in liver cells through estrogen receptor-alpha and signal transducers and activators of transcription 5a.

Na(+)/taurocholate cotransporting polypeptide (ntcp) mediates the uptake of bile salts from plasma across the basolateral domain of the hepatocyte. We have demonstrated that ntcp expression can be induced by prolactin (PRL) and placental lactogen via the PRL receptor and signal transducers and activators of transcription (Stat)5a pathway. However, elevated levels of placental lactogen do not increase the expression of ntcp in pregnant rats.

Insights into the regulation of heat shock transcription factor 1 SUMO-1 modification.

The transcriptional regulatory protein HSF1 is the key mediator of induced heat shock protein gene expression in response to elevated temperature and other stresses. Our previous studies identified stress-induced SUMO-1 modification of HSF1 as an important regulator of the DNA-binding activity of this factor. The underlying molecular mechanism by which stress leads to sumoylation of HSF1 was unknown.

Lowered temperature set point for activation of the cellular stress response in T-lymphocytes.

The induction of heat shock protein gene expression in response to stress is critical for the ability of organisms to cope with and survive exposure to these stresses. However, most studies on HSF1-mediated induction of hsp70 gene expression have utilized immortalized cell lines and temperatures above the physiologically relevant range. For these reasons much less is known about the heat shock response as it occurs in mammalian cells within tissues in the intact organism.