Biomedical subjects

P S Nelson

Publications and source records attributed to P S Nelson.

At least 19 recordsLinked to original sources

PubMed2000

PART-1: a novel human prostate-specific, androgen-regulated gene that maps to chromosome 5q12.

Genes regulated by androgenic hormones are of critical importance for the normal physiological function of the human prostate gland, and they contribute to the development and progression of prostate carcinoma. We used cDNA microarrays containing 1500 prostate-derived cDNAs to profile transcripts regulated by androgens in prostate cancer cells. This study identified a novel gene that we have designated PART-1 (prostate androgen-regulated transcript 1), which exhibited increased expression upon exposure to androgens in the LNCaP prostate cancer cell line. Northern analysis demonstrated that PART-1 is highly expressed in the prostate gland relative to other normal human tissues and is expressed as different transcripts using at least three different polyadenylation signals. The PART-1 cDNA and putative protein are not significantly homologous to any sequences in the nonredundant public sequence databases. Cloning and analysis of the putative PART-1 promoter region identified a potential binding site for the homeobox gene PBX-la, but no consensus androgen response element or sterol-regulatory element binding sites were identified. We used a radiation hybrid panel and fluorescence in situ hybridization to map the PART-1 gene to chromosome 5q12, a region that has been suggested to harbor a prostate tumor suppressor gene. These results identify a new gene involved in the androgen receptor-regulated gene network of the human prostate that may play a role in the etiology of prostate carcinogenesis.

Amino Acid Sequence↗

PubMed2000

The prostate expression database (PEDB): status and enhancements in 2000.

The Prostate Expression Database (PEDB) is an online resource designed to access and analyze gene expression information derived from the human prostate. PEDB archives >55 000 expressed sequence tags (ESTs) from 43 cDNA libraries in a curated relational database that provides detailed library information including tissue source, library construction methods, sequence diversity and sequence abundance. The differential expression of each EST species can be viewed across all libraries using a Virtual Expression Analysis Tool (VEAT), a graphical user interface written in Java for intra- and inter-library species comparisons. Recent enhancements to PEDB include: (i) the functional categorization of annotated EST assemblies using a classification scheme developed at The Institute for Genome Research; (ii) catalogs of expressed genes in specific prostate tissue sources designated as transcriptomes; and (iii) the addition of prostate proteome information derived from two-dimensional electrophoreses and mass spectrometry of prostate cancer cell lines. PEDB may be accessed via the WWW at http://www.mbt.washington.edu/PEDB/

Databases, Factual↗

PubMed2000

Comprehensive analyses of prostate gene expression: convergence of expressed sequence tag databases, transcript profiling and proteomics.

Several methods have been developed for the comprehensive analysis of gene expression in complex biological systems. Generally these procedures assess either a portion of the cellular transcriptome or a portion of the cellular proteome. Each approach has distinct conceptual and methodological advantages and disadvantages. We have investigated the application of both methods to characterize the gene expression pathway mediated by androgens and the androgen receptor in prostate cancer cells. This pathway is of critical importance for the development and progression of prostate cancer. Of clinical importance, modulation of androgens remains the mainstay of treatment for patients with advanced disease. To facilitate global gene expression studies we have first sought to define the prostate transcriptome by assembling and annotating prostate-derived expressed sequence tags (ESTs). A total of 55000 prostate ESTs were assembled into a set of 15953 clusters putatively representing 15953 distinct transcripts. These clusters were used to construct cDNA microarrays suitable for examining the androgen-response pathway at the level of transcription. The expression of 20 genes was found to be induced by androgens. This cohort included known androgen-regulated genes such as prostate-specific antigen (PSA) and several novel complementary DNAs (cDNAs). Protein expression profiles of androgen-stimulated prostate cancer cells were generated by two-dimensional electrophoresis (2-DE). Mass spectrometric analysis of androgen-regulated proteins in these cells identified the metastasis-suppressor gene NDKA/nm23, a finding that may explain a marked reduction in metastatic potential when these cells express a functional androgen receptor pathway.

DNA, Complementary↗

PubMed2000

Toll receptors: an expanding role in our understanding of human disease.

