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At least 685 records · Page 38Linked to original sources
PubMed2002

Single cell expression analysis--pharmacogenomic potential.

A fundamental challenge in biology is to correlate physiology with gene expression in specific cell types. This can only be achieved by understanding gene expression at the level of the single cell because, in many systems, each cell has the capacity to express a unique set of genes. Therefore, each cell can be considered to be functionally distinct. A clearer understanding of gene expression differences at such a discrete level provides an opportunity to develop drugs with more targeted pharmacologies or with decreased side effects.

Animals↗
PubMed2004

[Expression analysis of mouse homologous proteins with human aldose reductase like-1].

OBJECTIVE: To detect expression of mouse ARL-1 homologous proteins in mouse tissues, and analyze homology, genetic distance and phylogenetic relationship between human aldose reductase like-1 (ARL-1) and mouse homologous proteins. METHODS: Homology of mouse ARL-1 homologous proteins with human ARL-1 was analyzed by software Clustal X 1.8 using GenBank and Swiss-Prot database; genetic distance and phylogenetic relationship between mouse ARL-1 homologous proteins and human ARL-1 were analyzed by software Mega 2.0; mouse tissues were detected by Western blotting using polyclonal antibodies against ARL-1 protein from domestic rabbits. RESULTS: The amino acid sequence of human ARL-1 was 83%, 82%, 81%, 79%, 70%, 51%, 50% and 45% identical to that of the Chinese hamster ovary reductase (CHO-Red), the mouse fibroblast growth factor-regulated protein (FR-1), rat aldose reductase-like protein (rARLP), the mouse vas deferens protein (MVDP), rat lens aldose reductase (LeAR), delta4-3-ketosteroid-5beta-reductase (5beta-Red), rat aldo-keto reductase protein c (RaK-c) and 3alpha-hydroxysteroid dehydrogenase (3alpha-HSD). Among all the mouse ARL-1 homologous proteins, the genetic distance between CHO-Red and human ARL was the shortest (18.0%, P = 0.023), next was FR-1 (19.1%, P=0.023) and rARLP (19.9%, P = 0.025). From the phylogenetic tree, the protein whose relationship with human ARL-1 was the closest with CHO-Red, next was mouse FR-1, rARLP, MVDP and LeAR. Homologous proteins were found in mouse tissues including vas deferens, testis, bladder and uterus by Western blotting using polyclonal antibodies against ARL-1 protein from domestic rabbits. CONCLUSIONS: CHO-Red has the highest homology, the shortest genetic distance and the closest relationship with human ARL-1, next is FR-1, rARLP, MVDP. The major distribution of mouse ARL-1 homologous proteins is in vas deferens, testis, bladder and uterus, deducing they might be CHO-Red, FR-1, rARLP or MVDP

ADP-Ribosylation Factors↗
PubMed2005

Genome-wide expression analysis of yeast response during exposure to 4 degrees C.

Adaptation to temperature fluctuation is essential for the survival of all living organisms. Although extensive research has been done on heat and cold shock responses, there have been no reports on global responses to cold shock below 10 degrees C or near-freezing. We examined the genome-wide expression in Saccharomyces cerevisiae, following exposure to 4 degrees C. Hierarchical cluster analysis showed that the gene expression profile following 4 degrees C exposure from 6 to 48 h was different from that at continuous 4 degrees C culture. Under 4 degrees C exposure, the genes involved in trehalose and glycogen synthesis were induced, suggesting that biosynthesis and accumulation of those reserve carbohydrates might be necessary for cold tolerance and energy preservation. The observed increased expression of phospholipids, mannoproteins, and cold shock proteins (e.g., TIP1) is consistent with membrane maintenance and increased permeability of the cell wall at 4 degrees C. The induction of heat shock proteins and glutathione at 4 degrees C may be required for revitalization of enzyme activity, and for detoxification of active oxygen species, respectively. The genes with these functions may provide the ability of cold tolerance and adaptation to yeast cells.

Adaptation, Physiological↗
PubMed2003

[Cloning and tissue expression analysis of up-regulated cDNA fragment in human gastric cancer].

