Organ-specific Gene Expression Research Articles

Organ-specific patterns of P450arom gene isoforms are modulated by p,p′-DDE in adult male European common frog, Rana temporaria

The organ-specific gene expression patterns of P450arom isoforms have been studied in European common frog, Rana temporaria after exposure to DDE, using real-time PCR. Four groups of frogs were subcutaneously injected with DDE at 0.01, 0.1, 1 and 10 mg/kg body weight and one group, serving as the control was injected with pure corn oil. Brain, kidney, testis and liver P450aromA and P450aromB gene expressions were evaluated at day 14 after exposure. P450aromB data show that 0.1 and 10 mg DDE/kg doses caused 76% and 63% (testis) and 79% and 80% reductions, respectively, of mRNA levels. Brain P450aromB mRNA decreased 10% and 34%, respectively, after exposure to 0.01 and 0.1 mg DDE/kg. Thereafter, a 185% and 177% induction response of brain P450aromB was observed in the groups treated with 1 and 10 mg DDE/kg, respectively. In the kidney, 0.01, 0.1, 1 and 10 mg DDE/kg induced a 516%, 178%, 466% and 247% induction of P450aronB mRNA, respectively. P450aromA expression was not quantified in any of the organs. The relative abundance of P450aromB gene expression in different organs is in the order: kidney > brain > liver > testis. The present data suggest potential detrimental effect of organochlorine pesticides (OCs) and their metabolites on the European frog steroidogenic pathways. Given the high persistency in the environment and continued use in developing countries coupled with the tendency for global atmospheric transport, OCs and their metabolites such as DDE will remain a focus of concern both for scientific and societal reasons.

Beyond CAR and PXR

It is becoming increasingly evident that constitutive, induced, and regulated expression of genes important to the drug disposition process such as drug transporters, phase I and II metabolic enzymes are largely under the transcriptional control of certain nuclear receptor (NR) family members. In the past decade, important new insights regarding the role and relevance of ligand-activated nuclear receptors such as such as the pregnane X receptor (PXR) and constitutive androstane receptor (CAR) in terms of their activation by endogenous biochemicals, natural products, as well as synthetic compounds have led to a much better understanding of the xenobiotic-mediated induction process and the clinical relevance of such NRs to drug therapy in general. However, in addition to CAR and PXR, many orphan and adopted orphan NRs have recently been identified as key regulators of drug disposition genes. Indeed, nuclear receptors including farnesoid X receptor, peroxisome proliferator-activated receptor, and hepatocyte nuclear factors (1alpha, 3 and 4alpha) exhibit overlapping ligand specificities and regulate multiple gene targets, resulting in tissue- and organ-specific expression of drug disposition genes. In this review, the biology, pathophysiology, and the potential clinical relevance of such NRs to drug disposition and response are discussed.

Differential organ expression patterns of thyroid hormone receptor isoform genes in p, p′-DDE-treated adult male common frog, Rana temporaria

Using the European common frog, Rana temporaria, as a model, we have studied the organ-specific gene expression patterns of thyroid hormone receptor isoforms after exposure to an organochlorine (OC) compound, p, p′-DDE . Four groups of frogs were subcutaneously injected with p, p′-DDE at 0.01, 0.1, 1 and 10 mg/kg body weight, respectively. In addition, one group, serving as the control group, was injected with pure corn oil. TH receptor isoforms (TRα and TRβ) gene expressions were evaluated in the brain, kidney, testis and liver using real-time PCR with gene-specific primers. Our results show that p, p′-DDE doses induced slight elevations of TRα and TRβ mRNA in the brain. In the testis, p, p′-DDE induced an initial significant 3-fold increase of TRα mRNA at 0.01 mg/kg and thereafter clear dose-dependent decreases of TRα mRNA levels were observed. For testicular TRβ mRNA levels, p, p′-DDE induced a slight elevation at 0.01 mg/kg and thereafter significant decreases in TRβ mRNA levels were observed. p, p′-DDE induced significant 2–4-fold elevations of both TR isoforms in frog kidney. The strongest transcriptional effect of p, p′-DDE on TR isoforms was observed in the kidney. While TRα mRNA was not measurable in the liver, p, p′-DDE induced an initial 1.7-fold increase at 0.01 mg/kg of TRβ mRNA and thereafter an apparent dose-dependent decrease was observed. The relative abundance of TRα and TRβ gene expression in different organs are in the order: kidney > testis > brain > liver. While the induction TRα and TRβ might result to hypersensitivity and subsequent gain of biological functions, the inhibition might result to loss of biological function. Given the high persistency in the environment and continued use in developing countries coupled with the tendency for global atmospheric transport, DDT and its metabolites such as p, p′-DDE will remain a focus of concern both for scientific and societal reasons.

