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1.  Publishing confirming and non-confirming data 
F1000Research  2016;5:135.
This editorial introduces the Preclinical Reproducibility and Robustness channel on F1000Research, which has been created to encourage and facilitate open and transparent publication and discussion of confirmatory and non-confirmatory studies in biomedical research.
PMCID: PMC4786892  PMID: 26998234
Reproducibility; Robustness; Amgen
2.  Sphingosine Kinase Activity Is Not Required for Tumor Cell Viability 
PLoS ONE  2013;8(7):e68328.
Sphingosine kinases (SPHKs) are enzymes that phosphorylate the lipid sphingosine, leading to the formation of sphingosine-1-phosphate (S1P). In addition to the well established role of extracellular S1P as a mitogen and potent chemoattractant, SPHK activity has been postulated to be an important intracellular regulator of apoptosis. According to the proposed rheostat theory, SPHK activity shifts the intracellular balance from the pro-apoptotic sphingolipids ceramide and sphingosine to the mitogenic S1P, thereby determining the susceptibility of a cell to apoptotic stress. Despite numerous publications with supporting evidence, a clear experimental confirmation of the impact of this mechanism on tumor cell viability in vitro and in vivo has been hampered by the lack of suitable tool reagents. Utilizing a structure based design approach, we developed potent and specific SPHK1/2 inhibitors. These compounds completely inhibited intracellular S1P production in human cells and attenuated vascular permeability in mice, but did not lead to reduced tumor cell growth in vitro or in vivo. In addition, siRNA experiments targeting either SPHK1 or SPHK2 in a large panel of cell lines failed to demonstrate any statistically significant effects on cell viability. These results show that the SPHK rheostat does not play a major role in tumor cell viability, and that SPHKs might not be attractive targets for pharmacological intervention in the area of oncology.
PMCID: PMC3702540  PMID: 23861887
3.  Molecular Signatures of the Primitive Prostate Stem Cell Niche Reveal Novel Mesenchymal-Epithelial Signaling Pathways 
PLoS ONE  2010;5(9):e13024.
Signals between stem cells and stroma are important in establishing the stem cell niche. However, very little is known about the regulation of any mammalian stem cell niche as pure isolates of stem cells and their adjacent mesenchyme are not readily available. The prostate offers a unique model to study signals between stem cells and their adjacent stroma as in the embryonic prostate stem cell niche, the urogenital sinus mesenchyme is easily separated from the epithelial stem cells. Here we investigate the distinctive molecular signals of these two stem cell compartments in a mammalian system.
Methodology/Principal Findings
We isolated fetal murine urogenital sinus epithelium and urogenital sinus mesenchyme and determined their differentially expressed genes. To distinguish transcripts that are shared by other developing epithelial/mesenchymal compartments from those that pertain to the prostate stem cell niche, we also determined the global gene expression of epidermis and dermis of the same embryos. Our analysis indicates that several of the key transcriptional components that are predicted to be active in the embryonic prostate stem cell niche regulate processes such as self-renewal (e.g., E2f and Ap2), lipid metabolism (e.g., Srebp1) and cell migration (e.g., Areb6 and Rreb1). Several of the enriched promoter binding motifs are shared between the prostate epithelial/mesenchymal compartments and their epidermis/dermis counterparts, indicating their likely relevance in epithelial/mesenchymal signaling in primitive cellular compartments. Based on differential gene expression we also defined ligand-receptor interactions that may be part of the molecular interplay of the embryonic prostate stem cell niche.
We provide a comprehensive description of the transcriptional program of the major regulators that are likely to control the cellular interactions in the embryonic prostatic stem cell niche, many of which may be common to mammalian niches in general. This study provides a comprehensive source for further studies of mesenchymal/epithelial interactions in the prostate stem cell niche. The elucidation of pathways in the normal primitive niche may provide greater insight into mechanisms subverted during abnormal proliferative and oncogenic processes. Understanding these events may result in the development of specific targeted therapies for prostatic diseases such as benign prostatic hypertrophy and carcinomas.
PMCID: PMC2948007  PMID: 20941365
4.  Molecular Signatures of Prostate Stem Cells Reveal Novel Signaling Pathways and Provide Insights into Prostate Cancer 
PLoS ONE  2009;4(5):e5722.
The global gene expression profiles of adult and fetal murine prostate stem cells were determined to define common and unique regulators whose misexpression might play a role in the development of prostate cancer.