Toll receptor proteins in Drosophila are involved in establishing the dorsal-ventral axis in embryogenesis as well as participating in the innate immune response to invading pathogens. The basic mediators of this response show striking similarities in plants, insects, and vertebrates. The cytoplasmic signaling cascade is exemplified by the human interleukin-1 receptor complex (IL-1R), resulting in transcriptional activation of effector proteins through nuclear factor-kappaB (NF-kappaB). Six mammalian/human Toll-like receptors (TLR) have been described to date. The TLRs share the IL-1R cytoplasmic signaling cascade but are distinguished by their extracellular leucine-rich repeat (LRR) structure. The LRR superfamily comprises a diverse group of proteins, including a cohort involved in transmembrane signaling. Two of the human TLRs (TLR2, TLR4) have been shown to be involved in the innate response to bacterial pathogens and appear to provide a link between the innate and adaptive immune response. A better understanding of this response may provide improved therapeutic modalities in the treatment of bacterial and fungal sepsis, which continues to be a significant source of morbidity and mortality worldwide. In addition, similar to Drosophila, Toll receptors and related proteins in the LRR superfamily may also be involved in human development, as well as in noninfectious human disease.

Animals↗

PubMed1999

Negative selection: a method for obtaining low-abundance cDNAs using high-density cDNA clone arrays.

The identification of the entire complement of genes expressed in a cell, tissue, or organism provides a framework for understanding biological properties and establishes a tool set for subsequent functional studies. The large-scale sequencing of randomly selected clones from cDNA libraries has been successfully employed as a method for identifying a large fraction of these expressed genes. However, this approach is limited by the inherent redundancy of cellular transcripts reflecting widely variant levels of gene transcription. As a result, a high percentage of transcript duplications are encountered as the number of sequenced clones accrues. To address this problem, we have developed a negative hybridization selection method that employs the hybridization of complex cDNA probes to high-density arrays of cDNA clones and the subsequent selection of clones with a null or low hybridization signal. This approach was applied to a cDNA library constructed from normal human prostate tissue and resulted in the reduction of highly expressed prostate cDNAs from 6.8 to 0.57% with an overall decline in clone redundancy from 33 to 11%. The selected clones also reflected a more diverse cDNA population, with 89% of the clones representing distinctly different cDNAs compared with 67% of the randomly selected clones. This method compares favorably with cDNA library re-association normalization approaches and offers several distinct advantages, including the flexibility to use previously prepared libraries, and the ability to employ an iterative screening approach for continued accrual of cDNAs representing rare transcripts.

DNA, Complementary↗

PubMed1999

Comparative hybridization of an array of 21,500 ovarian cDNAs for the discovery of genes overexpressed in ovarian carcinomas.

Comparative hybridization of cDNA arrays is a powerful tool for the measurement of differences in gene expression between two or more tissues. We optimized this technique and employed it to discover genes with potential for the diagnosis of ovarian cancer. This cancer is rarely identified in time for a good prognosis after diagnosis. An array of 21,500 unknown ovarian cDNAs was hybridized with labeled first-strand cDNA from 10 ovarian tumors and six normal tissues. One hundred and thirty-four clones are overexpressed in at least five of the 10 tumors. These cDNAs were sequenced and compared to public sequence databases. One of these, the gene HE4, was found to be expressed primarily in some ovarian cancers, and is thus a potential marker of ovarian carcinoma.

Biomarkers, Tumor↗

PubMed1999

Prostate-localized and androgen-regulated expression of the membrane-bound serine protease TMPRSS2.

Genes regulated by androgenic hormones are of critical importance for the normal physiological function of the human prostate gland, and they contribute to the development and progression of prostate carcinoma. We used cDNA microarrays containing 1500 cDNAs to profile transcripts regulated by androgens in prostate cancer cells and identified the serine protease TMPRSS2 as a gene exhibiting increased expression upon exposure to androgens. The TMPRSS2 gene is located on chromosome 21 and contains four distinct domains, including a transmembrane region, indicating that it is expressed on the cell surface. Northern analysis demonstrated that TMPRSS2 is highly expressed in prostate epithelium relative to other normal human tissues. In situ hybridization of normal and malignant prostate tissues localizes TMMPRSS2 expression to prostate basal cells and to prostate carcinoma. These results suggest that TMPRSS2 may play a role in prostate carcinogenesis and should be investigated as a diagnostic or therapeutic target for the management of prostate cancers.

