Biomedical subjects

F Zhang

Publications and source records attributed to F Zhang.

At least 397 records · Page 22Linked to original sources

PubMed1997

Cyclo-oxygenase-2 gene expression in neurons contributes to ischemic brain damage.

Cyclo-oxygenase-2 (COX-2), a rate-limiting enzyme for prostanoid synthesis, is induced during inflammation and participates in inflammation-mediated cytotoxicity. Cerebral ischemia is followed by an inflammatory reaction that plays a role in the evolution of the tissue damage. We studied whether COX-2 is induced after cerebral ischemia and if so, whether such expression contributes to ischemic brain damage. The middle cerebral artery was occluded in rats, and the ischemic area was sampled for analysis 3-96 hr later. COX-2 mRNA was determined by the competitive reverse-transcription PCR. COX-2 mRNA was upregulated in the ischemic hemisphere, but not contralaterally, beginning 6 hr after ischemia. The upregulation reached a maximum at 12 hr, at which time a fivefold induction of the message occurred. Twenty-four hours after ischemia, the concentration of prostaglandin E2 was elevated in the injured brain by 292 +/- 57% (n = 6). COX-2 immunoreactivity was observed in neurons at the medial edge of the ischemic area. Administration of the COX-2 inhibitor NS-398 attenuated the elevation in prostaglandin E2 in the postischemic brain and reduced the volume of the infarct by 29 +/- 6% (p < 0.05). Thus, cerebral ischemia leads to upregulation of COX-2 message, protein, and reaction products in the injured hemisphere. The data implicate COX-2 in the mechanisms of delayed neuronal death at the infarct border and provide the rationale for neuroprotective strategies employing COX-2 inhibitors.

Animals↗

PubMed1997

Antiapoptotic compound to enhance hypothermic liver preservation.

BACKGROUND: Apoptosis (programmed cell death) occurs as a consequence of global organ ischemia during isolation and storage prior to transplantation. If apoptosis is inhibited during ischemia, organ preservation should be improved, and the length of time for permissible storage may be increased. The objective of this study was to test the effect of a newly developed antiapoptotic compound, LXR-015, during extended hypothermic liver preservation. METHODS: Three groups of 12 rats each were studied. In the normal group, liver function was studied immediately after harvesting. In the study group, harvested livers were flushed with Euro-Collins solution (30 ml/kg body weight) containing LXR-015 at a concentration equivalent to 9 mg/kg animal body weight (300 microg/ml). The livers were then stored at 4 degrees C for 24 hr before liver function was studied. In the control group, harvested livers were flushed with Euro-Collins solution without LXR-015 and then stored at 4 degrees C for 24 hr before liver function was studied. RESULTS: Portal venous flow was higher (P<0.05) in the normal and study groups compared with the control group. Portal venous resistance was lower (P<0.05) in the normal and study groups compared with the control group. Liver tissue oxygen consumption in the study group was significantly higher than in both the normal and control groups (P<0.05). Liver enzyme production (aspartate aminotransferase, alanine aminotransferase, lactate dehydrogenase, creatine kinase) was higher in the control group than in either the study or normal group (P<0.05). Bile production in both the normal and study groups was higher than in the control group (P<0.05). The liver tissue wet to dry weight ratio in both the normal and study groups was lower than in the control group (P<0.05). Histopathology studies revealed fewer apoptotic bodies (P<0.05) in both the normal (1.70+/-0.15 per high-power field) and study groups (2.08+/-0.10 per high-power field) than in the control group (7.92+/-.33 per high-power field). CONCLUSIONS: Adding an antiapoptotic compound, LXR-015, to Euro-Collins solution significantly improves hypothermic preservation of the rat liver compared with Euro-Collins solution alone.

Alanine Transaminase↗

PubMed1997

Role of transforming growth factor-beta in long-term synaptic facilitation in Aplysia.