Midkine regulates pleiotrophin organ-specific gene expression: Evidence for transcriptional regulation and functional redundancy within the pleiotrophin/midkine developmental gene family

Midkine (MK) and the highly related cytokine pleiotrophin (PTN) constitute the PTN/MK developmental gene family. The Mk and Ptn genes are essential for normal development of the catecholamine and renin–angiotensin pathways and the synthesis of different collagens. It is not known whether the Ptn and Mk genes regulate each other or whether PTN and MK are functionally redundant in development. We have now compared the levels of expression of Ptn and Mk in genetically deficient Mk −/− and Ptn −/− mice and found highly significant increases in Ptn gene expression in spinal cord, dorsal root ganglia, eye, heart, aorta, bladder, and urethra, but not in brain, bone marrow, testis, and lung of Mk −/− mice compared with wild type mice; a remarkable ∼230-fold increase in Ptn expression levels was found in heart of Mk −/− mice and highly significant but lesser increases were found in six other organs. Differences in levels of Mk gene expression in Ptn −/− mice could not be detected in any of the organs tested. The data demonstrate that MK regulates Ptn gene expression with a high degree of organ specificity, suggesting that Ptn gene expression follows Mk gene expression in development, that the increase in Ptn gene expression is compensatory for the absence of MK in Mk −/− mice, that PTN and MK share a high degree of functional redundancy, and that MK may be very important in the development of heart in mouse.

The auxin-responsive GH3 gene family in rice (Oryza sativa)

Auxin regulates plant growth and development by altering the expression of diverse genes. Among these, the genes of Aux/IAA, SAUR, and GH3 classes have been extensively studied in dicots, but little information is available on monocots. We have identified 12 members of GH3 gene family in rice using sequences of full-length cDNA clones available from KOME and analysis of the whole genome sequence of rice. The genomic organization as well as chromosomal location of all the OsGH3 genes is reported. The rice GH3 proteins can be classified in two groups (groups I and II) on the basis of their phylogenetic relationship with Arabidopsis GH3 proteins. Based upon the sequences available in the database, not a single group III GH3 protein could be identified in rice. An extensive survey of EST sequences of other monocots led to the conclusion that although GH3 gene family is highly conserved in both dicots and monocots but the group III is conspicuous by its absence in monocots. The in silico analysis has been complemented with experimental data to quantify transcript levels of all GH3 gene family members. Using real-time polymerase chain reaction, the organ-specific expression of individual OsGH3 genes in light- and dark-grown seedlings/plants has been examined. The transcript abundance of nearly all OsGH3 genes is enhanced on auxin treatment, with the effect more pronounced on OsGH3-1, -2, and -4. The functional validation of these genes in transgenics or analysis of gene-specific insertional mutants will help in elucidating their precise role in auxin signal transduction.

ASYMMETRIC LEAVES2-LIKE1gene, a member of the AS2/LOB family, controls proximal?distal patterning in Arabidopsis petals