OBJECTIVE: To identify novel human gastric cancer-associated susceptibility gene for early diagnosis and treatment of gastric cancer. METHODS: A primer was designed for 3'-rapid amplification of cDNA end(RACE) and amplified fragments were cloned, then they were analyzed by sequencing. Compared with ESTs in Genbank, the EST fragment represented a novel gene. Combination of Northern blot and virtual Northern and multiple tissues Northern blot, expression of the cDNA in multiple normal and carcinoma tissues were analyzed. RESULTS: One of the important cDNA bands with poly(A) tail was cloned. This band was named W41. Sequence analysis showed that W41 consists of 533 bp. Basic local alignment search tool analysis revealed that W41 has low identity with any genes from GenBank. This sequence data was submitted to GenBank with accession No. AF 325202. Northern blot revealed that W41 presented higher expression in gastric cancer tissue than in normal tissue. Multiple tissue Northern blot revealed that W41 presented higher expression in multiple cancers than in normal tissues. Virtual Northern revealed that the cDNA presented higher expression in tumor series analysis of gene expression libraries than in normal. CONCLUSION: A novel human gastric cancer-associated cDNA fragment was identified.

Blotting, Northern↗
PubMed1993

Plant dnaj homologue: molecular cloning, bacterial expression, and expression analysis in tissues of cucumber seedlings.

Proteins homologous to the bacterial dnaJ protein have been implicated as molecular chaperones in different compartments of eukaryotes. A plant equivalent is now described in various cucumber tissues. Using a cDNA library constructed from poly(A)-rich RNA of cotyledons of etiolated seedlings and dnaJ-specific probes, we isolated clones encoding full-length cDNAs: clone CSDNAJ-1 contained a 1.7-kb insert with a single open reading frame of 413 amino acid residues encoding a protein of M(r) 46, 012; pCSDNAJ-2 differed from pCSDNAJ-1 by an additional nucleotide and predicted a protein of M(r) 26, 104. Peptides of similar sizes were observed when the cDNA information was expressed by in vitro transcription and translation or expressed in vivo in bacteria. Using mRNA isolated by hybrid selection for in vitro translation we observed the formation of CSDNAJ-1 protein. Comparisons revealed that the similarity in primary structure between CSDNAJ-1 protein and dnaJ homologues in eukaryotes is most pronounced in the N-terminal region. The high degree of identity (39%) of CSDNAJ-1 protein with the yeast dnaJ homologue YDJ1 protein suggests that csdnaJ proteins assist intracellular protein transfer. The predicted CSDNAJ-1 protein is characterized by a fourfold repetition of the motif CXXCXGX(G) and by a C-terminus -QCAQQ. Analysis of gene expression at the level of mRNA showed that the putative dnaJ protein is expressed in cucumber seedlings in all tissues, but exceedingly high in hypocotyledons and roots. The level of dnaJ mRNA was transiently increased by a factor of 1.5-2.0 when heat shock was applied to the seedlings.

Amino Acid Sequence↗
PubMed2005

Cloning and expression analysis of retinoic acid receptors in the zebra finch brain.

The vitamin A derivative retinoic acid is produced postembryonically in discrete portions of the songbird brain, including some of the nuclei involved in song production and song learning, and its synthesis is required for the normal maturation of song behavior. To identify the brain targets for retinoic acid action, we cloned the zebra finch homologs of the alpha, beta, and gamma classes of retinoic acid receptors (RARs). In situ hybridization analysis revealed that the mRNAs for all three RARs are expressed at different levels in several brain areas, with a broader distribution than the mRNA for retinaldehyde-specific aldehyde dehydrogenase (zRalDH), a retinoic acid-synthesizing enzyme. Detectable RAR expression was found in all nuclei of the song control system, with the most marked expression occurring within the striatal song nucleus area X. These observations are consistent with a persistent action of retinoic acid in the postembryonic and adult songbird brain and provide further evidence for an involvement of retinoic acid signaling in the control of learned vocal behavior in a songbird species. They also suggest that the striatum is a major target of retinoic acid in songbirds.