The role of transforming growth factor-beta (TGF-beta) in long-term synaptic facilitation was examined in isolated Aplysia ganglia. Treatment with TGF-beta1 induced long-term facilitation (24 and 48 hours), but not short-term (5 to 15 minutes) or intermediate-term (2 to 4 hours) facilitation. The long-term effects of TGF-beta1 were not additive with those of serotonin. Moreover, serotonin-induced facilitation was blocked by an inhibitor of TGF-beta. Thus, activation of TGF-beta may be part of the cascade of events underlying long-term sensitization, consistent with the hypothesis that signaling molecules that participate in development also have roles in adult neuronal plasticity.

Animals↗

PubMed1997

Cloning, and molecular characterization of the GCV1 gene encoding the glycine cleavage T-protein from Saccharomyces cerevisiae.

We have isolated the gene encoding the glycine cleavage T-protein (GCV1) of the yeast Saccharomyces cerevisiae and shown through gene disruption and enzyme assays that inactivation of GCV1 destroys glycine cleavage function. A DNA fragment encoding the GCV1 gene was cloned by PCR amplification using degenerate oligodeoxyribonucleotides, and the cloned fragment was used as a probe to isolate the complete gene from a yeast genomic library. Growth with glycine stimulated expression of the GCV1 gene as determined by Northern analysis and increased the beta-galactosidase activity of a GCV1-lacZ fusion 30-fold. The URA3 gene was inserted into the coding sequence of GCV1 and the resulting construct was used to disrupt the chromosomal GCV1 gene in a diploid strain of yeast. gcv1::URA3 haploid derivatives grew normally or only slightly more slowly than the isogenic wild-type haploids. All gcv1 strains studied were unable to grow on glycine as a sole nitrogen source and lacked glycine cleavage enzyme activity. Growth of shm1 shm2 mutants was stimulated by glycine, whereas glycine could not supplement the growth of the isogenic gcv1 strain.

Amino Acid Oxidoreductases↗

PubMed1997

Regulation of ciprofloxacin uptake in human promyelocytic leukemia cells and polymorphonuclear leukocytes.

Polymorphonuclear leukocytes (PMNs) actively internalize ciprofloxacin, a capability that can enhance killing of intracellular bacteria and facilitate delivery of the antimicrobial agent to infection sites by migrating PMNs. In this study we investigated mechanisms for up-regulation of this process. Activation with N-formyl-methionyl-leucyl-phenylalanine (fMLP; 100 nM) enhanced PMN ciprofloxacin uptake by 50% (P < 0.05). Phorbol myristate acetate (PMA; > or = 10 nM) enhanced uptake by at least 36-fold, mainly by stimulating an increase in the Vmax of the ciprofloxacin transporter. This effect of PMA was inhibited by antagonists of protein kinase C (H7 and chelerythrine) and the mitogen-activated protein kinase cascade downstream (PD 098059). Under resting and PMA-activated conditions, ciprofloxacin uptake by immature human promyelocytic leukemia (HL-60) cells was much lower than in PMNs. However, when HL-60 cells were induced to mature into PMN-like cells, their ciprofloxacin uptake activity increased markedly. These findings implicate a role for protein kinase C in up-regulation of the ciprofloxacin transporter and suggest that myeloid cells acquire an enhanced ability to take up ciprofloxacin as they mature to end-stage PMNs.

Anti-Infective Agents↗

PubMed1997

Arterialization of the venous system in a rat lower limb model.

Creation of alternative vascular conduits has been one of the most challenging subjects in the history of vascular surgery. In the past, afferent arteriovenous fistulas have been used by vascular surgeons to rescue ischaemic extremities. We have undertaken a comprehensive investigation to characterize arterialization of the venous system at different levels of the rat lower limb. In Group 1 (n = 28), we performed an end-to-end anastomosis between the proximal saphenous artery and the distal saphenous vein (Superficial venous system), and ligated all other branches of the femoral artery. In Group 2 (n = 28), we performed an end-to-end anastomosis between the proximal saphenous artery and the distal femoral vein (Deep venous system). In Group 3 (n = 24), the control group, the femoral artery was ligated proximal to its trifurcation into the saphenous, epigastric, and distal femoral vessels. Group 4 (n = 10), the normal group, underwent no surgery. Animals were sacrificed at the first, third, and eighth postoperative weeks. Arteriographic, India ink injection, latex injection, and histological studies were performed on all groups. Successful arterialization of veins, significant neovascularization, and less ischaemic injury of muscles were noted in the arterialized vein groups (Groups 1 and 2) when compared with the ligated group (Group 3). This model appears to depict successfully arterialization of the lower limb venous system in a small animal model.

