bluuebunny/crossref_metadata_2025 huggingface dataset:
Contribution of the Residue Glu9, Glu46, and Phe49 to the Biological Activity of Insulin‐Like Growth Factor‐1
Authors: Chen Hui , Feng You‐Min
Published: September 2003

Three recombinant single‐chain insulin mutants, [B10Glu]Ins/IGF‐1(C), [A5Glu]PIP, and [A8Tyr]PIP were prepared by means of protein engineering. In the three mutants, the residue at position 10 of insulin B chain and positions 5 and 8 of insulin A chain were substituted by the corresponding residue Glu9, Glu46, and an aromatic residue at 49 of IGF‐1, respectively. The growth‐promoting activity of [B18Glu]Ins/IGF‐1(C) is 58‐fold higher than insulin and of [A5Glu]PIP, and [A8Tyr]PIP is 2.5‐ and 4.7‐fold higher than PIP, respectively, measured using a mouse mammary tumor cell line GR2H6 in vitro. The results show that the residues Glu9, Glu46, and Phe49, especially the Glu9, play an important role in the growth‐promoting function of IGF‐1. Based on the results, we suggest that the insulin could be used as a scaffold to investigate the contributions of residues of IGF‐1 to its biological function.
Molecular Basis of the Anaerobic Response in Plants
Authors: Dolferus Rudy , Klok Erik Jan , Ismond Kathleen , Delessert Christian , Wilson Sarah , Good Allen , Peacock Jim , Dennis Liz
Published: September 2003

The response of plants to flooding is complex and involves the induction of specific gene sets. A multidisciplinary approach by several research teams has led to a reasonably good understanding of the low oxygen response, and many of the genes and proteins that are involved are known. But the factors that are critical in determining tolerance or intolerance remain unknown. Microarray technology offers renewed hope to unravel the complex changes in gene expression occurring in plants upon low oxygen treatment and what mechanisms are involved in the response.
Protein Tyrosine Phosphatases: Dephosphorylating the Epidermal Growth Factor Receptor
Authors: Tiganis Tony
Published: September 2003

Protein tyrosine phosphatases (PTPs) are a large and structurally diverse family of enzymes that are found in eukaryotes, prokaryotes, viruses, and plants. PTPs catalyse the dephosphorylation of tyrosyl phosphorylated proteins and can either antagonise or potentiate protein tyrosine kinase signalling. PTPs regulate fundamental cellular processes and have been implicated in the etiology and pathogenesis of various human diseases. The epidermal growth factor receptor (EGFR) is a widely distributed protein tyrosine kinase that regulates both normal development and plays a role in pathological conditions such as cancer. This review discusses the structure and function of PTPs and focuses on the PTPs that have been implicated in the dephosphorylation of the EGFR and the consequent suppression of EGFR signalling.
The Myotubularin Family: Novel Phosphoinositide Regulators
Authors: Nandurkar Harshal H. , Huysmans Richard
Published: September 2003

Phosphatidylinositol 3‐phosphate [PtdIns(3)P] acts as a second messenger via the recruitment of diverse signalling proteins to various cellular compartments. Recent advances have highlighted the association of human diseases with mutations in phosphatases that regulate PtdIns(3)P levels. Myotubularin, the gene mutated in myotubular myopathy, functions as a lipid phosphatase with specificity for PtdIns(3)P. It is now apparent that there is an increasing family of proteins that are defined by their significant homology with myotubularin. The myotubularin‐related gene family includes proteins that exhibit a lipid phosphatase catalytic motif, those that contain mutations of the critical catalytic residues, and at least one protein that functions as an adapter subunit for PtdIns(3)P‐3‐phosphatase activity. The present challenge is to understand how deregulation of phosphoinositide metabolism causes human disease.
From Cytosol to Organelles: 14‐3‐3 Proteins as Multifunctional Regulators of Plant Cell
Authors: Aducci Patrizia , Camoni Lorenzo , Marra Mauro , Visconti Sabina
Published: September 2003

14‐3‐3 proteins are a class of highly conserved proteins widespread in eukaryotes. They regulate several cellular processes through phosphorylation‐dependent interaction with their targets. Since their discovery in plants, a number of peculiar functions have been ascertained, such as regulation of primary metabolism, ion transport, cellular trafficking, chloroplast and mitochondrial enzyme activities and gene transcription. The still increasing body of evidence suggests that 14‐3‐3s may function as versatile proteins able to move from cytosol to different cellular organelles. This review will focus on the broad range of regulatory tasks carried out by 14‐3‐3s in the different compartments.
Inositol Polyphosphate 5‐Phosphatases: Lipid Phosphatases With Flair
Authors: Mitchell Christina A. , Gurung Rajendra , Kong Anne M. , Dyson Jennifer M. , Tan April , Ooms Lisa M.
Published: September 2003