Amino Acid Sequence↗
PubMed1998

Cloning and expression analysis of a novel salicylate suppressible gene, Hs-CUL-3, a member of cullin/Cdc53 family.

By using a mRNA differential display technique to search for salicylate suppressible genes, we identified a cDNA in human foreskin fibroblasts, which by GenBankTM DNA data base search shows sequence homology to the recently reported cullin/Cdc53 (CUL) family genes, especially CUL-3. We have cloned the full-length human CUL-3 (Hs-CUL-3) cDNA. It encodes a 768-amino acid polypeptide and has a predicted molecular weight of 88,939. The amino acid sequence of Hs-CUL-3 shows 46% homology to that of its Caenorhabditis elegans ortholog, Ce-CUL-3, and 27 and 23% to that of Hs-CUL-1 and Hs-CUL-2, respectively. Northern blot analysis showed that phorbol 12-myristate 13-acetate increased the expression of Hs-CUL-3 mRNA in a concentration- and time-dependent manner, and this increase was inhibited by sodium salicylate. Hs-CUL-3 widely expressed in human tissues and its expression in cultured COLO205 colon cancer cells was increased when compared with that in normal colon cells. It is likely that Hs-CUL-3 is involved in cell proliferation control.

Amino Acid Sequence↗
PubMed2003

Gene expression analysis of human middle ear cholesteatoma using complementary DNA arrays.

OBJECTIVE: To identify genes regulated in human cholesteatoma compared with normal skin tissue using complementary DNA arrays. STUDY DESIGN: In vitro analysis. METHODS: Eight cholesteatoma and retroauricular skin samples were obtained from the same patients during surgery. Upregulated and downregulated genes were highlighted using complementary DNA arrays for screening. Reverse transcriptase-polymerase chain reaction and immunohistochemical staining were performed to confirm the results of the complementary DNA array. RESULTS: Twelve genes were found to be induced or upregulated in cholesteatoma compared with skin samples. These included genes involved in cell proliferation and differentiation (eg, calgranulin A, calgranulin B, psoriasin, thymosin beta-10) and cell invasion (eg, cathepsin C, cathepsin D, cathepsin H). Analyses by means of reverse transcription-polymerase chain reaction showed enhanced expression of several genes including calgranulin A, calgranulin B, psoriasin, thymosin beta-10, cathepsin C, cathepsin D, and cathepsin H in cholesteatoma, supporting the findings from the gene array. In addition, it was verified by immunohistochemical analysis that the expressions of Calgranulin A, Calgranulin B, and Cathepsin D were mainly located in cholesteatoma epithelium. CONCLUSION: The observed alteration in gene expression may play a role in various mechanisms of pathogenesis in cholesteatoma.

Actins↗
PubMed2004

Single-cell gene expression analysis: implications for neurodegenerative and neuropsychiatric disorders.

Technical and experimental advances in microaspiration techniques, RNA amplification, quantitative real-time polymerase chain reaction (qPCR), and cDNA microarray analysis have led to an increase in the number of studies of single-cell gene expression. In particular, the central nervous system (CNS) is an ideal structure to apply single-cell gene expression paradigms. Unlike an organ that is composed of one principal cell type, the brain contains a constellation of neuronal and noneuronal populations of cells. A goal is to sample gene expression from similar cell types within a defined region without potential contamination by expression profiles of adjacent neuronal subpopulations and noneuronal cells. The unprecedented resolution afforded by single-cell RNA analysis in combination with cDNA microarrays and qPCR-based analyses allows for relative gene expression level comparisons across cell types under different experimental conditions and disease states. The ability to analyze single cells is an important distinction from global and regional assessments of mRNA expression and can be applied to optimally prepared tissues from animal models as well as postmortem human brain tissues. This focused review illustrates the potential power of single-cell gene expression studies within the CNS in relation to neurodegenerative and neuropsychiatric disorders such as Alzheimer's disease (AD) and schizophrenia, respectively.

Cell Physiological Phenomena↗
PubMed2006

Genome-wide gene expression analysis of Pseudomonas syringae pv. tomato DC3000 reveals overlapping and distinct pathways regulated by hrpL and hrpRS.