The formation and the development of the floral organs require an intercalate expression of organ-specific genes. At the same time, meristem-specific genes are repressed to complete the differentiation of the organs in the floral whorls. In an Arabidopsis activation tagging population, a mutant affected in inflorescence architecture was identified. This gain-of-function mutant, designated downwards siliques1 (dsl1-D), has shorter internodes and the lateral organs such as flowers are bending downwards, similar to the loss-of-function brevipedicellus (bp) mutant. The affected gene in dsl1-D appeared to be ASYMMETRIC LEAVES2-LIKE1 (ASL1)/LATERAL ORGAN BOUNDARIES domain gene 36 (LBD36), which is a member of the ASYMMETRIC LEAVES2 (AS2)/LATERAL ORGAN BOUNDARIES (LOB) domain gene family. Analysis of the loss-of-function mutant asl1/lbd36 did not show morphological aberration. Double mutant analysis of asl1/lbd36 together with as2, the ASL1/LBD36 closest homologue, demonstrates that these two members of the AS2/LOB family act partially redundant to control cell fate determination in Arabidopsis petals. Moreover, molecular analysis revealed that overexpression of ASL1/LBD36 leads to repression of the homeobox gene BP, which supports the model that an antagonistic relationship between ASL/LBD and homeobox members is required for the differentiation of lateral organs.

The inositol oxygenase gene family of Arabidopsis is involved in the biosynthesis of nucleotide sugar precursors for cell-wall matrix polysaccharides

The nucleotide sugar UDP-glucuronic acid (UDP-GlcA) is the principal precursor for galacturonic acid, xylose, apiose and arabinose residues of the plant cell-wall polymers. UDP-GlcA can be synthesized by two different functional pathways in Arabidopsis involving either UDP-glucose dehydrogenase or inositol oxygenase as the initial enzyme reaction to channel carbohydrates into a pool of UDP sugars used for cell-wall biosynthesis. The genes for the enzyme myo-inositol oxygenase (MIOX) were analyzed in Arabidopsis. They represent a small gene family containing four members. The transcription of all those members indicates a transient and organ-specific gene expression pattern in growing plant tissues as analyzed by RT-PCR and in promoter::GUS reporter gene lines. Two isoforms (MIOX1, MIOX2) are expressed in almost all tissues of the plant, whereas the expression of MIOX4 and MIOX5 is largely restricted to flowers, particularly maturing pollen. T-DNA insertion lines in MIOX genes were isolated; however, single knock-outs show growth phenotypes similar to the wild type. The monosaccharide composition of the cell wall in these mutants is not significantly changed compared to wild type plants. However, the incorporation of 3H-inositol into wall polymers of seedlings is greatly impaired in the mutant lines (Delta)MIOX1 and (Delta)MIOX2, which are the only isoforms that are expressed in seedlings.

Genome-wide transcriptome mapping analysis identifies organ-specific gene expression patterns along human chromosomes

The Human Genome Project has revealed that there about 32,000 protein-encoding genes, which are distributed throughout the genome. It is unclear, however, whether genes are distributed on the chromosomes according to patterns linked to organ specificity. To explore the relationship between genes actively transcribed in normal tissues and their chromosomal locations, we analyzed serial analysis of gene expression libraries of normal human liver, brain, breast, and colon tissues. Transcriptome mapping analysis revealed that transcriptional activity in each tissue varied according to the chromosomal domains, and a weak positive correlation was observed between transcription density and gene density. We identified six liver-related and five colon-related chromosomal domains highly transcribed in each tissue, whereas no brain-related or breast-related chromosomal domains were identified. Representative genes located on these chromosomal domains were associated with the function of each organ and were highly conserved in both mouse and rat genomes. These data revealed that the transcriptional activities of normal human tissues are well orchestrated at chromosomal levels, suggesting that highly expressed genes may share physical proximity.

Hepatocyte-specific gene expression from integrated lentiviral vectors.

