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Y Ji

Publications and source records attributed to Y Ji.

At least 19 recordsLinked to original sources
PubMed2000

Disruption of a single copy of the SERCA2 gene results in altered Ca2+ homeostasis and cardiomyocyte function.

A mouse model carrying a null mutation in one copy of the sarcoplasmic reticulum (SR) Ca(2+)-ATPase isoform 2 (SERCA2) gene, in which SERCA2 protein levels are reduced by approximately 35%, was used to investigate the effects of decreased SERCA2 level on intracellular Ca(2+) homeostasis and contractile properties in isolated cardiomyocytes. When compared with wild-type controls, SR Ca(2+) stores and Ca(2+) release in myocytes of SERCA2 heterozygous mice were decreased by approximately 40-60% and approximately 30-40%, respectively, and the rate of myocyte shortening and relengthening were each decreased by approximately 40%. However, the rate of Ca(2+) transient decline (tau) was not altered significantly, suggesting that compensation was occurring in the removal of Ca(2+) from the cytosol. Phospholamban, which inhibits SERCA2, was decreased by approximately 40% in heterozygous hearts, and basal phosphorylation of Ser-16 and Thr-17, which relieves the inhibition, was increased approximately 2- and 2.1-fold. These results indicate that reduced expression and increased phosphorylation of phospholamban provides compensation for decreased SERCA2 protein levels in heterozygous heart. Furthermore, both expression and current density of the sarcolemmal Na(+)-Ca(2+) exchanger were up-regulated. These results demonstrate that a decrease in SERCA2 levels can directly modify intracellular Ca(2+) homeostasis and myocyte contractility. However, the resulting deficit is partially compensated by alterations in phospholamban/SERCA2 interactions and by up-regulation of the Na(+)-Ca(2+) exchanger.

Animals↗
PubMed2000

Overexpression of SERCA2b in the heart leads to an increase in sarcoplasmic reticulum calcium transport function and increased cardiac contractility.

The sarcoplasmic reticulum calcium ATPase SERCA2b is an alternate isoform encoded by the SERCA2 gene. SERCA2b is expressed ubiquitously and has a higher Ca(2+) affinity compared with SERCA2a. We made transgenic mice that overexpress the rat SERCA2b cDNA in the heart. SERCA2b mRNA level was approximately approximately 20-fold higher than endogenous SERCA2b mRNA in transgenic hearts. SERCA2b protein was increased 8-10-fold in the heart, whereas SERCA2a mRNA/protein level remained unchanged. Confocal microscopy showed that SERCA2b is localized preferentially around the T-tubules of the SR, whereas SERCA2a isoform is distributed both transversely and longitudinally in the SR membrane. Calcium-dependent calcium uptake measurements showed that the maximal velocity of Ca(2+) uptake was not changed, but the apparent pump affinity for Ca(2+) (K(0.5)) was increased in SERCA2b transgenic mice (0.199 +/- 0.011 micrometer) compared with wild-type control mice (0.269 +/- 0.012 micrometer, p < 0.01). Work-performing heart preparations showed that SERCA2b transgenic hearts had a higher rates of contraction and relaxation, shorter time to peak pressure and half-time for relaxation than wild-type hearts. These data show that SERCA2b is associated in a subcompartment within the sarcoplasmic reticulum of cardiac myocytes. Overexpression of SERCA2b leads to an increase in SR calcium transport function and increased cardiac contractility, suggesting that SERCA2b plays a highly specialized role in regulating the beat-to-beat contraction of the heart.

Animals↗
PubMed2000

Effects of cholinergic agents on spontaneous activity of nucleus basalis magnocellularis neurons in different age rats.