Recent studies have identified the inositol polyphosphate 5‐phosphatases as a large family of signal modifying enzymes comprising 10 mammalian and 4 yeast family members. A number of investigations including gene‐targeted deletion of 5‐phosphatases in mice have demonstrated that these enzymes regulate many important cellular events including hematopoietic cell proliferation and activation, insulin signaling, endocytosis, and actin polymerization.
Genomic Structure and Promoter Analysis of the Bovine Leptin Gene
Authors: Taniguchi Yukio , Itoh Tomohiro , Yamada Takahisa , Sasaki Yoshiyuki
Published: September 2003

The product of the leptin gene is a 16‐kDa protein secreted by adipose tissue and regulates adiposity. The leptin gene could be a potential candidate gene controlling some proportion of adipose and lean accretion in cattle, and thus, may be referred to as one of genetic factor controlling meat quality determinants such as marbling. We have isolated the bovine leptin gene including its promoter region. We have determined the exon‐intron organization of the bovine leptin gene, which consisted of three exons and two introns and spanned about 18.9 kb, equivalent to that of human or mouse gene. A ∼3‐kb 5'‐flanking region upstream from the putative transcription start site of the gene contained consensus Sp1 and CCAAT/enhancer‐binding protein (C/EBP) motifs, and transient transfection assay with secreted alkaline phosphatase reporter confirmed the promoter activity in 3T3‐L1 cells that possessed expression of the cotransfected C/EBP αexpression plasmid. Cotransfection with C/EBP αcaused 24‐fold activation in leptin reporter expression, as compared to cotransfection with control plasmid, consistent with existence of the putative C/EBP αbinding site in the proximal 5'‐flanking region of the bovine leptin gene.
Animals are Dependent on Preformed α‐Amino Nitrogen as an Essential Nutrient
Authors: Katagiri Masayuki , Nakamura Masahiko
Published: September 2003

It has traditionally been thought that animals can utilize ammonia for amino acid biosynthesis, and that for them some amino acids are nutritionally nonessential. Presumably this idea originates from the notions of Schoenheimer (G. L. Foster et al. [1939] J. Biol. Chem. 127, 319‐327) and of Rose (W. C. Rose et al. [1948] J. Biol. Chem. 176, 753‐762), which we question for the following reasons. First, Schoenheimer's experiments only showed the incorporation of ammonia into amino acids. This may occur simply as an exchange between ammonia and the α‐amino group of endogenous amino acids and reflects the enzymatic properties of glutamate dehydrogenase, which is a reversible enzyme. Second, Rose's nutritional experiments were concerned with whether carbon skeletons of particular amino acids can (nonessential) or cannot (essential) be synthesized from common intermediates of carbohydrate metabolism. We propose that mammals, living as they do at the top of the food web, are absolutely dependent directly or indirectly on higher plants and microorganisms for preformed α‐amino nitrogen per se and that the first joining of C ‐ and N ‐atoms to make glutamate constitutes a basic anabolic system in nature after the fixation of CO 2 and N 2 .
tRNALys3: The Primer tRNA for Reverse Transcription in HIV‐1
Authors: Kleiman Lawrence
Published: September 2003

During the assembly of HIV‐1, tRNA Lys isoacceptors are selectively packaged into the virion, and one of these, tRNA Lys3, is annealed to the viral RNA genome where it acts to prime the reverse transcriptase (RT)‐catalyzed synthesis of viral DNA. We review herein what is known about the selective packaging and annealing of primer tRNA Lys3 . Current evidence suggests that a complex of two major precursor viral proteins, Pr55 gag and Pr160 gag‐pol, interact with a tRNA Lys /lysyl‐tRNA synthetase (LysRS) complex during viral assembly, with Pr55 gag interacting with both LysRS and Pr160 gag‐pol, and RT sequences within Pr160 gag‐pol binding to tRNA Lys . LysRS appears to target the tRNA Lys isoacceptors for incorporation into HIV‐1. In the virion, the 3' terminal 18 nucleotides of tRNA Lys3 anneals to an 18‐nucleotide sequence at the 5' terminal region of viral RNA termed the primer binding site (PBS). Evidence is presented that other regions on the tRNA Lys3 also anneal with other regions in viral RNA upstream of the PBS, resulting in a destabilized tRNA Lys3 structure. Both viral and tRNA Lys3 regions need to be denatured to establish annealing, and the roles of both viral and cellular proteins in this process are discussed.
Nitric Oxide Limits Parasite Development in Vectors and in Invertebrate Intermediate Hosts
Authors: Gradoni Luigi , Ascenzi Paolo
Published: September 2003