For many applications, efficient gene therapy will require long-term, organ-specific therapeutic gene expression. Lentiviral vectors based on HIV-1 are promising gene delivery vehicles due to their ability to integrate transgenes into non-dividing cells. Many experimental vectors express transgenes under the control of the cytomegalovirus (CMV) immediate-early gene promoter. Although this promoter directs strong gene expression in vitro, it may be shut off rapidly in vivo. This study explores the potential of HIV-1-based vectors to transduce hepatocytes and compares gene expression from different promoters in integrated vectors. HIV-1-based vector plasmids expressing the green fluorescent protein (GFP) under the control of the CMV promoter, the alpha-1 antitrypsin gene promoter or promoters derived from the hepatitis B virus (HBV) genome were used to compare expression in transfected and transduced cell lines. Hepatocyte cell lines differed strikingly in their transfectability. Transduction with replication-deficient HIV-1-based vector particles incorporating the different promoter elements was uniformly effective in hepatocyte and non-hepatocyte lines. However, in hepatocytes, only the CMV, alpha-1 antitrypsin and HBV core but not HBV surface promoters were able to produce GFP expression. Addition of the HBV enhancer 2 element improved the transducing ability of the HBV surface promoter and suppressed expression in non-hepatocytes increasing specificity for hepatocytes. Integrated lentiviral vectors can be used to direct transgene expression in liver cells both promiscuously and specifically. Promoters derived from the alpha-1 antitrypsin gene or HBV are alternatives to the CMV promoter. Inclusion of the HBV enhancer 2 permits strong liver-specific gene expression in vitro.

Distinctive features of plant organs characterized by global analysis of gene expression in Arabidopsis.

The distinctive features of plant organs are primarily determined by organ-specific gene expression. We analyzed the expression specificity of 8809 genes in 7 organs of Arabidopsis using a cDNA macroarray system. Using relative expression (RE) values between organs, many known and unknown genes specifically expressed in each organ were identified. We also analyzed the organ specificity of various gene groups using the GRE (group relative expression) value, the average of the REs of all genes in a group. Consequently, we found that many gene groups even ribosomal protein genes, have strong organ-specific expression. Clustering of the expression profiles revealed that the 8809 genes were classified into 9 major categories. Although 3451 genes were clustered into the largest category, which showed constitutive gene expression, 266 and 1005 genes were found to be root- and silique-specific genes, respectively. By this clustering, particular gene groups which showed multi-organ-specific expression profiles, such as bud-flower-specific, stem-silique-specific or bud-flower-root-specific profiles, could be effectively identified. From these results, major features of plant organs could be characterized by their distinct profiles of global gene expression. These data of organ-specific gene expression are available at our web site: Arabidopsis thaliana Tissue-Specific Expression Database, ATTED (http://www.atted.bio.titech.ac.jp/).

ArabidopsisMSI1 is required for epigenetic maintenance of reproductive development

WD40 repeat proteins similar to yeast MSI1 are conserved in animals and plants, in which they participate in complexes involved in chromatin metabolism. Although MSI1-like proteins are well characterised biochemically, their function in the development of multicellular eukaryotes is not well understood. We constructed Arabidopsis plants in which the AtMSI1 protein level was altered. Strong ectopic expression of AtMSI1 produced no visible altered phenotype, but reduction of AtMSI1 dramatically affected development. The primary shoot apical meristem was unable to develop organs after the transition to flowering. Flowers that developed on floral shoots from axillary meristems experienced a progressive loss of floral morphology, including a reduction in size of the petals and stamens and the development of carpel-like sepals. Ovule development was disrupted in all flowers, resulting in complete female sterility. Molecular analysis of the mutant plants revealed that AtMSI1 is required to maintain the correct temporal and organ-specific expression of homeotic genes, including AGAMOUS and APETALA2. In contrast, FAS1 and FAS2, which together with AtMSI1 form the chromatin assembly complex CAF-1, are not required for repression of these genes. Therefore, AtMSI1 has specific functions in addition to CAF-1-mediated chromatin assembly. Efficient formation of heterochromatin, but not methylation of centromeric DNA repeats, depends on AtMSI1 presence demonstrating a key role of AtMSI1 in maintenance of chromatin structure.

Bioluminescent detection of endotoxin effects on HIV-1 LTR-driven transcription in vivo.