Methodology/Principal Findings
A distinctive core of transcriptional regulators common to both fetal and adult primitive prostate cells was identified as well as molecules that are exclusive to each population. Elements common to fetal and adult prostate stem cells include expression profiles of Wnt, Shh and other pathways identified in stem cells of other organs, signatures of the aryl-hydrocarbon receptor, and up-regulation of components of the aldehyde dehydrogenase/retinoic acid receptor axis. There is also a significant lipid metabolism signature, marked by overexpression of lipid metabolizing enzymes and the presence of the binding motif for Srebp1. The fetal stem cell population, characterized by more rapid proliferation and self-renewal, expresses regulators of the cell cycle, such as E2f, Nfy, Tead2 and Ap2, at elevated levels, while adult stem cells show a signature in which TGF-β has a prominent role. Finally, comparison of the signatures of primitive prostate cells with previously described profiles of human prostate tumors identified stem cell molecules and pathways with deregulated expression in prostate tumors including chromatin modifiers and the oncogene, Erg.
Our data indicate that adult prostate stem or progenitor cells may acquire characteristics of self-renewing primitive fetal prostate cells during oncogenesis and suggest that aberrant activation of components of prostate stem cell pathways may contribute to the development of prostate tumors.
PMCID: PMC2684642  PMID: 19478945
5.  Consequences of Nonadaptive Alterations in Cancer 
Molecular Biology of the Cell  2003;14(6):2201-2205.
PMCID: PMC194870  PMID: 12808022
6.  A simple method for statistical analysis of intensity differences in microarray-derived gene expression data 
BMC Biotechnology  2001;1:8.
Microarray experiments offer a potent solution to the problem of making and comparing large numbers of gene expression measurements either in different cell types or in the same cell type under different conditions. Inferences about the biological relevance of observed changes in expression depend on the statistical significance of the changes. In lieu of many replicates with which to determine accurate intensity means and variances, reliable estimates of statistical significance remain problematic. Without such estimates, overly conservative choices for significance must be enforced.
A simple statistical method for estimating variances from microarray control data which does not require multiple replicates is presented. Comparison of datasets from two commercial entities using this difference-averaging method demonstrates that the standard deviation of the signal scales at a level intermediate between the signal intensity and its square root. Application of the method to a dataset related to the β-catenin pathway yields a larger number of biologically reasonable genes whose expression is altered than the ratio method.
The difference-averaging method enables determination of variances as a function of signal intensities by averaging over the entire dataset. The method also provides a platform-independent view of important statistical properties of microarray data.
PMCID: PMC59472  PMID: 11690545
7.  Transcriptional transactivation by selected short random peptides attached to lexA-GFP fusion proteins 
Transcriptional transactivation is a process with remarkable tolerance for sequence diversity and structural geometry. In studies of the features that constitute transactivating functions, acidity has remained one of the most common characteristics observed among native activation domains and activator peptides.
We performed a deliberate search of random peptide libraries for peptides capable of conferring transcriptional transactivation on the lexA DNA binding domain. Two libraries, one composed of C-terminal fusions, the other of peptide insertions within the green fluorescent protein structure, were used. We show that (i) peptide sequences other than C-terminal fusions can confer transactivation; (ii) though acidic activator peptides are more common, charge neutral and basic peptides can function as activators; and (iii) peptides as short as 11 amino acids behave in a modular fashion.
These results support the recruitment model of transcriptional activation and, combined with other studies, suggest the possibility of using activator peptides in a variety of applications, including drug development work.
PMCID: PMC56998  PMID: 11580863
8.  Oligonucleotide-conjugated beads for transdominant genetic experiments 
Nucleic Acids Research  2000;28(2):534-543.
Transdominant genetics using expression libraries can identify proteins and peptides that affect cell division. In conjunction with these libraries, oligonucleotide-conjugated beads and flow cytometry were used to test a strategy that potentially expands the range of such genetic studies. The experimental approach involved creation of tagged expression libraries, introduction of these libraries into cells, growth of the cultured cells for several generations and recovery on oligonucleotide-conjugated beads of sequences that encode growth-modulatory proteins or peptides. Experiments in Saccharomyces cerevisiae demonstrating the feasibility of the strategy are presented.
PMCID: PMC102503  PMID: 10606653

Results 1-8 (8)