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At least 37 records · Page 2Linked to original sources
PubMed2000

Tissue-specific localization of cytochrome P450 aromatase in the equine embryo by in situ hybridization and immunocytochemistry.

Estrogen production by the preimplantation equine embryo is presumed to be important in maternal-conceptus communication in the mare. The synthesis of C(18) estrogens from C(19) androgens requires cytochrome P450 aromatase (P450(arom)) in the conceptus, but little information is available on the specific tissue location or potential developmental patterns of expression for the horse. The goal of this research was to localize P450(arom) in the equine conceptus by immunocytochemistry and in situ hybridization. Intact blastocyst-stage embryos were collected by nonsurgical flush on Days 12-15 of pregnancy, fixed in 4% paraformaldehyde, and paraffin-embedded. Aromatase protein was localized using rabbit anti-human placental aromatase antiserum with a detection system utilizing peroxidase and 3-amino-9-ethylcarbazole. For in situ hybridization, tissue sections were incubated with sense or antisense [(35)S]UTP-labeled cRNA probes prepared from equine aromatase cDNA. Aromatase protein and transcript were abundant in the extraembryonic trophectoderm but absent from embryonic ectoderm. No P450(arom) expression was detected in abembryonic endoderm or mesoderm. Aromatase expression was demonstrated in the endoderm beneath the disc (hypoblast). This pattern of P450(arom) expression in the equine blastocyst closely resembles that seen transiently in the porcine embryo, suggesting that regulatory mechanisms conferring tissue specificity may be conserved.

Animals↗
PubMed1991

Characterization of muscle and non muscle Xenopus laevis tropomyosin mRNAs transcribed from the same gene. Developmental and tissue-specific expression.

We have isolated and characterized three Xenopus laevis tropomyosin cDNAs (XTM alpha 2, XTM alpha 4 and XTM05). XTM alpha 2 and XTM alpha 4 were isolated from a stage-42 embryo cDNA library and XTM05 from an oocyte cDNA library. XTM alpha 2 and XTM alpha 4 both code for a 284-amino-acid protein homologous to the skeletal alpha tropomyosin but diverge in their 3' untranslated sequences. This divergence is due to an absence of splicing in the early embryo. XTM05 codes for a 248-amino-acid non-muscle tropomyosin. Sequence analysis indicates that the three cDNAs are generated from a single gene which uses two promoters and possibly three sets of alternate exons. Northern blot analysis, with specific probes against each cDNA, was used to study the expression of the X. laevis alpha tropomyosin gene during development and in specific tissues. This analysis showed a different temporal usage of the two promoters and characterized the tissue-specific expression of the different transcripts.

Amino Acid Sequence↗
PubMed1996

Binding specificity and tissue-specific expression pattern of the Arabidopsis bZIP transcription factor TGA2.

The binding specificity and tissue-specific expression pattern of TGA2 (AHBP-1b), an Arabidopsis bZIP transcription factor have been determined. Filter-binding and gel-shift assays showed that TGA2 has high affinity for C-boxes (ATGACGTCAT). In this respect TGA2 is similar to other members of the Arabidopsis TGA family (such as TGA1, TGA3 and OBF4) and to tobacco TGA1a. Genomic Southern blot analysis confirmed that TGA2 is a member of the gene family. Northern blot analysis showed that the gene is expressed at similar levels in root, stem, leaf and flower t at somewhat lower levels in siliques. TGA3 was also found to be expressed at the same level throughout the plant, whereas genes encoding TGA1 and OBF4 have relatively high RNA expression levels in root. The differential expression of these genes suggests that they have distinct functions.

Arabidopsis↗
PubMed1998

Insertion of two independent enhancers in the long terminal repeat of a self-inactivating vector results in high-titer retroviral vectors with tissue-specific expression.