We investigated the effects of Gram-negative bacterial lipopolysaccharide (LPS) on luciferase expression in transgenic reporter mice in which luciferase expression is driven by the nuclear factor kappaB (NF-kappaB)-dependent portion of the human immunodeficiency virus-1 (HIV-1) long terminal repeat (HIV-1 LTR). Using these mice, we dissected the sources of luciferase activity at the organ level by (a) assessing luciferase activity in organ homogenates, (b) bioluminescence imaging in vivo, and (c) bioluminescence imaging of individual organs ex vivo. Luciferin dosage was a critical determinant of the magnitude of photon emission from these reporter mice. Photon emission increased at doses from 0.5-6 mg of luciferin given by intraperitoneal (IP) injection. The differential between basal and LPS-induced bioluminescence was maximal at 3-6 mg of luciferin. Luciferase expression was highly inducible in lungs, liver, spleen, and kidneys after a single IP injection of LPS, as assessed by luciferase activity measurements in organ homogenates. Luciferase activity was also induced in the forebrain by treatment with IP LPS. In contrast, aerosolized LPS produced a response localized to the lungs as assessed by both bioluminescence and ex vivo luciferase assay measurements. These studies demonstrate the utility of luciferase reporter mice for determining organ-specific gene expression in response to local and systemic stimuli.

The identification of candidate genes for a reverse genetic analysis of development and function in the Arabidopsis gynoecium.

The screening for mutants and their subsequent molecular analysis has permitted the identification of a number of genes of Arabidopsis involved in the development and functions of the gynoecium. However, these processes remain far from completely understood. It is clear that in many cases, genetic redundancy and other factors can limit the efficiency of classical mutant screening. We have taken the alternative approach of a reverse genetic analysis of gene function in the Arabidopsis gynoecium. A high-throughput fluorescent differential display screen performed between two Arabidopsis floral homeotic mutants has permitted the identification of a number of genes that are specifically or preferentially expressed in the gynoecium. Here, we present the results of this screen and a detailed characterization of the expression profiles of the genes identified. Our expression analysis makes novel use of several Arabidopsis floral homeotic mutants to provide floral organ-specific gene expression profiles. The results of these studies permit the efficient targeting of effort into a functional analysis of gynoecium-expressed genes.

Органоспецифічна та світлозалежна експресія генів у трансгенних рослинах: одержання і клонування корене-, бульбо- та листспецифічних промоторів

Органоспецифічна та світлозалежна експресія генів у трансгенних рослинах: одержання і клонування корене-, бульбо- та листспецифічних промоторів

Far-upstream elements are dispensable for tissue-specific proenkephalin expression using a Cre-mediated knock-in strategy.

Several cis-regulatory DNA elements are present in the 5' upstream regulatory region of the enkephalin gene (ENK) promoter. To determine their role in conferring organ-specificity of ENK expression in mice and to circumvent the position effects from random gene insertion that are known to often frustrate such analysis in transgenic mice, we used a Cre-mediated gene knock-in strategy to target reporter constructs to a "safe haven" loxP-tagged locus in the hypoxanthine phosphoribosyltransferase (HPRT) gene. Here we report reliable and reproducible reporter gene expression under the control of the 5' upstream regulatory region of the mouse ENK gene in gene-modified mice using this Cre-mediated knock-in strategy. Comparison of two 5'ENK regulatory regions (one with and the other without known cis-regulatory DNA elements) in the resulting adult mice showed that conserved far-upstream cis-regulatory DNA elements are dispensable for correct organ-specific gene expression. Thus the proximal 1.4 kb of the murine ENK promoter region is sufficient for organ-specificity of ENK gene expression when targeted to a safe-haven genomic locus. These results suggest that conservation of the far-upstream DNA elements serves more subtle roles, such as the developmental or cell-specific expression of the ENK gene.

Promoter analysis in transient assays using a GUS reporter gene construct in creeping bentgrass ( Agrostis palustris)