Nitric oxide (NO) possesses antiparasitic effects on both Protozoa and Metazoa in vertebrate definitive and intermediate hosts. Inducible NO limits parasite development also in Rhodnius prolixus and Anopheles stephensi, the natural vectors of human trypanosomiasis and malaria respectively, and in the snail Biomphalaria glabrata, a natural invertebrate intermediate host of human schistosomiasis. Therefore, NO limits Trypanosoma, Plasmodium, and Schistosoma development at all stages of the parasite life cycle.
BAG‐1, An Anti‐Apoptotic Tumour Marker
Authors: Tang Shou‐Ching
Published: September 2003

BAG‐1 is a multifunctional and anti‐apoptotic or anti‐cell death protein that interacts with a variety of cellular proteins and affects their functions. On the cell surface, it binds to the cytosolic domain of the growth factor receptors and enhances the protection from cell death triggered by growth factor receptors. In the cytosol, it binds to Bcl‐2 and heat shock protein, and modulates their functions. In the nucleus, it binds to a variety of nuclear hormone receptors and inhibits hormone‐induced apoptosis. BAG‐1 is widely overexpressed in a variety of tumour cell lines and cancer tissues. In addition, differential expression of BAG‐1 isoforms has been observed. Preclinical studies indicate that overexpression of BAG‐1, especially its nuclear and cytoplasmic isoforms, may be useful as a prognostic and/or predictive biomarker. Pilot clinical studies have demonstrated that overexpression of nuclear BAG‐1 may be associated with a shorter survival in breast and laryngeal carcinomas. Conversely, overexpression of cytoplasmic BAG‐1 may be associated with a better clinical outcome in early stage breast cancer and in non‐small cell lung cancer. Further large‐scale clinical studies are warranted to establish the role of BAG‐1 as a novel prognostic and/or predictive biomarker in the clinical management of these common malignancies.
Oxalate, Germins, and Higher‐Plant Pathogens
Authors: Lane Byron G.
Published: September 2003

Earlier surveys ( 1, B. G. Lane. [1991] FASEB J. 5, 2983‐2901; 2, B. G. Lane. [1994] FASEB J. 8, 294‐301) helped to uproot entrenched views of plant oxalate as a static substance. It is now recognized that oxalate oxidases (OXOs) found in the “true cereals” (barley, maize, oat, rice, rye, wheat), the so‐called germin OXOs (G‐OXOs), or simply germins, are involved in cereal defence responses to invasion by fungal pathogens and that they show promise of being valuable agents of plant defence in dicotyledons, where they are not found naturally. G‐OXOs have very peculiar properties: (a) their water‐soluble oligomeric structures and enzymic activity are stable during SDS‐PAGE and nitrocellulose blotting, (b) their undenatured water‐soluble forms are refractory to the action of broad‐specificity proteases, (c) their water‐insoluble forms occur abundantly (∼50%) in the extracellular matrix (cell walls) of wheat, and probably in varying amounts in the cell walls of other true cereals. Transfer of the wheat G‐OXO coding element to dicotyledons has been found, in all cases so far examined, to result in improved resistance to fungal pathogens. The possible nature of the improved resistance is discussed in relation to (a) generation of microcidal concentrations of hydrogen peroxide when the G‐OXOs act on oxalate, (b) elicitation of hypersensitive cell death at lower concentrations of hydrogen peroxide, (c) formation of effective barriers against predator penetration by the hydrogen‐peroxide‐mediated lignification of cell walls, and (d) destruction of oxalate, which is an inhibitor of the hypersensitive response, a strategy of particular importance in the case of ubiquitous predator organisms such as Sclerotinia sclerotiorum, which secrete high concentrations of oxalate as a toxin.
Atrial Natriuretic Peptide Prohormone Gene Expression: Hormones and Diseases That Upregulate its Expression
Authors: Vesely David L.
Published: September 2003