The present study investigated the effects of intracerebroventricular (i.c.v.) injection of acetylcholine (Ach), atropine and tubocurarine on the spontaneous activity of nucleus basalis Magnocellularis (nbM) neurons in young, adult and old rats. I.c.v. injection of Ach (1, 10 and 100 mM) dose-dependently increased the spontaneous firing rate in most of the nbM neurons (66.7%). The Ach-induced excitation effect on the spontaneous firing of nbM neurons was decreased with aging. I.c.v. injection of atropine (2.5, 25 and 250 mM) or tubocurarine (0.1, 1 and 10 mM) not only antagonized the Ach-induced excitation, but also inhibited the spontaneous firing of nbM neurons. The inhibitory effects were gradually decreased with aging. These results suggested that there might be some functional changes in nbM neurons with aging, which impair their responsive ability to stimuli of drugs.

Acetylcholine↗
PubMed2000

Nucleotide sequence analyses of partial envgp46 gene of human T-lymphotropic virus type I from inhabitants of Fujian Province in Southeast China.

Partial sequences from the env(gp46) gene of two human T-lymphotropic virus type I (HTLV-I) isolates (LIN and WEN) obtained from inhabitants of Fujian Province in southeast China were analyzed. A phylogenetic tree was constructed from these sequence data and those of other known HTLV-I isolates from all over the world. Comparisons of the LIN and WEN nucleotide sequences with other HTLV-I isolates showed diversity ranging from 0.73 to 7.00% for LIN and from 0.87 to 7.00% for WEN. Sequences of isolates LIN, WEN, MT-2, TSP1, and CH were most closely related, and the phylogenetic tree showed that all belong to the widespread subtype A of the cosmopolitan group. These preliminary data indicate that HTLV-I isolates from Fujian Province, China are closely related to HTLV-I strains from Japan and the Caribbean.

Base Sequence↗
PubMed2000

The expression of SR calcium transport ATPase and the Na(+)/Ca(2+)Exchanger are antithetically regulated during mouse cardiac development and in Hypo/hyperthyroidism.

The mouse has been used extensively for generating transgenic animal models to study cardiovascular disease. Recently, a number of transgenic mouse models have been created to investigate the importance of sarcoplasmic reticulum (SR) Ca(2+)transport proteins in cardiac pathophysiology. However, the expression and regulation of cardiac SR Ca(2+)ATPase and other Ca(2+)transport proteins have not been studied in detail in the mouse. In this study, we used multiplex RNase mapping analysis to determine SERCA2, phospholamban (PLB), and Na(+)/Ca(2+)-exchanger (NCX-1) gene expression throughout mouse heart development and in hypo/hyperthyroid animals. Our results demonstrate that the expression of SERCA2 and PLB mRNA increase eight-fold from fetal to adult stages, indicating that SR function increases with heart development. In contrast, the expression of the Na(+)/Ca(2+)-exchanger gene is two-fold higher in fetal heart compared to adult. Our study also makes the important observation that in hypothyroidic hearts the NCX-1 mRNA and protein levels were upregulated, whereas the SERCA2 mRNA/protein levels were downregulated. In hyperthyroidic hearts, however, an opposite response was identified. These findings are important and point out that the expression of NCX-1 is regulated antithetically to that of SERCA2 during heart development and in response to alterations in thyroid hormone levels.

Animals↗
PubMed2000

Structure of the highly conserved HERC2 gene and of multiple partially duplicated paralogs in human.

Recombination between chromosome-specific low-copy repeats (duplicons) is an underlying mechanism for several genetic disorders. Recently, a chromosome 15 duplicon was discovered in the common breakpoint regions of Prader-Willi and Angelman syndrome deletions. We identified previously the large HERC2 transcript as an ancestral gene in this duplicon, with approximately 11 HERC2-containing duplicons, and demonstrated that recessive mutations in mouse Herc2 lead to a developmental syndrome, juvenile development and fertility 2 (jdf2). We have now constructed and sequenced a genomic contig of HERC2, revealing a total of 93 exons spanning approximately 250 kb and a CpG island promoter. A processed ribosomal protein L41 pseudogene occurs in intron 2 of HERC2, and putative VNTRs occur in intron 70 (28 copies, approximately 76-bp repeat) and 3' exon 40 through intron 40 (6 copies, approximately 62-bp repeat). Sequence comparisons show that HERC2-containing duplicons have undergone several deletion, inversion, and dispersion events to form complex duplicons in 15q11, 15q13, and 16p11. To further understand the developmental role of HERC2, a highly conserved Drosophila ortholog was characterized, with 70% amino acid sequence identity to human HERC2 over the carboxy-terminal 743 residues. Combined, these studies provide significant insights into the structure of complex duplicons and into the evolutionary pathways of formation, dispersal, and genomic instability of duplicons. Our results establish that some genes not only have a protein coding function but can also play a structural role in the genome.

