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Y L Sun

Publications and source records attributed to Y L Sun.

At least 37 records · Page 2Linked to original sources
PubMed1998

The influence of mutation at Glu44 and Glu56 of cytochrome b5 on the protein's stabilization and interaction between cytochrome c and cytochrome b5.

To characterize the roles played by Glu44 and Glu56 of cytochrome b5 in the formation of the electrostatic complex between cytochrome c and cytochrome b5, the Glu44, Glu56, or both sites were changed to alanine by site-directed mutagenesis. The influence of these two residues on the protein stability was probed by investigating the kinetic behaviors of protein denaturation in urea or upon heating and the heme-transfer reactions between apo-myoglobin and the variants of cytochrome b5. It has been found that when the Glu44 and/or Glu56 are mutated to alanine, the protein stability increases slightly due to the fact that the hydrophilic residue is changed to a hydrophobic one, resulting in the two pairs of helices surrounding the heme taking a more compact conformation. The difference in voltammetric behavior of cytochrome c, cytochrome b5, and its three mutants, Cyt b5 E44A, E56A, and E44/56A, alone and in 1:1 protein complexes demonstrates that both Glu44 and Glu56 of cytochrome b5 take part in the electrostatic interaction with cytochrome c. The entropy changes, DeltaS degreesrc and enthalpy changes, DeltaH degrees, derived from the temperature dependence of the formal reduction potentials of each protein in different protein systems suggest that, because of the mutual interaction with cytochrome c, cytochrome b5 mutants, especially the E44A-containing mutants, in the protein complexes suffer greater conformational changes upon reduction than that of the wild type. The variation of these thermodynamic parameters indicates that the strength of mutual interactions between cytochrome c and cytochrome b5 or its mutants has the following order: Cyt c/Cyt b5 > Cyt c/Cyt b5 E56A > Cyt c/Cyt b5 E44A > Cyt c/Cyt b5 E44/56A.

Animals↗
PubMed1998

Newborn and adult recombinant inbred strains: a tool to search for genetic determinants of target organ damage in hypertension.

It has been proposed that one of the primary events in the development of essential hypertension is a growth-related process initiated as early as during fetal development. Differences in kidney size have been observed between most rat models of hypertension and their respective controls. In this study, we analyzed relative kidney size (kidney weight/body wt) in a set of rat recombinant inbred strains (RIS) (N = 27) and their progenitors, the spontaneously hypertensive rat strain (SHR/Ola) and Brown Norway congenic strain (BN.1x), at two different ages, at birth and at 15 weeks. In the progenitors, the relative kidney weight was higher in the hypertensive than in the normotensive strain of both the newborn (P < 0.001) and adult (P < 0.001) animals. In the RIS, a significant correlation was found between the newborn and adult relative kidney weight (r = 0.49, P = 0.01), indicating that the two phenotypes share some of their genetic determinants. A total genome search of newborn and adult relative kidney weight was performed with a total of 453 genetic markers. These analyses revealed several suggestive quantitative trait loci (QTL), some of which were, indeed, significant for both newborn and adult relative kidney weight (such as, D3Mit9 on rat chromosome 3; r = -0.50, P < 0.01; r = -0.47, P < 0.01; respectively). Others, such as the locus on rat chromosome 1 (Rt6; r = -0.43, P < 0.05), were significant only for the adult relative kidney size. This QTL was found in close proximity to a region previously related to susceptibility to hypertensive renal disease in the fawn-hooded rat and, similarly to that study, its effect was found to be independent of blood pressure. Furthermore, a growth pattern of the kidneys after birth, evaluated as the difference between the newborn and adult relative kidney weight, was also subjected to total genome scan. Several suggestive QTL were identified. One of the most significant loci was found at the D1a marker on rat chromosome 17 (r = -0.51, P < 0.01), which was previously related to the determination of adult heart weight in the RIS. In conclusion, the current study demonstrates the usefulness of RIS in studies of hypertension-related phenotypes, some of which are abnormal before the development of high blood pressure. To better understand their role in the pathogenesis of hypertension, studies at different ages are needed, which are uniquely feasible in RIS.

Aging↗
PubMed1997

Alteration of endogenous antioxidant enzymes in naturally occurring hypertrophic cardiomyopathy.