Pseudomonas syringae pv. tomato DC3000 is a model pathogen infecting tomato and Arabidopsis plants. Genes encoding the type III secretion system and substrate proteins (collectively called TTSS genes) of this bacterium are induced in plants and in minimal medium (MM). The induction of TTSS genes is mediated by HrpL, an alternative sigma factor recognizing the hrp box in the promoter of TTSS genes. The transcription of hrpL is activated by HrpR and HrpS, two homologous DNA-binding proteins encoded by the hrpRS operon. Microarray analysis was conducted to evaluate the DC3000 genes regulated by hrpL and hrpRS in MM. The analysis identified a number of novel hrpL-activated genes with a putative TTSS-independent function. Genes regulated by hrpL were mostly regulated by hrpRS in the same manner, but a large number of genes regulated by hrpRS were hrpL-independent, indicating that hrpL represents one branch of the regulatory pathways downstream of hrpRS. The induction of the TTSS genes was associated with downregulation of the housekeeping genes, indicating that the activation of the TTSS has a cost on the basic cellular activities. The novel genes and pathways identified by the microarray provide new insight into the bacterial functions coordinating with the TTSS.

Bacterial Proteins↗
PubMed2003

Gene expression analysis of purified hematopoietic stem cells and committed progenitors.

Lifelong self-renewal is a unique property of somatic stem cells. Recently, several primitive multipotent yet committed (non-self-renewing) hematopoietic progenitor populations were identified in mouse bone marrow. We have characterized the expression of 1200 selected mouse genes using the Atlas cDNA array in highly purified hematopoietic stem cells (HSCs) and 6 closely related progenitor populations: common myeloid progenitors (CMPs), granulocyte-macrophage progenitors (GMPs), megakaryocyte-erythrocyte progenitors (MEPs), common lymphoid progenitors (CLPs), and pro-T and pro-B cells. Cluster analysis revealed that nearly half of all differentially expressed transcripts are associated with HSCs, supporting the notion of an active transcriptional status of HSCs. Genes found enriched in the HSC cluster encompass many developmentally regulated genes, some previously associated with HSC self-renewal. In contrast, genes that are enriched in committed progenitors are mostly associated with hematopoietic differentiation, immune regulation, and metabolism. Thus, the transition from HSCs toward committed progenitors correlates with the down-regulation of a large number of HSC-associated genes and progressive up-regulation of a limited number of lineage-specific genes. These genetic analyses revealed both quantitative and qualitative differences between the transcripts associated with HSCs versus downstream progenitors and produced a list of the candidate genes, potentially involved in HSC self-renewal.

Animals↗
PubMed2002

Characterization of the porcine sperm adhesion molecule gene SPAM1- expression analysis, genomic structure, and chromosomal mapping.

Sequence analysis of cDNA products, derived from adult porcine testis mRNA, gave overlapping nucleotide sequence correlating to 1952 bp of the sperm adhesion molecule 1 (SPAM1) gene. This sequence was shown to be homologous to SPAM1 genes known in other mammalian species and contained an open reading frame encoding a 493-amino acid protein. Fluorescence in situ hybridization (FISH), using a bacterial artificial chromosome (BAC) clone from the PigE BAC library, was used to map SPAM1 to chromosome 18 of the pig. This finding is consistent with comparative mapping experiments performed between pig and human chromosomes. Polymerase chain reaction (PCR) analysis of genomic DNA has shown that the 1952 bp of cDNA sequence spans approximately 9 kb of genomic DNA and comprises of at least four exons, with its size and structure being relatively conserved between mouse, human and pig. Reverse transcriptase (RT)-PCR analysis of mRNA from nine porcine tissues has also suggested that expression of SPAM1 is limited to the testis.

Animals↗
PubMed2006

Identification and expression analysis of cDNAs encoding channel catfish type I interferons.

