Medical Specialties Commons^™

Characterization Of Hsp70 Binding And Nucleotide Exchange By The Yeast Hsp110 Chaperone Sse1, Lance Shaner, Rui Sousa, Kevin A Morano

Journal Articles

SSE1 and SSE2 encode the essential yeast members of the Hsp70-related Hsp110 molecular chaperone family. Both mammalian Hsp110 and the Sse proteins functionally interact with cognate cytosolic Hsp70s as nucleotide exchange factors. We demonstrate here that Sse1 forms high-affinity (Kd approximately 10-8 M) heterodimeric complexes with both yeast Ssa and mammalian Hsp70 chaperones and that binding of ATP to Sse1 is required for binding to Hsp70s. Sse1.Hsp70 heterodimerization confers resistance to exogenously added protease, indicative of conformational changes in Sse1 resulting in a more compact structure. The nucleotide binding domains of both Sse1/2 and the Hsp70s dictate interaction specificity and …

Role Of The Cytoplasmic Domain In Anabaena Sensory Rhodopsin Photocycling: Vectoriality Of Schiff Base Deprotonation, Oleg A Sineshchekov, Elena N Spudich, Vishwa D Trivedi, John L Spudich

Journal Articles

Light-induced electric signals in intact E. coli cells generated by heterologously expressed full-length and C-terminally truncated versions of Anabaena sensory rhodopsin (ASR) demonstrate that the charge movements within the membrane-embedded part of the molecule are stringently controlled by the cytoplasmic domain. In particular, truncation inverts the direction of proton movement during Schiff base deprotonation from outward to cytoplasmic. Truncation also alters faster charge movements that occur before Schiff base deprotonation. Asp(217) as previously shown by FTIR serves as a proton acceptor in the truncated ASR but not in the full-length version, and its mutation to Asn restores the natural outward …

Acute And Chronic Methylphenidate Dose-Response Assessment On Three Adolescent Male Rat Strains, Pamela B Yang, Alan C Swann, Nachum Dafny

Journal Articles

Methylphenidate (MPD), commonly known as Ritalin, is the most frequently prescribed drug to treat children and adults with attention deficit hyperactivity disorder (ADHD). Adolescence is a period of development involving numerous neuroplasticities throughout the central nervous system (CNS). Exposure to a psychostimulant such as MPD during this crucial period of neurodevelopment may cause transient or permanent changes in the CNS. Genetic variability may also influence these differences. Thus, the objective of the present study was to determine whether acute and chronic administration of MPD (0.6, 2.5, or 10.0mg/kg, i.p.) elicit effects among adolescent WKY, SHR, and SD rats and to …

Proteomic Identification Of In Vivo Substrates For Matrix Metalloproteinases 2 And 9 Reveals A Mechanism For Resolution Of Inflammation, Kendra J Greenlee, David B Corry, David A Engler, Risë K Matsunami, Philippe Tessier, Richard G Cook, Zena Werb, Farrah Kheradmand

Journal Articles

Clearance of allergic inflammatory cells from the lung through matrix metalloproteinases (MMPs) is necessary to prevent lethal asphyxiation, but mechanistic insight into this essential homeostatic process is lacking. In this study, we have used a proteomics approach to determine how MMPs promote egression of lung inflammatory cells through the airway. MMP2- and MMP9-dependent cleavage of individual Th2 chemokines modulated their chemotactic activity; however, the net effect of complementing bronchoalveolar lavage fluid of allergen-challenged MMP2(-/-)/MMP9(-/-) mice with active MMP2 and MMP9 was to markedly enhance its overall chemotactic activity. In the bronchoalveolar fluid of MMP2(-/-)/MMP9(-/-) allergic mice, we identified several chemotactic …

Endocarditis And Biofilm-Associated Pili Of Enterococcus Faecalis, Sreedhar R Nallapareddy, Kavindra V Singh, Jouko Sillanpää, Danielle A Garsin, Magnus Höök, Stanley L Erlandsen, Barbara E Murray