Angiography↗

PubMed1997

Non-linear effects in dose-time analyses: application of modern statistical techniques.

BACKGROUND AND PURPOSE: The non-linear effect of overall treatment time and total dose on the outcome of fractionated head and neck radiotherapy is of interest. This and other non-linear effects can be investigated using recently developed statistical techniques. This article provides an illustration of the ability of these statistical methods. MATERIALS AND METHODS: Recently developed statistical methods (Hastie, T.J. and Tibshirani, R.J. Generalised Additive Models. Chapman-Hall, London, 1990), called generalized additive models, are applied to data from the patterns of fractionation study (Withers, H.R., Peters, L.J., Taylor, J.M.G., Owen, J.B., Morrison, W.H., Schultheiss, T.E., Keane, T., O'Sullivan, B., van Dyk, J., Gupta, N., Wang, C.C., Jones, C.U., Doppke, K.P., Myint, S., Thompson, M., Parsons, J.T., Mendenhall, W.M., Dische, S., Aird, E.G.A., Henk, J.M., Bidmead, M.A.M., Svoboda, V., Chon, Y., Hanlon, A.L., Peters, T.L. and Hanks, G.E. Local control of carcinoma of the tonsil by radiation therapy: an analysis of patterns of fractionation in nine institutions. Int. J. Radiat. Oncol. Biol. Phys. 33: 549-562, 1995) of tonsil cancer. These techniques enable one to develop models which more accurately represent the relationship between multivariate predictor variables and the outcome variable in a regression analysis. These data driven methods allow the effect of each predictor variable on the outcome to be non-linear and estimated from the data. This is achieved by replacing the standard linear model combination of predictor variables, such as 'alpha0 + alpha1dose + alpha2time' by 'S1(dose) + S2(time)', where S1 and S2 are smooth non-linear functions of dose and time, respectively, which are estimated from the data. RESULTS: In the pattern of fractionation study these methods indicate that the effect of total dose on the probability of local recurrence is linear, but there is a suggestion that the effect of overall treatment time is non-linear. There is no effect of dose and time on the latency time to recurrence of those who do recur, but there is a weak suggestion of a non-linear effect of patients age, with younger patients recurring earlier. CONCLUSIONS: Generalized additive models provide a flexible and powerful means of exploring non-linear effects in experimental data.

Adult↗

PubMed1997

Taurine is an osmolyte in rat liver macrophages (Kupffer cells).

BACKGROUND/AIMS: The availability of betaine as an osmolyte was recently shown to interfere strongly with important cell functions of liver macrophages (Kupffer cells), such as eicosanoid and tumor necrosis factor-alpha production or phagocytosis. We therefore investigated whether taurine is also used as an osmolyte by Kupffer cells and whether it is involved in the control of Kupffer cell functions. METHODS/RESULTS: Hyperosmotic (hypoosmotic) exposure of cultured rat liver macrophages (Kupffer cells) for 6-12 h led to an increase (decrease) in the mRNA levels of the taurine transporter (TAUT) and an increase (decrease) in taurine transport into the cells. The hyperosmolarity-induced increase in TAUT-mRNA levels was diminished by 37+/-10% upon addition of taurine, but not upon addition of betaine. When Kupffer cells were preloaded with taurine, hypoosmotic exposure led to a rapid efflux of taurine from the cells, which was significantly delayed in the presence of the anion exchanger inhibitor 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS). Taurine efflux was also stimulated during phagocytosis of Latex particles; however, Latex was without effect on the hyperosmolarity-induced increase of TAUT mRNA levels. Lipopolysaccharide (LPS) led to an induction of cyclooxygenase-2, which was markedly enhanced during hyperosmotic conditions. Taurine diminished the induction of cyclooxygenase-2 and inhibited the LPS/hyperosmolarity-induced stimulation of prostaglandin E2 formation. CONCLUSIONS: The data suggest that, in addition to betaine, taurine also acts as an osmolyte in Kupffer cells, and that taurine availability may be an important modulater of Kupffer cell functions such as eicosanoid synthesis.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗

PubMed1997

Elements both 5' and 3' to the murine Hoxd4 gene establish anterior borders of expression in mesoderm and neurectoderm.