Amino Acid Sequence↗
PubMed2000

Structure of chromosomal duplicons and their role in mediating human genomic disorders.

Chromosome-specific low-copy repeats, or duplicons, occur in multiple regions of the human genome. Homologous recombination between different duplicon copies leads to chromosomal rearrangements, such as deletions, duplications, inversions, and inverted duplications, depending on the orientation of the recombining duplicons. When such rearrangements cause dosage imbalance of a developmentally important gene(s), genetic diseases now termed genomic disorders result, at a frequency of 0.7-1/1000 births. Duplicons can have simple or very complex structures, with variation in copy number from 2 to >10 repeats, and each varying in size from a few kilobases in length to hundreds of kilobases. Analysis of the different duplicons involved in human genomic disorders identifies features that may predispose to recombination, including large size and high sequence identity between the recombining copies, putative recombination promoting features, and the presence of multiple genes/pseudogenes that may include genes expressed in germ cells. Most of the chromosome rearrangements involve duplicons near pericentromeric regions, which may relate to the propensity of such regions to accumulate duplicons. Detailed analyses of the structure, polymorphic variation, and mechanisms of recombination in genomic disorders, as well as the evolutionary origin of various duplicons will further our understanding of the structure, function, and fluidity of the human genome.

Animals↗
PubMed1999

Hepatic scavenger receptor BI promotes rapid clearance of high density lipoprotein free cholesterol and its transport into bile.

The clearance of free cholesterol from plasma lipoproteins by tissues is of major quantitative importance, but it is not known whether this is passive or receptor-mediated. Based on our finding that scavenger receptor BI (SR-BI) promotes free cholesterol (FC) exchange between high density lipoprotein (HDL) and cells, we tested whether SR-BI would effect FC movement in vivo using [(14)C]FC- and [(3)H]cholesteryl ester (CE)-labeled HDL in mice with increased (SR-BI transgenic (Tg)) or decreased (SR-BI attenuated (att)) hepatic SR-BI expression. The initial clearance of HDL FC was increased in SR-BI Tg mice by 72% and decreased in SR-BI att mice by 53%, but was unchanged in apoA-I knockout mice compared with wild-type mice. Transfer of FC to non-HDL and esterification of FC were minor and could not explain differences. The hepatic uptake of FC was increased in SR-BI Tg mice by 34% and decreased in SR-BI att mice by 22%. CE clearance and uptake gave similar results, but with much slower rates. The uptake of HDL FC and CE by SR-BI Tg primary hepatocytes was increased by 2.2- and 2.6-fold (1-h incubation), respectively, compared with control hepatocytes. In SR-BI Tg mice, the initial biliary secretion of [(14)C]FC was markedly increased, whereas increased [(3)H]FC appeared after a slight delay. Thus, in the mouse, a major portion of the clearance of HDL FC from plasma is mediated by SR-BI.

Animals↗
PubMed1999

A transgene insertion creating a heritable chromosome deletion mouse model of Prader-Willi and angelman syndromes.

Prader-Willi syndrome (PWS) and Angelman syndrome (AS) result from the loss of function of imprinted genes in human chromosome 15q11-q13. The central part of mouse chromosome 7 is homologous to human 15q11-q13, with conservation of both gene order and imprinted features. We report here the characterization of a transgene insertion (Epstein-Barr virus Latent Membrane Protein 2A, LMP2A) into mouse chromosome 7C, which has resulted in mouse models for PWS and AS dependent on the sex of the transmitting parent. Epigenotype (allelic expression and DNA methylation) and fluorescence in situ hybridization analyses indicate that the transgene-induced mutation has generated a complete deletion of the PWS/AS-homologous region but has not deleted flanking loci. Because the intact chromosome 7, opposite the deleted homolog, maintains the correct imprint in somatic cells of PWS and AS mice and establishes the correct imprint in male and female germ cells of AS mice, homologous association and replication asynchrony are not part of the imprinting mechanism. This heritable-deletion mouse model will be particularly useful for the identification of the etiological genes and mechanisms, phenotypic basis, and investigation of therapeutic approaches for PWS.