Atrial natriuretic peptides consist of a family of peptide hormones that are synthesized by three separate genes and then stored as three different prohormones (i.e., 126‐amino acid [a.a.]) atrial natriuretic peptide (ANP), 108‐a.a. brain natriuretic peptide (BNP), and 126‐a.a. C‐natriuretic peptide (CNP) prohormones. The gene encoding for the synthesis of the atrial natriuretic peptide prohormone (proANP) consists of three exons and two introns. Exon 1 encodes the signal peptide and the first 16 a.a. of the ANP prohormone. These 16 a.a. form the N ‐terminus of a peptide hormone named long‐acting natriuretic hormone (LANH). A valine‐to‐methionine substitution in LANH results in a 2‐fold increased incidence of strokes in humans. Exon 2 of the proANP gene encodes for three peptide hormones, i.e., vessel dilator, kaliuretic hormone, and ANP. Each of the proANP gene products have vasodilatory, diuretic, natriuretic, and/or kaliuretic properties. Stretch, glucocorticoids, thyroid hormone(s), mineralocorticoids, and calcium enhance proANP gene expression. Enhanced proANP gene expression is found in congestive heart failure, hypertension, and cirrhosis with ascites. The proANP gene is present with invertebrates and plants as well as in humans and other vertebrates.
Prediction of Drug Resistance in M. tuberculosis: Molecular Mechanisms, Tools, and Applications
Authors: Victor Thomas C. , van Helden Paul D. , Warren Robin
Published: September 2003

Management of Tuberculosis is complicated by the emergence of drug resistant strains of Mycobacterium tuberculosis and this poses a threat to the success of Tuberculosis control programmes. Drug susceptibility testing by culture is time‐consuming and technically difficult. It is known that resistance to drugs is due to a number of genomic mutations in specific genes of M. tuberculosis . These mutations in combination with molecular techniques can be used as markers for drug resistance, since drug susceptible isolates lack the corresponding gene mutations. This review focuses on molecular mechanisms, methods and applications as a possible new diagnostic tool for the early molecular detection of drug resistance in M. tuberculosis .
Oesophageal Cancer in Africa
Authors: Hendricks Denver , Parker M. Iqbal
Published: September 2003

Oesophageal cancer, the eighth most frequent cancer in the world occurs as two main subtypes, squamous cell carcinoma (more prevalent in developing countries) and adenocarcinoma (more common in developed countries). Certain populations of central, eastern, and southern Africa display very high frequencies of oesophageal squamous cell carcinoma, presenting a serious health burden to the continent. Most patients are diagnosed at a late stage because of the asymptomatic development of the disease, with associated poor prognosis. A better understanding of the aetiological agents and molecular mechanisms involved in the development of oesophageal squamous cell carcinoma may offer opportunities to reduce exposure to environmental risk factors and also allow early diagnosis or predict response to therapy. Epidemiologic studies have identified smoking, alcohol consumption, diets poor in fresh fruit and vegetables, consumption of foods contaminated with Fusarium verticillioides, and HPV infection as risk factors associated with the development of this disease in Africa. Although we have an incomplete understanding of the molecular events involved in the development of oesophageal squamous cell carcinoma, advances have been made that suggest lines of future exploration. South African patients with oesophageal squamous cell carcinoma display a lower incidence of point mutations in the p53 gene than described elsewhere, suggesting that the profile of aetiological agents may be different than described for other high‐risk areas for oesophageal cancer. Recent studies suggest that RAR βand COX II is frequently downregulated and upregulated, respectively, in oesophageal squamous cell carcinomas. These results suggest potential therapeutic opportunities that can be exploited to combat the high incidence of this disease in Africa.
Human Genetic Susceptibility to Tuberculosis and Other Mycobacterial Diseases
Authors: Hoal Eileen G.
Published: September 2003

The existence of a genetic component in mycobacterial disease susceptibility is no longer in doubt and the investigations now being conducted aim to determine which genes are involved, to what extent, and in which disease phenotype they are relevant. In certain rare instances of susceptibility to poorly pathogenic mycobacteria, the genetic component is clear. The approaches employed to elucidate common disease susceptibility include linkage studies, particularly genome‐wide linkage analysis of both tuberculosis and leprosy, and association studies. A number of candidate genes have shown association with tuberculosis, and in many cases, on replication of the study, association has been confirmed in a disparate population, indicating the wider importance of the gene in the disease process. In other instances, associations appear to be particular to a population or a subtype of disease.
Bacterial Multidrug Resistance Mediated by a Homologue of the Human Multidrug Transporter P‐glycoprotein
Authors: Konings Wil N. , Poelarends Gerrit J.
Published: September 2003