We have recently developed a porcine model with naturally occurring hypertrophic cardiomyopathy (HCM). Similar to humans, occluded intramural coronary artery and damaged mitochondria are frequently observed in these animals in which the disease is thought to be associated with the local ischemia of myocardium. In view of antioxidant functions involved in the ischemic injury, we measured the expression of endogenous antioxidant enzymes in the tissues with and without HCM. The results showed a significant increase of Cu,Zn-superoxide dismutase (SOD), but not Mn-SOD, and decrease of catalase (CAT) activities in the various areas of HCM hearts. It was demonstrated that SOD/CAT ratios in the HCM hearts were significantly higher than those in normals and were found to be dramatically correlated with the severity of cardiac hypertrophy. The altered SOD/CAT ratio was also consistent with increase in lipid damage. We hypothesize that the elevated SOD combined with an inadequate amount of H2O2 scavenging enzyme may lead HCM heart at oxidative stress risk. However, the pathogenic role of imbalanced antioxidant enzyme needs to be further explored.

Animals↗
PubMed1997

Importance of a conserved phenylalanine-35 of cytochrome b5 to the protein's stability and redox potential.

Phenylalanine-35, which is a residue of the hydrophobic patch on the surface of cytochrome b5, has been mutated into Tyr35, His35 and Leu35 to elucidate the functions of the Phe35 and give further insight into the roles of the hydrophobic patch and/or aromatic network. The effects of these mutations on the heme environment, denaturation towards heating and the denaturant urea, redox potential and stability of protein were studied. The relative stability of cytochrome b5 and its mutants towards heating has the order Phe35Tyr > wild type > Phe35Leu > Phe35His in the oxidized state and wild type > Phe35Tyr > Phe35Leu > Phe35His in the reduced state. All the mutants exhibit decreased reduction potentials: Phe35Tyr -66 mV, Phe35His -51 mV and Phe35Leu -28 mV, which are more negative than that of the wild type. The order of redox potential reflects the relative stability in the oxidized and reduced states. A method of producing multiple mutants at a single site of a gene is also described for the first time.

Amino Acid Sequence↗
PubMed1996

Cooperative DNA binding and sequence-selective recognition conferred by the STAT amino-terminal domain.

STAT proteins (signal transducers and activators of transcription) activate distinct target genes despite having similar DNA binding preferences. The transcriptional specificity of STAT proteins was investigated on natural STAT binding sites near the interferon-gamma gene. These sites are arranged in multiple copies and required cooperative interactions for STAT binding. The conserved amino-terminal domain of STAT proteins was required for cooperative DNA binding, although this domain was not essential for dimerization or binding to a single site. Cooperative binding interactions enabled the STAT proteins to recognize variations of the consensus site. These sites can be specific for the different STAT proteins and may function to direct selective transcriptional activation.

Animals↗
PubMed1996

Impaired IL-12 responses and enhanced development of Th2 cells in Stat4-deficient mice.

Interactions between cytokine and receptor lead to the activation of multiple signalling molecules, including the family of signal transducer and activator of transcription (STAT) proteins. STAT4 is one member of this family, and is activated only in response to the cytokine interleukin (IL)-12 (refs 5, 6). By gene targeting, we have generated mice deficient in STAT4 to determine whether the function of this transcription factor is redundant with other signalling molecules activated by IL-12. IL-12-induced increases in the production of interferon (IFN)-gamma cellular proliferation and natural killer (NK) cell cytotoxicity are abrogated in lymphocytes from STAT4-deficient mice. The development of Th1 cells in response to either IL-12 of Listeria monocytogenes is also impaired in the absence of Stat4. Furthermore, Stat4-deficient lymphocytes demonstrate a propensity towards the development of Th2 cells. These results demonstrate that Stat4 is essential for mediating responses to IL-12 in lymphocytes, and regulating the differentiation of both Th1 and Th2 cells.

Animals↗
PubMed1996

The influence of Glu44 and Glu56 of cytochrome b5 on the protein structure and interaction with cytochrome c.

The gene encoding trypsin-solubilized bovine liver microsomal cytochrome b5 (82 residues in length) has been mutated, in which the codons of Glu44 and Glu56 were changed to those of Ala. The mutated genes were expressed in Escherichia coli successfully and three mutant proteins (E44A, E56A and E44/56A) were obtained. The UV-visible, CD and 1H NMR spectra of proteins have been studied. The results show that the mutagenesis at surface residues does not alter the secondary and tertiary structures of cytochrome b5 significantly. The interactions between recombinant cytochrome b5 and its mutants with cytochrome c were studied by using optical difference spectra. The results demonstrated that both Glu44 and Glu56 of cytochrome b5 participate in the formation of a complex between cytochrome b5 and cytochrome c.

Animals↗
PubMed1995

Isolation of two new members of the NF-AT gene family and functional characterization of the NF-AT proteins.