The use of retroviral vectors (RVs) derived from the murine oncoretroviruses for gene therapy is associated with the risk of malignant transformation of infected cells and ectopic expression of the proteins of interest. Targeting retroviral vectors to specific tissues would increase their safety and clinical applicability. To explore the potential of targeting vector expression to skeletal muscle, the murine leukemia virus broad transcriptional tropism was modified by substituting the viral promoter and/or enhancer with a transcriptional cassette containing the human T cell leukemia virus type I Tax-responsive element and the minimal muscle creatine kinase enhancer and promoter. The resulting retroviral vectors could be transcriptionally trans-activated by tax. In the absence of Tax, however, the viruses showed muscle-specific expression. Trans-complementing packaging and indicator cells stably expressing Tax were used to isolate high-titer producer cell clones (10(6) CFU/ml). In vitro, the levels of expression of these RVs in Tax-expressing fibroblasts were 10,000-fold higher than in normal fibroblasts and 1000-fold higher in C2C12 myotubes than in C2C12 myoblasts. Expression of the vectors and the endogenous muscle creatine kinase gene was similarly dependent on the maturity of the muscle cultures. One vector with modified LTRs was also tested in vivo in regenerating muscle and showed a delayed pattern of expression in myofibers compared with the vector containing the wild-type LTRs. These vectors can be easily modified to contain different tissue-specific enhancer and promoter elements and the availability of complementing packaging and indicator cells expressing Tax should allow their application in a variety of gene therapy settings.

3T3 Cells↗
PubMed2001

Tissue-specific overexpression of lipoprotein lipase causes tissue-specific insulin resistance.

Insulin resistance in skeletal muscle and liver may play a primary role in the development of type 2 diabetes mellitus, and the mechanism by which insulin resistance occurs may be related to alterations in fat metabolism. Transgenic mice with muscle- and liver-specific overexpression of lipoprotein lipase were studied during a 2-h hyperinsulinemic-euglycemic clamp to determine the effect of tissue-specific increase in fat on insulin action and signaling. Muscle-lipoprotein lipase mice had a 3-fold increase in muscle triglyceride content and were insulin resistant because of decreases in insulin-stimulated glucose uptake in skeletal muscle and insulin activation of insulin receptor substrate-1-associated phosphatidylinositol 3-kinase activity. In contrast, liver-lipoprotein lipase mice had a 2-fold increase in liver triglyceride content and were insulin resistant because of impaired ability of insulin to suppress endogenous glucose production associated with defects in insulin activation of insulin receptor substrate-2-associated phosphatidylinositol 3-kinase activity. These defects in insulin action and signaling were associated with increases in intracellular fatty acid-derived metabolites (i.e., diacylglycerol, fatty acyl CoA, ceramides). Our findings suggest a direct and causative relationship between the accumulation of intracellular fatty acid-derived metabolites and insulin resistance mediated via alterations in the insulin signaling pathway, independent of circulating adipocyte-derived hormones.

Animals↗
PubMed2004

Moderate caloric restriction, but not physiological hyperleptinemia per se, enhances mitochondrial oxidative capacity in rat liver and skeletal muscle--tissue-specific impact on tissue triglyceride content and AKT activation.

The study aimed at determining, in lean tissues from nonobese rats, whether physiological hyperleptinemia with leptin-induced reduced caloric intake and/or calorie restriction (CR) per se: 1) enhance mitochondrial-energy metabolism gene transcript levels and oxidative capacity; and 2) reduce triglyceride content. Liver and skeletal muscles were collected from 6-month-old Fischer 344 rats after 1-wk leptin sc infusion (0.4 mg/kg . d: leptin + approximately 3-fold leptinemia vs. ad libitum-fed control) or moderate CR (-26% of those fed ad libitum) in pair-fed animals (CR). After 1 wk: 1) leptin and CR comparably enhanced transcriptional expression of mixed muscle mitochondrial genes (P < 0.05 vs. control); 2) CR independently increased (P < 0.05 vs. leptin-control) hepatic mitochondrial-lipooxidative gene expression and oxidative capacity; 3) hepatic but not muscle mitochondrial effects of CR were associated (P < 0.01) with increased activated insulin signaling at AKT level (P < 0.05 vs. leptin-control); 4) liver and muscle triglyceride content were comparable in all groups. In additional experiments, assessing time course of posttranscriptional CR effects, 3-wk superimposable CR (P < 0.05): 1) increased both liver and muscle mitochondrial oxidative capacity; and 2) selectively reduced muscle triglyceride content. Thus, in nonobese adult rat: 1) moderate CR induces early increments of mitochondrial-lipooxidative gene expression and time-dependent increments of oxidative capacity in liver and mixed muscle; 2) sustained moderate CR alters tissue lipid distribution reducing muscle but not liver triglycerides; 3) mitochondrial-lipid metabolism changes are tissue-specifically associated with hepatic AKT activation; 4) short-term physiological hyperleptinemia has no independent stimulatory effects on muscle and liver mitochondrial-lipooxidative gene expression. Increased lean tissue oxidative capacity could favor substrate oxidation over storage during reduced nutrient availability.