Most ATP‐binding cassette (ABC) multidrug transporters known to date are of eukaryotic origin, such as the P‐glycoproteins (Pgps) and multidrug resistance‐associated proteins (MRPs). Only one well‐characterized ABC multidrug transporter, LmrA, is of bacterial origin. On the basis of its structural and functional characteristics, this bacterial protein is classified as a member of the P‐glycoprotein cluster of the ABC transporter superfamily. LmrA can even substitute for P‐glycoprotein in human lung fibroblast cells, suggesting that this type of transporter is conserved from bacteria to man. The functional similarity between bacterial LmrA and human P‐glycoprotein is further exemplified by their currently known spectrum of substrates, consisting mainly of hydrophobic cationic compounds. In addition, LmrA was found to confer resistance to eight classes of broad‐spectrum antibiotics, and homologs of LmrA have been found in pathogenic bacteria, supporting the clinical and academic value of studying this bacterial protein. Current studies are focused on unraveling the mechanism by which ABC multidrug transporters, such as LmrA, couple the hydrolysis of ATP to the translocation of drugs across the membrane. Recent evidence indicates that LmrA mediates drug transport by an alternating two‐site transport mechanism.
Thiol Metabolism of the Trypanosomatids as Potential Drug Targets
Authors: Steenkamp Daniel J.
Published: September 2003

Trypanosomatids produce significant amounts of four major low molecular mass thiols, trypanothione, glutathionylspermidine, glutathione, and ovothiol A. Of these, only glutathione is present in cells of the host. All four low molecular mass thiols are directly or indirectly maintained in a reduced state by trypanothione reductase. Available evidence, from gene disruption studies, indicate that this is an essential enzyme. Attempts to exploit trypanothione reductase as a chemotherapeutic target lead to the design of competitive and irreversible inhibitors of the enzyme. A promising route involves the design of redox cyclers interacting specifically with trypanothione reductase as subversive substrates. Progress in studies on the biosynthesis of ovothiol A is summarized.
HIV Receptors and Cellular Tropism
Authors: Weiss Robin A.
Published: September 2003

Viruses use specific cell surface receptors to bind to and subsequently gain entry into their host cells. Some retroviruses such as HIV‐1 and HIV‐2 utilize one receptor for high‐affinity binding (CD4), and a separate coreceptor to mediate fusion of the viral envelope with the cell membrane (CCR5 or CXCR4). The identification of these receptors explains the cellular tropism of HIV, and hence its pathogenesis leading to immune deficiency (T‐helper cell depletion), the wasting syndrome (macrophage infection), and dementia (microglia infection). HIV can infect cells by membrane fusion at the cell surface and by receptor‐mediated endocytosis. Knowledge of the HIV receptors has led to practical developments such as inhibitory drugs, reasons for genetic resistance to infection, and should inform the judicious choice of candidate vaccines.
Human Papillomavirus (HPV) Infection in Southern Africa: Prevalence, Immunity, and Vaccine Prospects
Authors: Williamson Anna‐Lise , Marais Dianne , Passmore Jo‐Ann , Rybicki Ed
Published: September 2003

Human papillomavirus (HPV) associated cancers are more prevalent in developing countries compared to developed countries. The major cancer caused by HPV is cervical cancer. The humoral immune response to HPV can be a marker of past infection but may also reflect persistent infection and cervical disease. IgA antibodies to HPV in oral fluid were also found to be markers of cervical disease. Cell mediated immunity is important in clearing HPV infection and for regression of the associated lesions: this means that women infected with HIV have a high prevalence of co‐infection with HPV. Good cervical screening programmes can control HPV associated cervical neoplasia. However, in countries such as South Africa, where these programmes are inadequate, there is a need for an HPV vaccine. The development of HPV vaccines is reviewed. There is a call for an inexpensive vaccine that will be accessible to the women that do not have access to adequate screening programmes and are therefore at the greatest risk of cervical cancer.
Alphavirus Replicon Particles as Candidate HIV Vaccines
Authors: Davis Nancy L. , West Ande , Reap Elizabeth , MacDonald Gene , Collier Martha , Dryga Sergey , Maughan Maureen , Connell Mary , Walker Chris , McGrath Kathryn , Cecil Chad , Ping Li‐Hua , Frelinger Jeff , Olmsted Robert , Keith Paula , Swanstrom Ronald , Williamson Carolyn , Johnson Philip , Montefiori David , Johnston Robert E.
Published: September 2003