Previously a cDNA encoding a putative interferon gene, designated CF IFN-1, was identified from a catfish EST library. However, its constitutive expression, absence of a signal peptide, and apparently low level of biological activity suggested that this cDNA likely encoded an expressed pseudogene. Since Southern blot analysis suggested the presence of two to three IFN genes, additional cDNAs were generated from catfish fibroblast and lymphoid cell lines using primers designed to conserved regions of zebrafish and catfish interferon. Using this approach, three novel CF IFN genes, two of which likely encode functional interferon molecules, were identified. At the amino acid level, similarity among CF IFNs ranged from 71% to 82%, whereas similarity to other fish IFNs ranged from 15% to 35%. Although CF IFN-3, like CF IFN-1, lacks a signal peptide, CF IFN-2 and -4 appear to encode full-length, signal sequence-bearing genes. Consistent with their putative identification as functional genes, CF IFN-2 and -4 were not expressed in unstimulated cell lines, and CF IFN-2 was rapidly upregulated in CCO cells in response to virus infection or treatment with dsRNA. Moreover, as with salmon, fugu, and zebrafish interferon genes, CF IFN-1 contained four introns whose locations were conserved not only with respect to other fish IFNs, but also with respect to mammalian IFN-lambda. While it is likely that CF IFNs represent Type I IFNs, several characteristics preclude assigning these cytokines to any particular subfamily.

Amino Acid Sequence↗
PubMed2006

Protein identification and expression analysis using mass spectrometry.

The identification and quantification of the proteins that a whole organism expresses under certain conditions is a main focus of high-throughput proteomics. Advanced proteomics approaches generate new biologically relevant data and potent hypotheses. A practical report of what proteome studies can and cannot accomplish in common laboratory settings is presented here. The review discusses the most popular tandem mass-spectrometry-based methods and focuses on how to produce reliable results. A step-by-step description of proteome experiments is given, including sample preparation, digestion, labeling, liquid chromatography, data processing, database searching and statistical analysis. The difficulties and bottlenecks of proteome analysis are addressed and the requirements for further improvements are discussed. Several diverse high-throughput proteomics-based studies of microorganisms are described.

Amino Acid Sequence↗
PubMed1992

S100 alpha, CAPL, and CACY: molecular cloning and expression analysis of three calcium-binding proteins from human heart.

Elevated levels of intracellular calcium are a major cause of myocardial dysfunction. To find possible mediators of the deregulated calcium we searched for EF-hand calcium-binding proteins of the S100 family. By PCR technology we identified three members of the S100 protein family (S100 alpha, CACY, and CAPL) in the human heart. We cloned the corresponding cDNAs and examined their expression levels in various human tissues by Northern blot analysis. All three proteins are expressed at high levels in the human heart. Whereas CACY and CAPL mRNAs are expressed ubiquitously, S100 alpha mRNA is restricted to heart, skeletal muscle, and brain. Interestingly, the expression pattern of S100 alpha, CACY, and CAPL in human tissues differs significantly from that in rodent tissues.

Amino Acid Sequence↗
PubMed2003

Isolation and expression analysis of the canine insulin-like factor 3 gene.

The insulin-like factor 3 (INSL3 or relaxin-like factor) is a hormone produced mainly in gonadal tissues in males and females. Deletion of INSL3 or its receptor in male mice leads to the undescended testes, or cryptorchidism. Here we describe an isolation and analysis of full-length canine INSL3 gene. The INSL3 gene is composed of two exons within a small genomic region. Putative translation of the isolated cDNA yields 132 amino acid preproINSL3 that has the domain structure characteristic for the insulin-relaxin peptide superfamily with a well-conserved receptor-binding domain. Northern blot hybridization showed stronger expression of INSL3 in testis than in ovary. Reverse transcription-polymerase chain reaction analysis of the INSL3 expression revealed a minor splice variant of INSL3 potentially encoding 105 amino acids peptide. We established that the medium, conditioned with recombinant canine INSL3, produced from the full-length cDNA, but not from the minor splice variant, activated human GREAT/LGR8 receptor in vitro. In addition to the functional allele of INSL3, genomic DNA of one of the analyzed dogs contained an intronless nonexpressed pseudogene of INSL3. We isolated canine INSL3 promoter and showed that its activity was strongly mediated by steroidogenic factor-1 in vitro. Using site-specific mutagenesis, we identified a well-conserved steroidogenic factor-1 binding site within canine INSL3 promoter.

Animals↗