Journal Articles

Increasing multidrug resistance in Enterococcus faecalis, a nosocomial opportunist and common cause of bacterial endocarditis, emphasizes the need for alternative therapeutic approaches such as immunotherapy or immunoprophylaxis. In an earlier study, we demonstrated the presence of antibodies in E. faecalis endocarditis patient sera to recombinant forms of 9 E. faecalis cell wall-anchored proteins; of these, we have now characterized an in vivo-expressed locus of 3 genes and an associated sortase gene (encoding sortase C; SrtC). Here, using mutation analyses and complementation, we demonstrated that both the ebp (encoding endocarditis and biofilm-associated pili) operon and srtC are important for biofilm production …

Treatment Evolution In High-Risk Congenital Diaphragmatic Hernia: Ten Years' Experience With Diaphragmatic Agenesis, Kevin P Lally, Pamela A Lally, Krisa P Van Meurs, Desmond J Bohn, Carl F Davis, Bradley Rodgers, Jatinder Bhatia, Golde Dudell

Journal Articles

OBJECTIVE: The objective of this study was to evaluate the impact of newer therapies on the highest risk patients with congenital diaphragmatic hernia (CDH), those with agenesis of the diaphragm.

SUMMARY BACKGROUND DATA: CDH remains a significant cause of neonatal mortality. Many novel therapeutic interventions have been used in these infants. Those children with large defects or agenesis of the diaphragm have the highest mortality and morbidity.

METHODS: Twenty centers from 5 countries collected data prospectively on all liveborn infants with CDH over a 10-year period. The treatment and outcomes in these patients were examined. Patients were followed until death …

Ligand-Signaled Upregulation Of Enterococcus Faecalis Ace Transcription, A Mechanism For Modulating Host-E Faecalis Interaction, Sreedhar R Nallapareddy, Barbara E Murray

Journal Articles

Enterococcus faecalis, the third most frequent cause of bacterial endocarditis, appears to be equipped with diverse surface-associated proteins showing structural-fold similarity to the immunoglobulin-fold family of staphylococcal adhesins. Among the putative E. faecalis surface proteins, the previously characterized adhesin Ace, which shows specific binding to collagen and laminin, was detectable in surface protein preparations only after growth at 46 degrees C, mirroring the finding that adherence was observed in 46 degrees C, but not 37 degrees C, grown E. faecalis cultures. To elucidate the influence of different growth and host parameters on ace expression, we investigated ace expression using E. …

Minimizing Radiation Injury And Neoplastic Effects During Pediatric Fluoroscopy: What Should We Know?, Louis Kenneth Wagner

Journal Articles

Radiation-induced injuries from fluoroscopic procedures in pediatric patients have occurred, and young patients are at greatest risk of many radiation-induced neoplasms. Some fluoroscopists have been injured from their use of fluoroscopy, and they are known to be at risk of radiation-induced neoplasm when radiation is not well-controlled. This article reviews the circumstances that lead to radiation injury and delineates some procedural methods to avoid injury and limit radiation exposure to both the patient and the fluoroscopist.

Role Of A2b Adenosine Receptor Signaling In Adenosine-Dependent Pulmonary Inflammation And Injury, Chun-Xiao Sun, Hongyan Zhong, Amir Mohsenin, Eva Morschl, Janci L Chunn, Jose G Molina, Luiz Belardinelli, Dewan Zeng, Michael R Blackburn

Journal Articles

Adenosine has been implicated in the pathogenesis of chronic lung diseases such as asthma and chronic obstructive pulmonary disease. In vitro studies suggest that activation of the A2B adenosine receptor (A2BAR) results in proinflammatory and profibrotic effects relevant to the progression of lung diseases; however, in vivo data supporting these observations are lacking. Adenosine deaminase-deficient (ADA-deficient) mice develop pulmonary inflammation and injury that are dependent on increased lung adenosine levels. To investigate the role of the A2BAR in vivo, ADA-deficient mice were treated with the selective A2BAR antagonist CVT-6883, and pulmonary inflammation, fibrosis, and airspace integrity were assessed. Untreated and …

Transcriptional Regulation Of The Borrelia Burgdorferi Antigenically Variable Vlse Surface Protein, Tomasz Bykowski, Kelly Babb, Kate Von Lackum, Sean P Riley, Steven J Norris, Brian Stevenson