Animals↗
PubMed2005

Modular genes with metazoan-specific domains have increased tissue specificity.

We have systematically examined the domain composition across a comprehensive set of tissue-specific, midrange and housekeeping genes as defined by their mode of expression in 52 normal mouse tissues. We show a definite correlation between the number of domains and the degree of tissue specificity. This trend is further supported by a novel analysis involving the time of origin of each domain. Genes containing metazoan-specific domains are more prevalent in signal transduction and cell-communication pathways, and are depleted in primary metabolism. Our analyses suggest that highly modular gene products have been recruited for tissue-specific functions that are required in complex organisms.

Animals↗
PubMed1983

20-hydroxyecdysone stimulates tissue-specific yolk-protein gene transcription in both male and female Drosophila.

The yolk polypeptides of Drosophila are normally synthesized in the fat body and ovarian follicle cells of adult females. In response to 20-hydroxyecdysone males synthesize yolk polypeptides. The actual level of yolk polypeptides synthesized in males is not always a direct reflection of the YP-transcripts present. Initially YP-transcripts are efficiently translated into polypeptides whereas later they are not and the YP-transcripts can have a half-life of less than 8 h in males. We suggest that the expression of the genes coding for the yolk polypeptides in males may be regulated at transcriptional and translational levels. Treatment of females with 20-hydroxyecdysone leads to a transient increase in YP-transcript accumulation, but the response is difficult to assess in whole flies due to the high variability in transcript levels during normal development. Analysing the response to 20-hydroxyecdysone at the level of specific tissues shows that transcript accumulation is dramatically increased in body walls (fat-body cells, epidermis and oenocytes) of both males and females. Gut, Malpighian tubules, testis and ovaries are not affected. Treatment of females with 20-hydroxyecdysone followed by measuring YP-transcript accumulation over the next 24 h in ovaries and body walls separately, confirms that only body walls respond to the hormone. There is an increase in yolk-polypeptide synthesis during the period of increased YP-transcript accumulation in females. We conclude that the response of the YP-genes to 20-hydroxyecdysone is tissue-, but not sex-specific.

Animals↗
PubMed2004

Tissue-specific imprinting of the G protein Gsalpha is associated with tissue-specific differences in histone methylation.

The G protein Gsalpha is imprinted in a tissue-specific manner, being primarily expressed from the maternal allele in some tissues, such as renal proximal tubules. The Gsalpha promoter is unmethylated, but is downstream of a differentially methylated region [the exon 1A differentially methylated region (DMR)] that is methylated on the maternal allele. Maternal Gsalpha null mutations or loss of maternal-specific exon 1A methylation leads to pseudohypoparathyroidism types 1A or 1B, respectively. We now have examined the chromatin state of each parental allele within the exon 1A-Gsalpha promoter region by chromatin immunoprecipitation of samples derived from mice with heterozygous deletions within the region using antibodies to covalently modified histones. The exon 1A DMR had allele-specific differences in histone acetylation and methylation, with histone acetylation and H3 lysine 4 (H3K4) methylation of the paternal allele, and H3 lysine 9 (H3K9) methylation of the maternal allele. Both parental alleles had similar levels of histone acetylation and H3K4 methylation within the Gsalpha promoter and first exon, with no H3K9 methylation. In liver, where Gsalpha is biallelically expressed, both parental alleles had similar levels of tri- and dimethylated H3K4 within the Gsalpha first exon. In contrast, in renal proximal tubules there was a greater ratio of tri- to dimethylated H3K4 of Gsalpha exon 1 in the more transcriptionally active maternal as compared with the paternal allele. These results show that allele-specific differences in Gsalpha expression correlate in a tissue-specific manner with allele-specific differences in the extent of H3K4 methylation, and are the first demonstration that chronic transcriptional activation in mammals is correlated with trimethylation of H3K4.