Replicon particles based on Venezuelan equine encephalitis virus (VEE) contain a self‐replicating RNA encoding the VEE replicase proteins and expressing a gene of interest in place of the viral structural protein genes. Structural proteins for packaging of replicon RNA into VEE replicon particles (VRPs) are expressed from separate helper RNAs. Aspects of the biology of VEE that are exploited in VRP vaccines include 1) expression of very high levels of immunogen, 2) expression of immunizing proteins in cells in the draining lymph node, and 3) the ability to induce mucosal immunity from a parental inoculation. Results of experiments with VRPs expressing green fluorescent protein or influenza virus hemagglutinin (HA) demonstrated that specific mutations in the VRP envelope glycoproteins affect both targeting in the draining lymph node and efficiency of the immune response in mice. VRPs expressing either the matrix‐capsid portion of Gag, the full‐length envelope gp160, or the secreted gp140 of cloned SIVsm H‐4i were mixed in a cocktail and used to immunize macaques at 0, 1, and 4 months. Neutralizing antibodies against SIVsm H‐4 were induced in 6 of 6 vaccinates and CTL in 4 of 6. An intrarectal challenge with the highly pathogenic SIVsm E660 was given at 5 months. A vaccine effect was seen in reduced peak virus loads, reduced virus loads both at set point and at 41 weeks postchallenge, and preserved or increased CD4 counts compared to controls. A candidate VRP HIV vaccine expressing Clade C Gag contains a sequence that is very close to the South African Clade C consensus and was selected from a recent seroconverter in the Durban cohort to represent currently circulating genotypes in South Africa. A GMP lot of this vaccine has been manufactured and tested for a phase I trial in the first months of 2002.
Medicinal Chemistry and Chemical Biology of New Generation Taxane Antitumor Agents
Authors: Ojima Iwao , Geney Raphaël , Ungureanu Ioana Maria , Li Dansu
Published: September 2003

P‐glycoprotein (P‐GP)‐based multidrug resistance (MDR) and undesirable side effects are significant drawbacks to the clinical use of paclitaxel and docetaxel. Extensive SAR studies of taxanes in these laboratories led to the discovery of new generation taxanes that are highly active against not only drug‐sensitive but also drug‐resistant human cancer cell lines as well as tumor xenografts in mice. One of these second generation taxanes, SB‐T‐110131 (IDN5109), exhibited excellent pharmacological profile in the preclinical studies and has been selected for clinical development (re‐coded as Bay 59‐8862), which is currently in the phase II clinical trials. Bay 59‐8862 is orally active with high bioavailability, showing excellent activity against a variety of drug‐resistant tumors. “Advanced second generation taxanes” show essentially no difference in cytotoxicity against drug‐resistant and drug‐sensitive cell lines, virtually overcoming MDR. Photoaffinity labeling of P‐GP using photoreactive radiolabeled paclitaxel analogs has disclosed the paclitaxel‐binding domain of P‐GP. Highly efficient taxane‐based MDR reversal agents (TRAs) have also been developed, which can recover the cytotoxicity of paclitaxel to practically the original level against paclitaxel‐resistant MDR expressing cancer cells. Highly promising results have emerged from the study of taxane‐monoclonal antibody (MAb) immunoconjugates, which have been proved to specifically deliver extremely cytotoxic agents to tumor in an animal model.
Suppression of Gastric Mucosal Inflammatory Responses to Helicobacter pylori Lipopolysaccharide by Peroxisome Proliferator‐Activated Receptor γ Activation
Authors: Slomiany Bronislaw L. , Slomiany Amalia
Published: September 2003

Peroxisome proliferator‐activated receptor‐ γ(PPAR γ) is a nuclear transcription factor that regulates the expression of genes associated with inflammation. We applied the animal model of H. pylori lipopolysaccharide(LPS)‐induced gastritis to assess the effect of a specific PPAR γligand, ciglitazone, on the apoptotic processes and the mucosal activity of inducible nitric oxide synthase (NOS‐2), and the expression of COX‐1 and ‐2 cyclooxygenases. In the absence of ciglitazone, the LPS‐elicited mucosal inflammatory responses were accompanied by a massive epithelial cell apoptosis, upregulation of NOS‐2 and COX‐2 protein expression, and a marked increase in the mucosal PGE 2 generation and NOS‐2 activity. The expression of COX‐1 protein, however, remained unchanged. Administration of ciglitazone led to dose‐dependent reduction (up to 48%) in the severity of mucosal inflammatory involvement elicited by the LPS and this effect of the agent was reflected in a 72.5% reduction in apoptosis, a 58.7% decline in the mucosal PGE 2 generation and a 75.6% drop in NOS‐2 activity, and produced a marked decrease inCOX‐2 and NOS‐2 protein expression. Our findings demonstrate that PPAR γactivation suppresses gastric mucosal inflammatory responses to H. pylori LPS, and suggest that pharmacological manipulation of PPAR γactivation may provide therapeutic benefits in the resolution of inflammation associated with H. pylori infection.
The Complex Nature of Protein Phosphatases
Authors: Sim Alistair T. R. , Ludowyke Russell I.
Published: September 2003