Journal Articles

The Lyme disease agent Borrelia burgdorferi can persistently infect humans and other animals despite host active immune responses. This is facilitated, in part, by the vls locus, a complex system consisting of the vlsE expression site and an adjacent set of 11 to 15 silent vls cassettes. Segments of nonexpressed cassettes recombine with the vlsE region during infection of mammalian hosts, resulting in combinatorial antigenic variation of the VlsE outer surface protein. We now demonstrate that synthesis of VlsE is regulated during the natural mammal-tick infectious cycle, being activated in mammals but repressed during tick colonization. Examination of cultured B. …

Mutations Affecting Beta-Tubulin Folding And Degradation, Yaqing Wang, Guoling Tian, Nicholas J Cowan, Fernando Cabral

Journal Articles

Revertants of a colcemid-resistant Chinese hamster ovary cell line with an altered (D45Y) beta-tubulin have allowed the identification of four cis-acting mutations (L187R, Y398C, a 12-amino acid in-frame deletion, and a C-terminal truncation) that act by destabilizing the mutant tubulin and preventing it from incorporating into microtubules. These unstable beta-tubulins fail to form heterodimers and are predominantly found in association with the chaperonin CCT, suggesting that they cannot undergo productive folding. In agreement with these in vivo observations, we show that the defective beta-tubulins do not stably interact with cofactors involved in the tubulin folding pathway and, hence, fail to …

Comparison Of Og1rf And An Isogenic Fsrb Deletion Mutant By Transcriptional Analysis: The Fsr System Of Enterococcus Faecalis Is More Than The Activator Of Gelatinase And Serine Protease, Agathe Bourgogne, Susan G Hilsenbeck, Gary M Dunny, Barbara E Murray

Journal Articles

The FsrABC system of Enterococcus faecalis controls the expression of gelatinase and a serine protease via a quorum-sensing mechanism, and recent studies suggest that the Fsr system may also regulate other genes important for virulence. To investigate the possibility that Fsr influences the expression of additional genes, we used transcriptional profiling, with microarrays based on the E. faecalis strain V583 sequence, to compare the E. faecalis strain OG1RF with its isogenic mutant, TX5266, an fsrB deletion mutant. We found that the presence of an intact fsrB influences expression of numerous genes throughout the growth phases tested, namely, late log to …

Hepatic And Renal Cytochrome P450 Gene Regulation During Citrobacter Rodentium Infection In Wild-Type And Toll-Like Receptor 4 Mutant Mice, Terrilyn A Richardson, Melanie Sherman, Leposava Antonovic, Sean S Kardar, Henry W Strobel, Daniel Kalman, Edward T Morgan

Journal Articles

Citrobacter rodentium is the rodent equivalent of human enteropathogenic Escherichia coli infection. This study investigated regulation of hepatic and renal cytochrome P450 (P450) mRNAs, hepatic P450 proteins, cytokines, and acute phase proteins during C. rodentium infection. Female C3H/HeOuJ (HeOu) and C3H/HeJ (HeJ) mice [which lack functional toll-like receptor 4 (TLR4)] were infected with C. rodentium by oral gavage and sacrificed 6 days later. Hepatic CYP4A10 and 4A14 mRNAs were decreased in HeOu mice (

Translational Regulation Of Nuclear Gene Cox4 Expression By Mitochondrial Content Of Phosphatidylglycerol And Cardiolipin In Saccharomyces Cerevisiae, Xuefeng Su, William Dowhan

Journal Articles

Previous results indicated that translation of four mitochondrion-encoded genes and one nucleus-encoded gene (COX4) is repressed in mutants (pgs1Delta) of Saccharomyces cerevisiae lacking phosphatidylglycerol and cardiolipin. COX4 translation was studied here using a mitochondrially targeted green fluorescence protein (mtGFP) fused to the COX4 promoter and its 5' and 3' untranslated regions (UTRs). Lack of mtGFP expression independent of carbon source and strain background was established to be at the translational level. The translational defect was not due to deficiency of mitochondrial respiratory function but was rather caused directly by the lack of phosphatidylglycerol and cardiolipin in mitochondrial membranes. Reintroduction of …

Functional Analysis Of The Amine Substrate Specificity Domain Of Pepper Tyramine And Serotonin N-Hydroxycinnamoyltransferases, Sei Kang, Kiyoon Kang, Gap Chae Chung, Doil Choi, Atsushi Ishihara, Dong-Sun Lee, Kyoungwhan Back