Androgens↗

PubMed1999

Monitoring gene expression profile changes in ovarian carcinomas using cDNA microarray.

The development of cancer is the result of a series of molecular changes occurring in the cell. These events lead to changes in the expression level of numerous genes that result in different phenotypic characteristics of tumors. In this report we describe the assembly and utilization of a 5766 member cDNA microarray to study the differences in gene expression between normal and neoplastic human ovarian tissues. Several genes that may have biological relevance in the process of ovarian carcinogenesis have been identified through this approach. Analyzing the results of microarray hybridizations may provides new leads for tumor diagnosis and intervention.

Cloning, Molecular↗

PubMed1999

Molecular cloning and characterization of prostase, an androgen-regulated serine protease with prostate-restricted expression.

The identification of genes with selective expression in specific organs or cell types provides an entry point for understanding biological processes that occur uniquely within a particular tissue. Using a subtraction approach designed to identify genes preferentially expressed in specific tissues, we have identified prostase, a human serine protease with prostate-restricted expression. The prostase cDNA encodes a putative 254-aa polypeptide with a conserved serine protease catalytic triad and an amino-terminal pre-propeptide sequence, indicating a potential secretory function. The genomic sequence comprises five exons and four introns and contains multiple copies of a chromosome 19q-specific minisatellite repeat. Northern analysis indicates that prostase mRNA is expressed in hormonally responsive normal and neoplastic prostate epithelial tissues, but not in prostate stromal constituents. Prostase shares 35% amino acid identity with prostate-specific antigen (PSA) and 78% identity with the porcine enamel matrix serine proteinase 1, an enzyme involved in enamel matrix degradation and with a putative role in the disruption of intercellular junctions. Radiation-hybrid-panel mapping localized prostase to chromosome 19q13, a region containing several other serine proteases, including protease M, pancreatic/renal kallikrein hK1, and the prostate-specific kallikreins hK2 and hK3 (PSA). The sequence homology between prostase and other well-characterized serine proteases suggests several potential functional roles for the prostase protein that include the degradation of extracellular matrix and the activation of PSA and other proteases.

Amino Acid Sequence↗

PubMed1999

PEDB: the Prostate Expression Database.

The Prostate Expression Database (PEDB) is a curated relational database and suite of analysis tools designed for the study of prostate gene expression in normal and disease states. Expressed Sequence Tags (ESTs) and full-length cDNA sequences derived from more than 40 human prostate cDNA libraries are maintained and represent a wide spectrum of normal and pathological conditions. Detailed library information including tissue source, library construction methods, sequence diversity and abundance are available in a library archive. Prostate ESTs are assembled into distinct species groups using the multiple alignment program CAP2 and are annotated with information from the GenBank, dbEST and Unigene public sequence databases. Annotated sequences in PEDB are searched using the BLAST algorithm. The differential expression of each EST species can be viewed across all libraries using a Virtual Expression Analysis Tool (VEAT), a graphical user interface written in Java for intra- and inter-library species comparisons. PEDB may be accessed via the World Wide Web at http://www.mbt.washington.edu/PEDB/

Base Sequence↗

PubMed1998

Cloning and characterization of two Toll/Interleukin-1 receptor-like genes TIL3 and TIL4: evidence for a multi-gene receptor family in humans.

Remarkable structural and functional similarities exist between the Drosophila Toll/Cactus/Dorsal signaling pathway and the mammalian cytokine-mediated interleukin-1 receptor (IL-1R)/I-kappaB/NF-kappaB activation cascade. In addition to a role regulating dorsal-ventral polarity in the developing Drosophila embryo, signaling through Drosophila Toll (dToll) activates the nonclonal, or innate, immune response in the adult fly. Recent evidence indicates that a human homologue of the dToll protein participates in the regulation of both innate and adaptive human immunity through the activation of NF-kappaB and the expression of the NF-kappaB-controlled genes IL-1, IL-6, and IL-8, thus affirming the evolutionary conservation of this host defense pathway. We report here the cloning of two novel human genes, TIL3 and TIL4 (Toll/IL-1R-like-3, -4) that exhibit homology to both the leucine-rich repeat extracellular domains and the IL-1R-like intracellular domains of human and Drosophila Toll. Northern analysis showed distinctly different tissue distribution patterns with TIL3 expressed predominantly in ovary, peripheral blood leukocytes, and prostate, and TIL4 expressed primarily in peripheral blood leukocytes and spleen. Chromosomal mapping by fluorescence in situ hybridization localized the TIL3 gene to chromosome 1q41-42 and TIL4 to chromosome 4q31.3-32. Functional studies showed that both TIL3 and TIL4 are able to activate NF-kappaB, though in a cell type-dependent fashion. Together with human Toll, TIL3 and TIL4 encode a family of genes with conserved structural and functional features involved in immune modulation.