The activation of cytokine genes in response to antigenic stimulation of T cells is mediated by NF-AT proteins. Previous studies have identified two NF-AT proteins, NF-ATp and NF-ATc, that are homologous within a 290 aa domain distantly related to the Rel domain. We have isolated two additional members of this gene family, NF-AT3 and NF-AT4, which encode proteins 65% identical to the other NF-AT proteins within the Rel domain. The four NF-AT genes are transcribed in different sets of tissues that included many sites of expression outside the immune system. The Rel homology domain is sufficient for DNA recognition and cooperative binding interactions with AP-1. Although other members of the Rel family bind DNA as dimers, NF-AT proteins are monomers in solution or bound to DNA. Transfection assays indicate that each of the four NF-AT proteins can activate the IL-2 promoter in T cells.

Amino Acid Sequence↗
PubMed1995

Novel NFAT sites that mediate activation of the interleukin-2 promoter in response to T-cell receptor stimulation.

The transcription factors NFAT and AP-1 have been shown to be essential for inducible interleukin-2 (IL-2) expression in activated T cells. NFAT has been previously reported to bind to two sites in the IL-2 promoter: in association with AP-1 at the distal antigen response element at -280 and at -135. On the basis of DNase I footprinting with recombinant NFAT and AP-1 proteins, gel shift assays, and transfection experiments, we have identified three additional NFAT sites in the IL-2 promoter. Strikingly, all five NFAT sites are essential for the full induction of promoter activity in response to T-cell receptor stimulation. Four of the five NFAT sites are part of composite elements able to bind AP-1 in association with NFAT. These sites display a diverse range of cooperativity and interdependency on NFAT and AP-1 proteins for binding. One of the NFAT sites directly overlaps the CD28-responsive element. We present evidence that CD28 inducibility is conferred by the AP-1 component in NFAT-AP-1 composite elements. These findings provide further insight into the mechanisms involved in the regulation of the IL-2 promoter.

Animals↗
PubMed1995

[Experimental test of the bacteriostatic and bactericidal effect of six disinfectants for Pseudomonas aeruginosa].

Minimal inhibitory concentration (MIC) and minimal bacteriocidal concentration (MBC) of six chosen disinfectants against pseudomonas aeruginosa were tested by the tubular liquid double dilution method. Results showed that 0.06% Chang Kou Jing and 1.5% Supertime are fairly good disinfectants against pseudomonas aeruginosa in infected wound surfaces, postoperative wound dressing, disinfection of surgeons' arms and hands before operation, sterilization of instruments and scrub medical workers' hands after contact with infected patients. The MIC and MBC of the former were 7.5 micrograms/ml and 30.0 micrograms/ml separately, while those of the latter were 11.7 micrograms/ml and 93 micrograms/ml respectively.

Disinfectants↗
PubMed1994

Genes of stress in experimental hypertension.

1. A significant portion of blood pressure variance is modified by the environment. 2. The present report summarizes evidence that: (i) the environmental response is genetically determined; (ii) various stressors can evoke a differential response in hypertensive animals and constitute its intermediate phenotypes; (iii) the response to heat stress can be assigned to a single 'thermosensitivity' locus; (iv) candidate genes of susceptibility to environmental stresses are member(s) of the heat stress gene (HSP) gene families; (v) a restriction fragment length polymorphism of hsp70 and a single base mutation in the 3'-untranslated region of hsp27 are associated with hypertension in recombinant inbred strains. 3. In conclusion, HSP gene variants may be causative in susceptibility to hypertension.

Animals↗
PubMed1993

Novel glucocorticoid receptor complex with DNA element of the hormone-repressed POMC gene.

Previous studies defined a DNA element necessary for glucocorticoid repression of the pro-opiomelanocortin (POMC) gene. The glucocorticoid receptor (GR) binds this negative glucocorticoid response element (nGRE) with an in vitro affinity similar to that of GR for positive GREs. However, whereas GR binds GREs as homodimers, a novel GR complex which forms with nGRE appears to contain three GR molecules. Biochemical characterization of this complex as well as equilibrium binding studies suggest that it is formed by sequential binding of a GR homodimer followed by binding of a GR monomer on the opposite side of the double helix. The DNA-binding domain (DBD) of GR is sufficient for differential binding of GRE and nGRE, as bacterially-expressed DBD formed unique nGRE complexes that contain three GR polypeptides. Thus, the POMC nGRE provides the first example of an interaction between GR and DNA in which GR binds otherwise than as a homodimer. Despite its high affinity for GR, the nGRE differs significantly from GREs in that it does not activate transcription in any context. As the nGRE appears insufficient on its own to confer hormone responsiveness, other POMC promoter elements are likely to be required to mediate glucocorticoid repression.

Animals↗
PubMed1993

Porcine pancreatic islets: isolation, microencapsulation, and xenotransplantation.