Transient expression profiles for several chimeric beta-glucuronidase (GUS) gene constructs were determined in tissues (young leaves, mature leaves and roots) of creeping bentgrass (Agrostis palustris, cv. Penn A4) following microprojectile bombardment. The constructs analyzed consisted of the uidA (GUS) reporter gene driven by four different promoters (ubiquitin 3-potato, ubiquitin corn, ubiquitin rice and CaMV 35S). The total number of GUS hits (or transient expression units; TEUs) were determined manually under a dissecting scope after histochemical staining for GUS. Results suggest that the ubiquitin rice promoter is most active in cells of turfgrass, regardless of the developmental stage or tissue-type. The ubiquitin corn promoter was the next best. Of the four promoter used, except for ubiquitin 3-potato, reporter gene activity was dramatically higher in mature leaves compared to young leaves. The relative efficiency of each promoter was about the same in roots and leaves. We have also analyzed uidA (GUS) reporter gene activity following microprojectile bombardment in transient expression assays with callus from two cultivars (Providence or Penn A4) of creeping bentgrass. Differences in the frequency of GUS positive hits were observed between cultivars up to 72 hours post-bombardment. However, this difference between cultivars disappeared after 72 hours post-bombardment. This information describing promoter functionality in bentgrass will be important when designing gene constructs for trait modification and when choosing appropriate cultivars for improvement through gene transfer experiments. This is the first in depth report on organ-specific and developmental gene expression profiles for transgenes in a turfgrass species.

Legumains - a family of asparagine-specific cysteine endopeptidases involved in propolypeptide processing and protein breakdown in plants

Legumains are a recently discovered family of plant and animal cysteine endopeptidases with a cleavage specificity for Asn in the P1 position of peptide bonds. Asp-flanked peptide bonds also are cleaved, but with a much lower efficiency. Legumains evolved from GPI transamidase-like progenitors. Sequence analysis revealed three major groups of plant legumains corresponding to differences in the developmental and organ-specific gene expression. With the exception of a single cell wall specific representative, all legumains occur in the vacuolar compartment. Legumains are either involved in protein degradation or play a role in the processing of precursor proteins by Asn/Asp-specific limited proteolysis. Which function legumains perform depends on the conformational state of the substrate protein. A legumain acts as a vacuolar processing enzyme when it only has access to the regular processing sites of a precursor polypeptide, but it acts as a degradative enzyme when an altered conformation opens the substrate for unlimited proteolysis. The specificity of these interactions seems to be the result of a co-evolution of enzyme and substrate. The double function of legumains is particularly evident in the events of deposition and mobilisation of storage globulins during seed maturation and germination/seedling growth and in senescing and dying cells.

Organ-specific expression of brassinosteroid-biosynthetic genes and distribution of endogenous brassinosteroids in Arabidopsis

Organ-specific expression of brassinosteroid-biosynthetic genes and distribution of endogenous brassinosteroids in Arabidopsis

Organ-specific expression of brassinosteroid-biosynthetic genes and distribution of endogenous brassinosteroids in Arabidopsis(Abstracts of The 37th Annual Meeting of the Japanese Society for Chemical Regulation of Plants)

Organ-specific expression of brassinosteroid-biosynthetic genes and distribution of endogenous brassinosteroids in Arabidopsis(Abstracts of The 37th Annual Meeting of the Japanese Society for Chemical Regulation of Plants)

Organ-Specific Molecular Classification of Primary Lung, Colon, and Ovarian Adenocarcinomas Using Gene Expression Profiles

Molecular classification of tumors based on their gene expression profiles promises to significantly refine diagnosis and management of cancer patients. The establishment of organ-specific gene expression patterns represents a crucial first step in the clinical application of the molecular approach. Here, we report on the gene expression profiles of 154 primary adenocarcinomas of the lung, colon, and ovary. Using high-density oligonucleotide arrays with 7129 gene probe sets, comprehensive gene expression profiles of 57 lung, 51 colon, and 46 ovary adenocarcinomas were generated and subjected to principle component analysis and to a cross-validated prediction analysis using nearest neighbor classification. These statistical analyses resulted in the classification of 152 of 154 of the adenocarcinomas in an organ-specific manner and identified genes expressed in a putative tissue-specific manner for each tumor type. Furthermore, two tumors were identified, one in the colon group and another in the ovarian group, that did not conform to their respective organ-specific cohorts. Investigation of these outlier tumors by immunohistochemical profiling revealed the ovarian tumor was consistent with a metastatic adenocarcinoma of colonic origin and the colonic tumor was a pleomorphic mesenchymal tumor, probably a leiomyosarcoma, rather than an epithelial tumor. Our results demonstrate the ability of gene expression profiles to classify tumors and suggest that determination of organ-specific gene expression profiles will play a significant role in a wide variety of clinical settings, including molecular diagnosis and classification.