Protein phosphatases are integrally associated with the regulation of cellular signaling. The mechanisms underlying the specific regulatory roles are likely to be unique to each cell system. Nevertheless, analysis of phosphatase regulation in a number of systems has identified phosphatase targeting through association with a wide range of binding partners to be a fundamental mechanism of regulation. Using protein phosphatase 2A (PP2A) as an example, this snapshot summarizes these fundamental mechanisms of protein phosphatase regulation.
E2F1 Activates the Human p53 Promoter and Overcomes the Repressive Effect of Hepatitis B Viral X Protein (HBx) on the p53 Promoter
Authors: Choi Murim , Lee Hyeon , Rho Hyune Mo
Published: September 2003

The functional effect of the interaction of E2F1 and hepatitis B virus X protein (HBx) on the promoter of human p53 gene was studied using chloramphenicol acetyl transferase (CAT) assay. E2F1 activated the p53 promoter through E2F1 binding site. As previously reported, HBx repressed the p53 promoter through E‐box. When E2F1 was cotransfected with HBx, E2F1 overcame the repressive effect of HBx on the p53 promoter through the E2F1 site. However, in the thymidine kinase ( tk ) heterologous promoter system with the E2F1 binding sites, cotransfection of E2F1 and HBx showed a strong synergistic activation. An in vitro interaction assay showed that E2F1 and HBx physically bind with each other. Analyses of the interaction domain with the GAL4 fusion protein showed that the pRb‐binding domain of E2F1 was necessary for the functional interaction of these two proteins. Taken together, these results imply the functional inhibitory action of E2F1 on the HBV life cycle and HBV‐mediated hepatocellular carcinogenesis (HCC). Therefore, the normal or enhanced function of E2F1 gene would be important in controlling the HBx function in HCC.
Do Hemoglobin and Hemocyanin Impair Schistosoma Killing by NO?
Authors: Ascenzi Paolo , Fasano Mauro , Gradoni Luigi
Published: September 2003

Nitric oxide (NO) possesses antiparasitic effects on both Protozoa and Metazoa in definitive and intermediate hosts as well as in vectors. Here, we postulate that hemoglobin and hemocyanin may impair Schistosoma killing by NO in the definitive and intermediate hosts. Interestingly, hemoglobin, myoglobin, and neuroglobin may protect Plasmodium and Trypanosoma from the antiparasitic effects of NO.
Biochemical and Cytochemical Characterization of Extracellular Proteoglycans in the Inner Circular Smooth Muscle Layer of Dog Small Intestine
Authors: de Toledo Olga Maria Szymanski , Marquezini Mônica Valéria , Jia Kaio Bin , Pinheiro Mônica de Campos , Mora Oswaldo Alves
Published: September 2003

Current literature concerning smooth muscle blood vessels has shown versican as the main proteoglycan (PG) component of the matrix. To show whether smooth muscle matrix has the same PG distribution when present in organs, other than the blood vessels, the inner circular smooth muscle layer of the small intestine was obtained by dissection as a highly purified tissue and analyzed by biochemical and cytochemical methods. The smooth muscle layer PGs were extracted from dog small intestine with 4 M guanidine‐HCl in the presence of proteinase inhibitors, purified by charge equilibrium, isolated by equilibrium CsCl density gradients, and analyzed in terms of anion exchange, size, and glycosaminoglycan (GAG) distribution. Proteoheparan sulfate itself represented 91.5% of the PGs present in this tissue. The remainder was proteodermatan sulfate. Cytochemical analyses using the cationic dye cuprolinic blue associated with enzymatic treatments with chondroitinases ABC and heparitinase III showed the arrangement and identification of PGs in basal lamina and intramuscular connective tissues. The PGs in the basal lamina were proteoheparan sulfate, and those associated with collagen fibrils in the endomysium and perimysium were rich in dermatan sulfate. In contrast to the blood vessels, inner circular muscle smooth tissue in intestine has, as the main PG, perlecan.
The Effects of Protein Hydrophobic Exposure on the Slope of the Line in the Δπ vs πi Plot
Authors: Zhang Fan , Lu Ying‐Jie , Wang Shao‐Xiong , Sui Sen‐Fang
Published: September 2003