Angelman Syndrome↗
PubMed1999

Analysis of sarcoplasmic reticulum Ca2+ transport and Ca2+ ATPase enzymatic properties using mouse cardiac tissue homogenates.

Recent studies have focused on developing transgenic mouse models to explore the physiological roles of sarcoplasmic reticulum (SR) calcium handling proteins. The goal of this study was to develop methodology to measure SR Ca2+ transport function and enzymatic properties of SR Ca2+ ATPase (SERCA) in individual mouse hearts. We describe here the procedures to specifically measure SR Ca2+ uptake, the formation and decomposition of SERCA phosphoenzyme intermediate (E-P) in mouse cardiac homogenates. The specificity of SERCA enzymatic activity in cardiac homogenates was established by (a) the selective inhibition of SERCA enzyme by inhibitor-thapsigargin, and (b) comparison of the kinetic parameters of SERCA activity between homogenates and isolated microsomes. Here we show that the apparent affinity of SERCA for Ca2+ and ATP, the time to reach steady-state levels of E-P, and the rate of E-P decomposition (turnover rate of SERCA enzyme) are similar in homogenates and microsomes. These studies demonstrate that SERCA Ca2+ transport and enzymatic properties can be accurately measured in mouse cardiac tissue homogenates. Additionally, we show that frozen cardiac homogenates can be used without significant loss of enzymatic activity. In conclusion, we have developed and established the methods to employ tissue homogenates to study SR Ca2+ transport function in individual mouse hearts.

Animals↗
PubMed1999

S-nitrosylation and S-glutathiolation of protein sulfhydryls by S-nitroso glutathione.

The modification of reactive protein sulfhydryls by S-nitrosoglutathione and other NO donors has been studied by gel isoelectric focusing. S-nitrosylated, unmodified, and S-glutathiolated protein forms are differentiated by this method. With specific antibodies for the protein of interest, both S-nitrosylation and S-glutathiolation of the protein were analyzed in mixtures obtained as soluble tissue or cell extracts. The effect of S-nitrosoglutathione (GSNO) on purified phosphorylase b, on carbonic anhydrase III in an extract from rat liver, and on H-ras expressed in Escherichia coli was examined. When fresh GSNO reacted with pure phosphorylase b, only S-nitrosylated forms of the protein were observed. Likewise the NO donors, amyl nitrite, spermine NONOate, and diethylamine NONOate, all generated S-nitrosylated phosphorylase b. When crude mixtures of proteins from rat liver (containing carbonic anhydrase III) or from E. coli (containing an overexpressed form of H-ras) were exposed to fresh GSNO, both the S-nitrosylated and the S-glutathiolated forms of the proteins were observed. It is suggested that reactive intermediates from the breakdown of GSNO are responsible for the observed S-glutathiolation. These experiments show that both S-nitrosylated and S-glutathiolated forms of proteins may be generated by the addition of GSNO to mixtures containing proteins with reactive sulfhydryls. These protein modifications may exhibit metabolic consequences independent of the release of nitric oxide.

Animals↗
PubMed1999

Impaired cardiac performance in heterozygous mice with a null mutation in the sarco(endo)plasmic reticulum Ca2+-ATPase isoform 2 (SERCA2) gene.