In this report, we show that a lacZ reporter spanning 12.5 kb of murine Hoxd4 genomic DNA contains the major regulatory elements controlling Hoxd4 expression in the mouse embryo. Mutational analysis revealed multiple regulatory regions both 5' and 3' to the coding region. These include a 3' enhancer region required for expression in the central nervous system (CNS) and setting the anterior border in the paraxial mesoderm, and a 5' mesodermal enhancer that directs expression in paraxial and lateral plate mesoderm. A previously defined retinoic acid response element (RARE) is a component of the 5' mesodermal enhancer. Our results support a model in which retinoic acid receptors (RARs) and HOX proteins mediate the initiation and maintenance of Hoxd4 expression.

Animals↗

PubMed1997

Environmental and occupational determinants of blood pressure in rural communities in China.

PURPOSE: To identify and characterize major environmental and occupational determinants of blood pressure in rural communities in China. METHODS: In 1993 we conducted a large cross-sectional, community-based study of 20,216 residents aged 15 years or older, from the Yijing area of Anhui Province (8022 men, 12,194 women), one of whom were receiving treatment for hypertension. The mean systolic blood pressure was 116.7 +/- 19.5 mmHg for men and 113.2 +/- 19.4 mmHg for women. RESULTS: The mean diastolic blood pressure was 72.4 +/- 12.1 mmHg for men and 70.4 +/- 11.6 mmHg for women. Age and body mass index were the two most important determinants of blood pressure in this population. With controls for age and body mass index, height and weight remained significant predictors of blood pressure. Multiple linear regression analysis indicated that alcohol consumption, self-reported exposure to noise, drinking of tap water and pond water, occupational exposure to dust/fumes/gases, rice consumption, inferior housing, household crowdedness, and being unmarried were related to increased blood pressure levels. Vegetable intake, frequent consumption of meat at meals, high level of physical activity, exposure to straw-combustion smoke, and pesticide use were negatively associated with blood pressure. CONCLUSIONS: Our study demonstrated that a broad array of demographic, ergonomic, nutritional, and environmental factors are critical determinants of blood pressure in this rural Chinese population.

Adolescent↗

PubMed1997

Oxidation of dopamine in the presence of cysteine: characterization of new toxic products.

Previous studies demonstrated that oxidation of dopamine (DA) in the presence of L-cysteine (CySH) at pH 7.4 gives a complex mixture of cysteinyl conjugates of the neurotransmitter that can be easily further oxidized to a number of dihydrobenzothiazines (DHBTs) along with unidentified yellow products. In this investigation, three of these products have been identified. 7-(2 Aimoethyl)-5-hydroxy-1,4-benzothiazine-3-carboxylic acid (BT-1) is formed as a result of oxidation of 5-S-cysteinyldopamine (5-S-CyS-DA) and 7-(2-aminoethyl)-3,4-dihydro-5-hydroxy-2H-1,4-benzothiazine-3-carboxylic acid (DHBT-1). Regioisomers 6-(2-aminoethyl)-1,8,9,10-tetrahydrobenzo[1,2-b:4,3-b']bis[1,4] thiazine-9-carboxylic acid (12) and 6-(2-aminoethyl)-1,2,3,10-tetrahydrobenzo[1,2-b:4,3-b']bis[1,4] thiazine-2-carboxylic acid (13) are formed by oxidation of 2,5-bi-S-CyS-DA), 6-S-cysteinyl-7-(2-aminoethyl)-3-4-dihydro-5-hydroxy-2H-1, 4-benzothiazine-3-carboxylic acid (DHBT-2) and 6-S-cysteinyl-8(2-aminoethyl)-3,4-dihydro-2H-1,4-benzothiazine-3-carboxy lic acid (DHBT-6). 2,5-Bi-S-CyS-DA, DHBT-2, and DHBT-6 are major early products of DA oxidation in the presence of CySH. However, because these three compounds are the most easily oxidized products formed in this reaction, they are subsequently transformed into 12 and 13, the latter regioisomer always being the major product. Both 12 (LD50 = 18.5 micrograms) and 13 (LD50 = 1.5 microgram) are lethal when administered into the brains of mice and evoke hyperactivity and tremor. The potential relevance of the in vitro chemistry described in this and earlier reports to reaction that might occur in neuromelanin-pigmented dopaminergic neurons in Parkinson's disease is discussed.