Amino Acid Sequence↗

PubMed1998

Hevin, an antiadhesive extracellular matrix protein, is down-regulated in metastatic prostate adenocarcinoma.

Hevin, a gene closely related to the extracellular matrix protein SPARC, is an acidic cysteine-rich glycoprotein shown to be important for the adhesion and trafficking of cells through the endothelium. Through the use of differential display and differential EST analysis, we identified Hevin as a gene whose transcription is down-regulated in transformed prostate epithelial cell lines and metastatic prostate adenocarcinoma. These results were confirmed by comparing expression levels between normal and neoplastic human prostate tissues using Northern analysis. In situ hybridization with an 35S-labeled antisense riboprobe demonstrated the loss of Hevin expression in metastatic prostate carcinoma. The expression pattern of Hevin in transformed and metastatic epithelium may provide further insights into the complex cell adhesion events involved in the metastatic progression of prostate carcinoma.

Adenocarcinoma↗

PubMed1998

An expressed-sequence-tag database of the human prostate: sequence analysis of 1168 cDNA clones.

The human prostate is a complex glandular organ with functional development under hormonal regulation. Diseases of the prostate result in significant morbidity and mortality in the form of benign prostatic hypertrophy and prostate adenocarcinoma. The characterization of the molecular framework of the human prostate at the level of expressed genes will facilitate the understanding of normal and pathological prostate biology. The purposes of this study were to acquire an initial assessment of the qualitative and quantitative diversity of gene expression in the normal human prostate and to determine the extent that genes with prostate-restricted expression can be assessed using an expressed sequence tag approach. We have constructed a directional cDNA library from normal adult human prostate tissue and partially sequenced the 5' end of 1168 randomly selected cDNA clones, resulting in more than 400 kb of DNA sequence. Homology searches of the sequenced cDNAs against the GenBank and dbEST databases revealed that 43% of the sequences are identical to human genes whose functions are known, 5% are similar but not identical to known genes in humans or lower organisms, 5% match the mitochondrial genome, 9% are composed of interspersed DNA repeats, 30% are homologous to sequences in the dbEST database without a described function, and 6% are novel sequences. A total of 780 distinct species were identified. In addition to the 74 novel transcripts, 4 genes, prostate-specific antigen (PSA), prostate secretory protein (PSP), prostate acid phosphatase (PAP), and human glandular kallekrein 2 (HK2), have no homologous sequences in the databases that originate from sources other than prostate and thus may represent genes with prostate-restricted expression. Sequences matching PSA, PSP, and PAP each accounted for > 1% of the total ESTs and represent highly abundant transcripts, correlating with the abundance of these proteins in the prostate gland. No novel transcripts were represented by more than one EST and thus are expressed at levels much lower than the known prostate-specific genes.

Adult↗

PubMed1997

Cell-cell interaction in prostate gene regulation and cytodifferentiation.

To examine the role of intercellular interaction on cell differentiation and gene expression in human prostate, we separated the two major epithelial cell populations and studied them in isolation and in combination with stromal cells. The epithelial cells were separated by flow cytometry using antibodies against differentially expressed cell-surface markers CD44 and CD57. Basal epithelial cells express CD44, and luminal epithelial cells express CD57. The CD57+ luminal cells are the terminally differentiated secretory cells of the gland that synthesize prostate-specific antigen (PSA). Expression of PSA is regulated by androgen, and PSA mRNA is one of the abundant messages in these cells. We show that PSA expression by the CD57+ cells is abolished after prostate tissue is dispersed by collagenase into single cells. Expression is restored when CD57+ cells are reconstituted with stromal cells. The CD44+ basal cells possess characteristics of stem cells and are the candidate progenitors of luminal cells. Differentiation, as reflected by PSA production, can be detected when CD44+ cells are cocultured with stromal cells. Our studies show that cell-cell interaction plays an important role in prostatic cytodifferentiation and the maintenance of the differentiated state.