The sarco(endo)plasmic reticulum Ca2+-ATPase isoform 2 (SERCA2) gene encodes both SERCA2a, the cardiac sarcoplasmic reticulum Ca2+ pump, and SERCA2b, which is expressed in all tissues. To gain a better understanding of the physiological functions of SERCA2, we used gene targeting to develop a mouse in which the promoter and 5' end of the gene were eliminated. Mating of heterozygous mutant mice yielded wild-type and heterozygous offspring; homozygous mutants were not observed. RNase protection, Western blotting, and biochemical analysis of heart samples showed that SERCA2 mRNA was reduced by approximately 45% in heterozygous mutant hearts and that SERCA2 protein and maximal velocity of Ca2+ uptake into the sarcoplasmic reticulum were reduced by approximately 35%. Measurements of cardiovascular performance via transducers in the left ventricle and right femoral artery of the anesthetized mouse revealed reductions in mean arterial pressure, systolic ventricular pressure, and the absolute values of both positive and negative dP/dt in heterozygous mutants. These results demonstrate that two functional copies of the SERCA2 gene are required to maintain normal levels of SERCA2 mRNA, protein, and Ca2+ sequestering activity, and that the deficit in Ca2+ sequestering activity due to the loss of one copy of the SERCA2 gene impairs cardiac contractility and relaxation.

Animals↗
PubMed1999

Molecular characterization of radiation- and chemically induced mutations associated with neuromuscular tremors, runting, juvenile lethality, and sperm defects in jdf2 mice.

The juvenile development and fertility-2 (jdf2) locus, also called runty-jerky-sterile (rjs), was originally identified through complementation studies of radiation-induced p-locus mutations. Studies with a series of ethylnitrosourea (ENU)-induced jdf2 alleles later indicated that the pleiotropic effects of these mutations were probably caused by disruption of a single gene. Recent work has demonstrated that the jdf2 phenotype is associated with deletions and point mutations in Herc2, a gene encoding an exceptionally large guanine nucleotide exchange factor protein thought to play a role in vesicular trafficking. Here we describe the molecular characterization of a collection of radiation- and chemically induced jdf2/Herc2 alleles. Ten of the 13 radiation-induced jdf2 alleles we studied are deletions that remove specific portions of the Herc2 coding sequence; DNA rearrangements were also detected in two additional mutations. Our studies also revealed that Herc2 transcripts are rearranged, not expressed, or are present in significantly altered quantities in animals carrying most of the jdf2 mutations we analyzed, including six independent ENU-induced alleles. These data provide new molecular clues regarding the wide range of jdf2 and p phenotypes that are expressed by this collection of recently generated and classical p-region mutations.

Alleles↗
PubMed1999

New ribosomal RNA gene locations in Gossypium hirsutum mapped by meiotic FISH.

In this study we have mapped newly identified rDNA loci in Gossypium hirsutum. Four new minor 18S-26S rDNA loci, in addition to the sites previously identified, were mapped using fluorescence in situ hybridization (FISH) to heterozygous translocation (NT) quadrivalents (IVs). The newly detected 18S-26S rDNA loci were mapped to the right arms of chromosomes 8, 9, 15, 17, 19, 20, and 23 and the left arms of chromosomes 5, 11, 12, and 14. Using the rDNA loci as common reference points, we detected several erroneous arm assignments in the previously published map of NT breakpoints. The data are summarized in the form of an integrated map for all 17 known rDNA loci, relative to centromeres, telomeres, and NT breakpoints. This information will facilitate future locus-specific research on rRNA gene evolution and function.

Chromosome Mapping↗
PubMed1999

Junctional diversity in Xenopus immunoglobulin light chains.

Xenopus cDNA sequences encoding the homolog of mammalian kappa (kappa) light (L) chains were isolated from isogenic tadpole and adult individuals to investigate whether there existed stage-specific immunoglobulin L chain expression and somatic diversification. In the course of these studies rearrangements to a sixth J(L) gene segment and a pseudogene (J(L)psi) were found, and it is suggested that the order of these gene segments with respect to the L chain constant (C) region exon is: J(L)6-J(L)1-J(L)2-J(L)3-J(L)4-J(L)5-J(L)psi-C(L). The cDNA junctional diversity was analyzed; few N and P regions were found and almost all the CDR3 were 9 codons in length. There were restricted patterns of recombination site resolution, and this is attributed to some constraint in JL coding end processing.

Animals↗