Animals↗
PubMed2003

Tissue-specific p53 responses to ionizing radiation and their genetic modification: the key to tissue-specific tumour susceptibility?

Although little is understood of the underlying mechanisms, there are tissue-specific responses to tumourigenic and therapeutic agents and these responses are influenced by genetic factors. Ionizing radiation is an important tumourigenic and therapeutic agent for which there is substantial evidence for such tissue-dependent and genotype-dependent responses. Because the p53 tumour suppressor protein is a major determinant of cellular responses to radiation, the present study has investigated whether modification of the p53 pathway contributes to tissue-dependent and genotype-dependent responses using inbred strains of mice. Comparison of responses in haemopoietic and epithelial cells in irradiated C57BL/6 and DBA/2 mice revealed significant differences in p53 and apoptotic responses in different cell types and in different cells of the same type, reflecting the complexity of damage responses operating in the whole organism. The data suggest that p53-mediated up-regulation of Bax is a major determinant of apoptosis in the spleen, but not in the intestine, whereas p53-mediated induction of p21(waf1) plays an anti-apoptotic role in the spleen, but not in the intestine. It is also shown that p53 stabilization and differential transactivational activities towards Bax or p21(waf1) are influenced by genetic factors that act in a tissue-specific manner. Analysis of ATM, a potential mediator of differential p53 activation, indicates that this key regulator of radiation responses is preferentially induced in epithelial cells, but is unlikely to account for genetic modification of p53 or apoptotic responses in the mouse strains studied. Polymorphisms in the p53 or DNA-PKcs genes are also unlikely to account for the genetic modifications that are reported here. There are numerous further potential modifiers of the p53 pathway, but analysis of backcross and inter-cross mice demonstrates that genes responsible for the complex modification of these in vivo responses can be identified by linkage analysis. This approach has the potential to reveal new or unexpected interactions involving the p53 pathway that determine both short-term and long-term effects of radiation exposure and the basis of tissue-specific responses and tumour susceptibility.

Animals↗
PubMed1996

Galectin-1 from bovine spleen: biochemical characterization, carbohydrate specificity and tissue-specific isoform profiles.

Selected biochemical properties, including the charge heterodispersity profile and carbohydrate specificity, of bovine galectin-1 were determined in detail. The lectin was purified through an improved purification protocol that yielded 35-40 mg/kg of wet tissue with a specific activity of 1.7-2 x 10(4) mg-1.ml. The galectin is a homodimer of approximately 14.5 kDa subunits with E(280)mg/ml of 0.65 ml.mg-1.cm-1. When stored in the presence of its carbohydrate ligand, the lectin's binding activity remained stable in a non-reducing environment even at room temperature. The optimal pH for binding to the ligand was 6.5-8.0. The overall carbohydrate specificity of the bovine galectin-1 isolated from spleen is similar to that of the galectin isolated from heart and to other mammalian galectins that exhibit "conserved" (Type I) carbohydrate recognition domains (CRDs) [Ahmed, H. and Vasta, G.R. (1994) Glycobiology 4, 545-549], but differs from those from Xenopus laevis and rat intestine domain I. The fluorescence of 4-methylumbelliferyl alpha-D-galactopyranoside was quenched on binding to bovine spleen galectin-1. Scatchard plots of data obtained at 5, 15, and 30 degrees C showed that the galectin has two sugar exothermic binding sites with association constants of 3.4 x 10(5), 1.0 x 10(5), and 0.3 x 10(5), respectively. Chemical modification studies indicated that histidine, tryptophan, carboxylic acid, and arginine, but not lysine or tyrosine, are involved in the binding to the carbohydrate ligand. On isoelectric focusing, the spleen galectin-1 appeared as six isoforms ranging from pI4.56-4.88 with main components at pI 4.63 (34.0%), 4.73 (42.6%), and 4.88 (16.6%). The galectin-1 isolated from heart yielded a quali- and quantitatively different profile with four isoforms ranging from pI 4.53-4.73, those with pIs of 4.56, 4.63, and 4.73 being common to the spleen homolog. Edman degradation of selected peptides purified from the spleen galectin-1 digest revealed amino acid sequences identical to those obtained for the heart galectin-1. This suggests that although point mutations in the subunit primary structure may not be the likely source of isolectins, as observed for X. laevis, tissue-specific co- or post-translational modifications may be the possible cause of the differences in the galectin isoform profile between bovine spleen and heart.