Journal Articles

Pepper (Capsicum annuum) serotonin N-hydroxycinnamoyltransferase (SHT) catalyzes the synthesis of N-hydroxycinnamic acid amides of serotonin, including feruloylserotonin and p-coumaroylserotonin. To elucidate the domain or the key amino acid that determines the amine substrate specificity, we isolated a tyramine N-hydroxycinnamoyltransferase (THT) gene from pepper. Purified recombinant THT protein catalyzed the synthesis of N-hydroxycinnamic acid amides of tyramine, including feruloyltyramine and p-coumaroyltyramine, but did not accept serotonin as a substrate. Both the SHT and THT mRNAs were found to be expressed constitutively in all pepper organs. Pepper SHT and THT, which have primary sequences that are 78% identical, were used as models …

The Dynamic Proteome Of Lyme Disease Borrelia, Steven J Norris

Journal Articles

The proteome of the spirochete bacterium Borrelia burgdorferi, the tick-borne agent of Lyme disease, has been characterized by two different approaches using mass spectrometry, providing a launching point for future studies on the dramatic changes in protein expression that occur during transmission of the bacterium between ticks and mammals.

Construction Of Improved Temperature-Sensitive And Mobilizable Vectors And Their Use For Constructing Mutations In The Adhesin-Encoding Acm Gene Of Poorly Transformable Clinical Enterococcus Faecium Strains, Sreedhar R Nallapareddy, Kavindra V Singh, Barbara E Murray

Journal Articles

Inactivation by allelic exchange in clinical isolates of the emerging nosocomial pathogen Enterococcus faecium has been hindered by lack of efficient tools, and, in this study, transformation of clinical isolates was found to be particularly problematic. For this reason, a vector for allelic replacement (pTEX5500ts) was constructed that includes (i) the pWV01-based gram-positive repAts replication region, which is known to confer a high degree of temperature intolerance, (ii) Escherichia coli oriR from pUC18, (iii) two extended multiple-cloning sites located upstream and downstream of one of the marker genes for efficient cloning of flanking regions for double-crossover mutagenesis, (iv) transcriptional terminator …

Chronic Exposure To Mdma (Ecstasy) Elicits Behavioral Sensitization In Rats But Fails To Induce Cross-Sensitization To Other Psychostimulants, Gunjan M Modi, Pamela B Yang, Alan C Swann, Nachum Dafny

Journal Articles

BACKGROUND: The recreational use of 3,4-methylenedioxymethamphetamine (MDMA, ecstasy) among adolescents and young adults has become increasingly prevalent in recent years. While evidence suggests that the long-term consequences of MDMA use include neurodegeneration to serotonergic and, possibly, dopaminergic pathways, little is known about susceptibility, such as behavioral sensitization, to MDMA.

METHODS: The objectives of this study were to examine the dose-response characteristics of acute and chronic MDMA administration in rats and to determine whether MDMA elicits behavioral sensitization and whether it cross-sensitizes with amphetamine and methylphenidate. Adult male Sprague-Dawley rats were randomly divided into three MDMA dosage groups (2.5 mg/kg, 5.0 …

Dose-Response Characteristics Of Methylphenidate On Locomotor Behavior And On Sensory Evoked Potentials Recorded From The Vta, Nac, And Pfc In Freely Behaving Rats, Pamela B Yang, Alan C Swann, Nachum Dafny

Journal Articles

BACKGROUND: Methylphenidate (MPD) is a psychostimulant commonly prescribed for attention deficit/hyperactivity disorder. The mode of action of the brain circuitry responsible for initiating the animals' behavior in response to psychostimulants is not well understood. There is some evidence that psychostimulants activate the ventral tegmental area (VTA), nucleus accumbens (NAc), and prefrontal cortex (PFC).

METHODS: The present study was designed to investigate the acute dose-response of MPD (0.6, 2.5, and 10.0 mg/kg) on locomotor behavior and sensory evoked potentials recorded from the VTA, NAc, and PFC in freely behaving rats previously implanted with permanent electrodes. For locomotor behavior, adult male Wistar-Kyoto …