Animals↗

PubMed1997

Solution structure of human intestinal fatty acid binding protein: implications for ligand entry and exit.

The human intestinal fatty acid binding protein (I-FABP) is a small (131 amino acids) protein which binds dietary long-chain fatty acids in the cytosol of enterocytes. Recently, an alanine to threonine substitution at position 54 in I-FABP has been identified which affects fatty acid binding and transport, and is associated with the development of insulin resistance in several populations including Mexican-Americans and Pima Indians. To investigate the molecular basis of the binding properties of I-FABP, the 3D solution structure of the more common form of human I-FABP (Ala54) was studied by multidimensional NMR spectroscopy. Recombinant I-FABP was expressed from E. coli in the presence and absence of 15N-enriched media. The sequential assignments for non-delipidated I-FABP were completed by using 2D homonuclear spectra (COSY, TOCSY and NOESY) and 3D heteronuclear spectra (NOESY-HMQC and TOCSY-HMQC). The tertiary structure of human I-FABP was calculated by using the distance geometry program DIANA based on 2519 distance constraints obtained from the NMR data. Subsequent energy minimization was carried out by using the program SYBYL in the presence of distance constraints. The conformation of human I-FABP consists of 10 antiparallel beta-strands which form two nearly orthogonal beta-sheets of five strands each, and two short alpha-helices that connect the beta-strands A and B. The interior of the protein consists of a water-filled cavity between the two beta-sheets. The NMR solution structure of human I-FABP is similar to the crystal structure of rat I-FABP. The NMR results show significant conformational variability of certain backbone segments around the postulated portal region for the entry and exit of fatty acid ligand.

Amino Acid Sequence↗

PubMed1997

Blood-flow velocity as a factor in postoperative microvascular patency.

The authors attempted to develop a reliable and reproducible new animal model in which the blood-flow velocity to a flap could be varied. This model was utilized to study the effects of different blood-flow velocities on the patency rate of small 1- to 2-mm vessels after common microsurgical procedures. Male Sprague-Dawley rats, weighing 450 to 550 gm, were used to develop a model creating either a "high blood flow" or a "low blood flow" state by ligating the rat femoral artery, either distally or proximally, to an epigastric artery based on a groin cutaneous flap. Blood-flow velocities were measured by microvascular flowmeter, and statistical analysis was performed on the data collected. The model was next used to determine the effects of different blood-flow velocities on the patency rates of rat femoral vessels after primary anastomosis vs interpositional vein grafting. Interpositional vein grafting was subsequently repeated by a more senior microsurgeon, to determine the potential effects of increased surgical experience. The animal model was reliable, easily reproducible, and efficacious in producing two separate groups of rats with significantly different blood-flow velocities (3.98 vs. 2.14 +/- 0.5 ml/min), as was confirmed by electromagnetic flowmeter and statistical analysis. In experienced hands, decreased blood-flow velocity did not result in decreased patency rates of these small vessels after primary anastomosis, or even after vein grafting. As long as microvascular vein grafting and primary anastomosis procedures are done properly, even 1-mm vessels can tolerate significantly decreased blood-flow velocity without a decreased patency rate. Although many known factors can contribute to thrombosis and failure of anastomoses in clinical microsurgery, blood-flow velocity appears not to be a significant factor. Also described is a new, reliable animal model that can be used in small-vessel blood-flow velocity studies.

Anastomosis, Surgical↗

PubMed1997

Comparison of muscle mass preservation in denervated muscle and transplanted muscle flaps after motor and sensory reinnervation and neurotization.