Amino Acid Sequence↗
PubMedDate not supplied

A novel full-scale flat membrane bioreactor utilizing porcine hepatocytes: cell viability and tissue-specific functions.

When designing an extracorporeal hybrid liver support device, special attention should be paid to providing the architectural basis for reconstructing a proper cellular microenvironment that ensures highest and prolonged functional activity of the liver cells. The common goal is to achieve high cell density culture and to design the bioreactor for full-scale primary liver cell cultures under adequate mass transfer conditions. An important aim of this study was to evaluate the biochemical performance of a flat membrane bioreactor that permits high-density hepatocyte culture and simultaneously to culture cells under sufficient oxygenation availability conditions comparable to the in vivo-like microenvironment. In such a bioreactor pig liver cells were cultured within an extracellular matrix between oxygen-permeable flat-sheet membranes. In this investigation we used a novel scaled-up prototype consisting of up to 20 modules in a parallel mode. Each module was seeded with 2 x 10(8) cells. Microscopic examination of the hepatocytes revealed morphological characteristics as found in vivo. Cell concentration increased in the first days of culture, as indicated by DNA measurements. The performance of the bioreactor was monitored for 18 days in terms of albumin synthesis, urea synthesis, ammonia elimination, and diazepam metabolism. The ability of the hepatocytes to synthesize albumin and urea increased during the first days of culture. Higher rates of albumin synthesis were obtained at day 9 and remained at a value of 1.41 pg/h/cell until day 18 of culture. The rate of urea synthesis increased from 23 ng/h/cell to 28 ng/h/cell and then remained constant. Cells eliminated ammonia at a rate of about 56 pg/h/cell, which was constant over the experimental period. Hepatocytes in the bioreactor metabolized diazepam and generated three different metabolites: nordiazepam, temazepam, and oxazepam. The production of such metabolites was sustained until 18 days of culture. These results demonstrated that the scale-up of the bioreactor was assessed, and it could be demonstrated that the device design aimed at the reconstruction of the liver-specific tissue architecture supported the expression of liver-specific functions of primary pig liver cells.

Albumins↗
PubMed1995

Molecular cloning of a small DNA binding protein with specificity for a tissue-specific negative element within the rps1 promoter.

A cDNA encoding a specific binding activity for the tissue-specific negative cis-element S1F binding site of spinach rps1 was isolated from a spinach cDNA expression library. This cDNA of 0.7 kb encodes an unusual small peptide of only 70 amino acids, with a basic domain which contains a nuclear localization signal and a putative DNA binding helix. This protein, named S1Fa, is highly conserved between dicotyledonous and monocotyledonous plants and may represent a novel class of DNA binding proteins. The corresponding mRNA is accumulated more in roots and in etiolated seedlings than in green leaves. This expression pattern is correlated with the tissue-specific function of the S1F binding site which represses the rps1 promoter preferentially in roots and in etiolated plants.

Amino Acid Sequence↗
PubMed1997

A novel juxtamembrane domain isoform of HER4/ErbB4. Isoform-specific tissue distribution and differential processing in response to phorbol ester.