Androgens↗

PubMed1997

Identification of differentially expressed prostate genes: increased expression of transcription factor ETS-2 in prostate cancer.

BACKGROUND: Little is known about the genetic events in the malignant transformation of prostatic cells. This is due in large measure to the cellular heterogeneity of the prostate. METHODS: An amplification method was devised to synthesize cDNA from small samples of cancer and benign tissues of the same resected glands. Differential gene expression of candidate informative markers between cancer and benign was screened by the polymerase chain reaction with gene-specific oligonucleotide primers. RESULTS: The expression of a transcription factor, ETS-2, was shown to be elevated in some cancer specimens. Elevated expression was also noted for neuron-specific enolase (NSE) and another transcription factor, SEF2. CONCLUSIONS: Our method can be used to identify quickly genes that are differentially expressed between benign and cancerous prostate cells. Transcription factors, such as ETS-2, may play a significant role in cancer progression.

Cell Transformation, Neoplastic↗

PubMed1997

Rainbow Universal CPG: a versatile solid support for oligonucleotide synthesis.

We have developed a universal solid support, termed Rainbow Universal CPG, for use in automated oligonucleotide synthesis. The universal solid support allows any oligodeoxyribonucleotide sequence to be synthesized from a single type of controlled pore glass (CPG) support. Deprotection of oligodeoxyribonucleotides was optimized using 0.5 M LiCl in concentrated ammonium hydroxide. PCR experiments using three different sets of primers proved that the 3'-hydroxyl function of oligodeoxyribonucleotides synthesized from Rainbow Universal CPG was retained. This universal solid support shows promise for replacing the standard nucleoside CPG supports.

Automation↗

PubMed1997

Inexpensive handheld device for the construction of high-density nucleic acid arrays.

We report an easy-to-use, 384-pin handheld arraying and replicating device (ARD) for constructing high-density replicas of nucleic acids and E. coli transformants. We have modified an existing 384-pin tool to include a novel guide system to ensure vertical pin movement and enhance reproducibility. An asymmetric rectangular multiplexing frame is designed to increase the array density to 1536 dots on a standard microplate-size nylon membrane and to reduce the time and effort involved in producing array replicas. Our initial studies used the ARD to construct 1536-dot arrays of ovarian cDNA clones. We have hybridized these arrays with 32P-labeled probes, which resulted in distinctive signals for either visual interpretation or semiautomatic spot detection and signal integration.

Autoradiography↗

PubMed1996

Chemoprevention for prostatic intraepithelial neoplasia.

OBJECTIVES: To evaluate the potential application of chemoprevention strategies in prostatic intraepithelial neoplasia. METHODS: Review of relevant literature on chemoprevention with emphasis on prostate cancer and premalignant lesions. RESULTS: Chemoprevention represents a strategy designed to inhibit or reverse the process of carcinogenesis by administering one or several noncytotoxic chemical compounds. The epidemiology of prostate carcinoma indicates that this cancer is a prime candidate for a strategy aimed at prevention due to the extremely high prevalence rate, rising annual incidence, and long latent interval between the cancer-initiating events and the development of invasive disease. Chemopreventive agents may exert their inhibitory effects at different stages of the multistep carcinogenic process broadly categorized as initiation, promotion, and progression. The synthetic retinoids, polyamine synthesis inhibitors, and antiandrogens are among the compounds shown to have in vitro or in vivo chemopreventive effects in prostate carcinogenesis. A major limitation in the evaluation of such agents in a human prostate cancer is the long duration of clinical trials required to assess the efficacy with an endpoint of cancer development. Premalignant epithelial changes such as prostate intraepithelial neoplasia, or PIN, are highly associated with prostate cancer, and share many molecular features of invasive cancer. If the reversal or inhibition of intraepithelial neoplasia translates to a concomitant reduction in clinically relevant prostate cancer, then the pharmacological modulation of PIN may provide a rapid means to evaluate the effects and benefits of potential chemopreventive agents. CONCLUSION: Men with PIN may represent ideal candidates for chemoprevention protocols. The ideal agent and duration of therapy remains to be defined.

Androgen Antagonists↗