The gracilis muscle model was used either as a denervated muscle in situ or as a transplanted flap in 273 rats to compare the trophic effects of muscle reinnervation and neurotization using sensory and motor nerves. The average gracilis muscle flap weighed 626 +/- 94 mg at the time of the initial procedure. Experimental muscles were examined 6 months following the procedure. In denervated, nontransplanted muscles, both motor nerve reinnervation and neurotization resulted in significantly preserved muscle mass, averaging 570 +/- 69 and 521 +/- 116 mg, respectively, compared with the denervated control average of 178 +/- 22 mg (p < 0.05). Sensory nerve reinnervation and neurotization produced much smaller trophic effects (p > 0.05). In transplanted gracilis free flaps, however, only direct reinnervation with motor or sensory nerves resulted in improved bulk preservation, with average weights of 313 +/- 83 and 327 +/- 91 mg compared with the control average of 201 +/- 76 mg (p < 0.05). Neither sensory nor motor neurotization was significantly effective in the free-flap model (p > 0.05). These data suggest that transplantation may alter the response of muscle to reinnervation.

Animals↗

PubMed1997

A defective signal peptide tethers the floury-2 zein to the endoplasmic reticulum membrane.

The maize (Zea mays L.) floury-2 (fl2) mutation is associated with a general decrease in storage protein synthesis, altered protein body morphology, and the synthesis of a novel 24-kD alpha-zein storage protein. Unlike storage proteins in normal kernels and the majority of storage proteins in fl2 kernels, the 24-kD alpha-zein contains a signal peptide that would normally be removed during protein synthesis and processing. The expected processing site of this alpha-zein reveals a putative mutation alanine-->valine (Ala-->Val) that is not found at other junctions between signal sequences and mature proteins. To investigate the impact of such a mutation on signal peptide cleavage, we have assayed the 24-kD fl2 alpha-zein in a co-translational processing system in vitro. Translation of RNA from fl2 kernels or synthetic RNA encoding the fl2 alpha-zein in the presence of microsomes yielded a 24-kD polypeptide. A normal signal peptide sequence, generated by site-directed mutagenesis, restored the capacity of the RNA to direct synthesis of a properly processed protein in a cell-free system. Both the fl2 alpha-zein and the fl2 alpha-zein (Val-->Ala) were translocated into the lumen of the endoplasmic reticulum. The processed fl2 alpha-zein (Val-->Ala) was localized in the soluble portion of the microsomes, whereas the fl2 alpha-zein co-fractionated with the microsomal membranes. By remaining anchored to protein body membranes during endosperm maturation, the fl2 zein may thus constrain storage protein packing and perturb protein body morphology.

Animals↗

PubMed1997

Susceptibility of nude mice carrying the Fv-4 gene to Friend murine leukemia virus infection.

Fv-4 is a mouse gene that dominantly confers resistance to infection with Friend murine leukemia virus (F-MuLV) (S. Suzuki, Jpn. J. Exp. Med. 45:473-478, 1975). Despite complete resistance to ecotropic MuLV infection in mice carrying the Fv-4 gene, it is known that cells carrying the resistance gene in tissue culture do not always show resistance as extensive as that in vivo (H. Yoshikura and T. Odaka, JNCI 61:461-463, 1978). To investigate the immunological effect on resistance in vivo, we introduced the Fv-4 gene into BALB/c nude mice (Fv-4-/- nude[nu/nu]) by mating them with Fv-4 congenic BALB/c mice (Fv-4r/r nude+/+) and examined the susceptibility of the F2 progeny to F-MuLV. All BALB/c nude mice without the Fv-4 gene (Fv-4-/- nude[nu/nu]) were permissive to F-MuLV and developed erythroleukemia within 2 weeks after virus inoculation. The BALB/c nude mice with the Fv-4 gene (Fv-4r/r nude[nu/nu]) did not develop leukemia, and no or little virus was detected in the spleen 7 weeks after virus inoculation. The resistance to F-MuLV was dominant in (Fv-4 congenic BALB/c x BALB/c nude) F1 mice with the Fv-4r/- nude(nu/+) genotype as strictly as in (Fv-4 congenic BALB/c x BALB/c) F1 mice with the Fv-4r/- nude+/+ genotype. However, almost all BALB/c nude mice with the Fv-4r/- nude(nu/nu) genotype developed the disease within 7 weeks, and the virus was detected in all of their spleens even in the mice without leukemia. These results show that the resistance caused by the Fv-4 gene is recessive in nude mice and dominant in BALB/c mice. Some immunological effects, perhaps cell-mediated immunity, may play important roles in the resistance to F-MuLV infection in vivo in addition to the dosage effect of the Fv-4 product.