Human epidermal growth factor receptor 4 (HER4) is a member of the epidermal growth factor (EGF) receptor subfamily of receptor tyrosine kinases that is activated by neuregulins (NRG), betacellulin (BTC), and heparin-binding EGF-like growth factor. Sequencing of full-length human HER4 cDNAs revealed the existence of two HER4 isoforms that differed by insertion of either 23 or 13 alternative amino acids in the extracellular juxtamembrane (JM) region. The 23-amino acid form (HER4 JM-a) and the 13-amino acid form (HER4 JM-b) were expressed in a tissue-specific manner, as demonstrated by reverse transcriptase-polymerase chain reaction analysis of mouse and human tissues. Both isoforms were expressed in neural tissues such as cerebellum, whereas kidney expressed HER4 JM-a only and heart HER4 JM-b only. In situ hybridization using specific oligonucleotides demonstrated transcription of both JM-a and JM-b isoforms in the mouse cerebellum. Tyrosine phosphorylation analysis indicated that both receptor isoforms were activated to the same extent by NRG-beta1 and BTC, and to a lesser extent by NRG-alpha1 and heparin-binding EGF-like growth factor. A functional difference was found, however, in response to phorbol ester treatment. Stimulation of cells with phorbol ester resulted in a loss of 125I-NRG-beta1 binding and in a reduction of total cell-associated HER4 protein in HER4 JM-a transfectants but not in HER4 JM-b transfectants. It was concluded that novel alternatively spliced isoforms of HER4 exist, that they are distributed differentially in vivo in mouse and human tissues, that they are both activated by HER4 ligands, and that they may represent cleavable and noncleavable forms of HER4.

Amino Acid Sequence↗
PubMedDate not supplied

Protein metabolism in alcoholism: effects on specific tissues and the whole body.

Ethanol is one of the few nutrients that is profoundly toxic. Alcohol causes both whole-body and tissue-specific changes in protein metabolism. Chronic ethanol missuse increases nitrogen excretion with concomitant loss of lean tissue mass. Even acute doses of alcohol elicit increased nitrogen excretion. The loss of skeletal muscle protein (i.e., chronic alcoholic myopathy) is one of several adverse reactions to alcohol and occurs in up to two-thirds of all ethanol misusers. There are a variety of other diseases and tissue abnormalities that are entirely due to ethanol-induced changes in the amounts of individual proteins or groups of tissue proteins; for example, increased hepatic collagen in cirrhosis, reduction in myosin in cardiomyopathy, and loss of skeletal collagen in osteoporosis. Ethanol induces changes in protein metabolism in probably all organ or tissue systems. Clinical studies in alcoholic patients without overt liver disease show reduced rates of skeletal muscle protein synthesis though whole-body protein turnover does not appear to be significantly affected. Protein turnover studies in alcohol misusers are, however, subject to artifactual misinterpretations due to non-abstinence, dual substance misuse (e.g., cocaine or tobacco), specific nutritional deficiencies, or the presence of overt organ dysfunction. As a consequence, the most reliable data examining the effects of alcohol on protein metabolism is derived from animal studies, where nutritional elements of the dosing regimen can be strictly controlled. These studies indicate that, both chronically and acutely, alcohol causes reductions in skeletal muscle protein synthesis, as well as of skin, bone, and the small intestine. Chronically, animal studies also show increased urinary nitrogen excretion and loss of skeletal muscle protein. With respect to skeletal muscle, the reductions in protein synthesis do not appear to be due to the generation of reactive oxygen species, are not prevented with nitric oxide synthase inhibitors, and may be indirectly mediated by the reactive metabolite acetaldehyde. Changes in skeletal muscle protein metabolism have profound implications for whole body physiology, while protein turnover changes in organs such as the heart (exemplified by complex alterations in protein profiles) have important implications for cardiovascular function and morbidity.

Alcoholism↗
PubMed2006

Variability of taurine concentrations in Mytilus galloprovincialis as a function of body size and specific tissue.

Variability of taurine (2-aminoethane sulfonic acid) was studied as a function of size in the mussel Mytilus galloprovincialis and tissue specificity. Isometric and/or allometric relationships were established with regard to total soft mass of the mussels between 20 and 60 mm shell length. Relative amounts of taurine dropped significantly with increasing mass of whole soft tissues with an allometric coefficient value of -0.15. The inverse relationship between taurine and increasing size of mussels was confirmed for gill epithelium and labial palp (allometric coefficient values of -0.16 and -0.10, respectively), tissues that, in turn, represented isometric functions with regard to total soft mass. Although relative amounts of taurine were significantly different in digestive gland, mantle and foot, relationships with increasing size of mussels remained unchanged in these tissues. Gill area of mussels was related to soft mass with an allometric coefficient of 0.70 by 2D Image Analysis, but increased to 0.85 when introducing a third dimension, i.e. gill thickness. Results are discussed according to gill structure analysis and taurine functionality.

Animals↗