To provide a plentiful source of pancreatic islets for future clinical transplants into diabetic patients, we have developed a simple and reliable method to isolate porcine islets of a high degree of purity. Porcine pancreata were perfused and digested with collagenase, and the islets were then purified on dextran density gradients. In order to avoid any damage to the islets, no mechanical devices nor any strenuous treatment was employed. As many as 5 x 10(5) islets were isolated from a single porcine pancreas. Islets were encapsulated in alginate-polylysine-alginate membranes with the aid of an electrostatic droplet generator. In vitro studies demonstrated that the isolated islets secreted insulin in response to glucose and 3-isobutyl-L-methylxanthine (IBMX) challenge for at least 4 weeks. Perifusion studies showed that the kinetics of insulin release from the encapsulated islets was similar to that exhibited by free islets. In in vivo studies, 18 diabetic BALB-c mice were transplanted with 1,500-2,500 encapsulated islets each. In 13 recipients, the diabetic condition was reversed for at least 85 days. When capsules were removed from 2 transplant recipients, their diabetic condition quickly recurred.

Animals↗
PubMed1992

Restriction fragment length polymorphism of hsp70 gene, localized in the RT1 complex, is associated with hypertension in spontaneously hypertensive rats.

Previous studies from our laboratory have demonstrated that the intermediate phenotype of thermosensitivity is present in hypertensive mice and rats. Increased expression of hsp70 caused by increased transcription rate was demonstrated in vivo, in organs, and in cultured cells from spontaneously hypertensive rats and hypertensive mice. In this study, a polymorphism of this gene was revealed with BamHI enzyme by using a human hsp70 probe. A 4.4-kb fragment was visualized in normotensive rats (Brown-Norway BN.lx and Sprague-Dawley), and a 3.0-kb fragment was found in spontaneously hypertensive rats (SHR) of three different origins and in Wistar and Buffalo rats. Both fragments were present in the Wistar-Kyoto rat strain. The present study mapped the polymorphism of hsp70 into the RT1 complex in BN.1K and SHR.1N congenic strains. The hsp70 restriction fragment length polymorphism is associated with a blood pressure difference of 15 mm Hg in recombinant inbred strains. These results justify the search for a mechanism by which hsp70 could influence blood pressure.

Animals↗
PubMed1992

Homodimer formation is rate-limiting for high affinity DNA binding by glucocorticoid receptor.

The glucocorticoid receptor (GR) is a hormone-inducible transcription factor which activates transcription of specific genes by binding to a DNA sequence present in the promoters of inducible genes. These glucocorticoid response elements (GREs) have a conserved palindromic sequence. Each half-GRE palindrome binds one subunit of GR. We have assessed the relative affinity of GR monomers and homodimers for GRE and determined whether homodimer formation is rate-limiting for high affinity GRE binding. The in vitro affinity of GRE binding by GR homodimers was approximately 2 x 10(-10) M, whereas it was approximately 1 nM for GR monomers. While homodimer:GRE complexes were very stable, monomer:GRE complexes appeared less stable in vitro. At low receptor concentration, GR preferentially bound GRE as a homodimer. Prior dilution of GR (equilibrium shifted to monomers) before addition to a GRE binding reaction resulted in slower kinetics of binding by comparison to parallel reactions in which concentrated (largely homodimeric) GR was added first. Taken together, these experiments suggest that homodimer formation is rate-limiting for high affinity GRE binding. A GRE mutant which contained only a half-binding site and which was unable to bind GR homodimers was also unable to confer glucocorticoid-inducible transcription. Taken together with previous work, these experiments support the model that GR homodimers are required for hormone-dependent activation of transcription and that receptor homodimer formation is rate-limiting for GRE binding.

Base Sequence↗
PubMed1992

Ultrastructural study of cerebral arteries in Moyamoya disease.

The cerebral arteries in seven cases of Moyamoya disease, demonstrated by angiography were observed by light, transmission and scanning electron microscopy. In two cases, the lumen of the terminal portion of the internal carotid artery was completely occluded by proliferated connective tissue. The characteristic pathologic change was extensive destruction of the smooth muscle cells and the elastica interna. Degeneration and necrosis of smooth muscle cells were obvious in the media. Many condensed organelles and other elements of cellular destruction were dispersed throughout the interstitium. The media was thinned. The proliferated smooth muscle cells in the intima showed the same destructive changes as in the media. The elastica interna was excessively infolded and was thickened or thinned, disrupted or disintegrated in some segments. Identical, but less severe and more localized changes were seen in three middle cerebral arteries (M2 and M5 segments) and two collateral arteries at the base of brain. These changes suggest that Moyamoya disease is a progressive disorder characterized by repeated destruction of smooth muscle cells occurRing over a long time period. The pathogenesis of these changes is discussed.

Cerebral Arteries↗