Animals↗

PubMed1997

Evidence that complex formation by Bas1p and Bas2p (Pho2p) unmasks the activation function of Bas1p in an adenine-repressible step of ADE gene transcription.

Bas1p and Bas2p (Pho2p) are Myb-related and homeodomain DNA binding proteins, respectively, required for transcription of adenine biosynthetic genes in Saccharomyces cerevisiae. The repression of ADE genes in adenine-replete cells involves down-regulation of the functions of one or both of these activator proteins. A LexA-Bas2p fusion protein was found to activate transcription from a lexAop-lacZ reporter independently of both BAS1 function and the adenine levels in the medium. In contrast, a LexA-Bas1p fusion activated the lexAop reporter in a BAS2-dependent and adenine-regulated fashion. The DNA binding activity of Bas2p was not needed for its ability to support activation of the lexAop reporter by LexA-Bas1p, indicating that LexA-Bas1p recruits Bas2p to this promoter. The activation functions of both authentic Bas1p and LexA-Bas1p were stimulated under adenine-repressing conditions by overexpression of Bas2p, suggesting that complex formation by these proteins is inhibited in adenine-replete cells. Replacement of Asp-617 with Asn in Bas1p or LexA-Bas1p allowed either protein to activate transcription under repressing conditions in a manner fully dependent on Bas2p, suggesting that this mutation reduces the negative effect of adenine on complex formation by Bas1p and Bas2p. Deletions of N-terminal and C-terminal segments from the Bas1p moiety of LexA-Bas1p allowed high-level activation by the truncated proteins independently of Bas2p and adenine levels in the medium. From these results we propose that complex formation between Bas1p and Bas2p unmasks a latent activation function in Bas1p as a critical adenine-regulated step in transcription of the ADE genes.

Adenine↗

PubMed1997

IGF-I regulation of Na(+)-K(+)-ATPase in rat arterial smooth muscle.

Insulin-like growth factor I (IGF-I) is vasodilatory and mitogenic for vascular smooth muscle cells (VSMC). Alteration in VSMC Na(+)-K(+)-adenosinetriphosphatase (Na(+)-K(+)-ATPase) activity is hypothesized to underlie abnormal vascular tone and growth in hypertension and diabetes. Therefore, we investigated effects of IGF-I on Na(+)-K(+)-ATPase activity in rat aortic VSMC. IGF-I increases pump activity in a dose- and time-dependent manner: the minimal dose required was 10(-10) M, and the minimal time required was 20 min (at 10(-8) M) to increase activity. Similar effects persisted through 12 h. In Na(+)-loaded cells, IGF-I does not further stimulate activity. Blockade of Na+/H+ exchange attenuates IGF-I-induced increases in activity after 30 min but has no effect after 12 h. Northern blot analyses reveal that expression of the alpha 1- and the alpha 2-subunits of the pump were unaffected by IGF-I. Plasma membrane alpha 1- and alpha 2-protein were also unaffected, suggesting translocation of preformed pools was not responsible for the increases. Inhibitors revealed that neither tyrosine kinase activity, RNA transcription, protein synthesis, nitric oxide synthase activity, or protein kinase C activity mediated this IGF-I effect. Therefore, IGF-I regulates Na pump activity in the short term by an Na+/H+ exchange-dependent but transcription/translocation-independent mechanism. These data suggest that IGF-I, known to be produced by VSMC, may regulate tone and growth responses abnormal in disease states such as hypertension and diabetes.

Animals↗