The Δπvs πi curve from monolayer surface pressure detection is a powerful method to characterize the membrane insertion. The Δπvs πi curve is fixed by two parameters, the πc and the slope. The intersected point ( πc ) of Δπvs πi curve with abscissa is generally used as a quantitative measure of the membrane insertion ability of a protein. In the current work we demonstrate a correlation between the variation in the slope of the Δπvs πi curve with protein hydrophobic exposure by performing monolayer experiments on three different proteins, human apolipoprotein H, trichosanthin, and mutant trichosanthin. The value of |slope| increases gradually following the degree of hydrophobic exposure. These findings suggest that the two parameters, the πc and the slope, will complement each other to interpret the lipid/protein interaction involved in membrane insertion.
Swapping of Three‐Dimensional Domains as a Molecular Mechanism of Dimerization of Aminoacyl‐tRNA Synthetases
Authors: Deniziak Marzanna A. , Barciszewski Jan
Published: September 2003

It has been shown recently that many proteins undergo oligomerization through exchange of structural elements. That process, termed a 3D domain swapping, is the replacement of a portion of the tertiary structure of a protein with an identical piece from a second polypeptide chain. When the exchange is reciprocated, domain‐swapped dimers embrace with the exchange of elements of secondary structure or domains; however, if the exchange is not reciprocated but propagated along multiple polynucleotide chains, higher‐order assemblies may form. In this paper we discuss swapping as a general mechanism of aminoacyl‐tRNA synthetases dimerization, specifically plant methionyl‐tRNA synthetase.
Regulation of Mitochondrial Gene Expression by Retinoids
Authors: Everts Helen B. , Berdanier Carolyn D.
Published: September 2003

Several nuclear hormone receptors have been localized to the mitochondrial compartment. Evidence supports the hypothesis that these receptors directly regulate mitochondrial transcription. Retinoic acid has also been shown to regulate mitochondrial transcription and function. This review discusses mechanisms of mitochondrial transcription and how retinoic acid may either indirectly or directly regulate mitochondrial transcription. How retinoic acid may affect individual nutrient requirements is also discussed.
Thymocyte Apoptosis Induced by Phosphorylation of Histones is Associated with the Change in Chromatin Structure to Allow Easy Accessibility of DNase
Authors: Enomoto Riyo , Tatsuoka Hiroyuki , Yoshida Yukari , Komai Tomoe , Node Kaori , Nogami Rieko , Yamauchi Aiko , Lee Eibai
Published: September 2003

The inhibitors of protein phosphatase such as calyculin A and okadaic acid induce the apoptotic cell death in rat thymocytes. To clarify the molecular mechanism of these inhibitor‐induced apoptosis, the effect of calyculin A on DNA fragmentation in the isolated nuclei were studied. A significant increase in DNA fragmentation was observed in the nuclei prepared from the cells treated with calyculin A that caused histone hyperphosphorylation. No changes of the activities of caspase‐8 and ‐3 were observed in the extract from the cells treated with calyculin A. The circular dichroism analysis of soluble chromatin from calyculin A‐treated thymocyte nuclei indicated that phosphorylation of histones decreased its α‐helical content. Thus, the change in the chromatin structure may be due to the chemical modification of histones. Moreover, the structural change in chromatin preceded DNA fragmentation in the nuclei. Therefore, these results suggest that the change of chromatin structure allow easy accessibility of nuclear DNase to chromosomal DNA.
In Vitro Selection of Self‐Interacting Transmembrane Segments‐‐Membrane Proteins Approached from a Different Perspective
Authors: Langosch Dieter , Lindner Eric , Gurezka Rolf
Published: September 2003

The principles underlying the folding of integral membrane proteins are uncovered in an increasingly detailed way. Experimental determination of high‐resolution structures followed by analysis of packing reveal structural similarities as well as differences to soluble globular proteins. At the same time, protein/protein interactions at the level of membrane‐embedded domains have been investigated for different model proteins. More recently, self‐interacting transmembrane helices have been selected from combinatorial libraries in vitro to study the mechanistic basis of protein/protein interaction in membranes in a systematic way. With an emphasis on the latter approach, this review discusses insights emerging from an integrated view on the recent advances.
Biosynthesis of Chondroitin/Dermatan Sulfate
Authors: Silbert Jeremiah E. , Sugumaran Geetha
Published: September 2003

Chondroitin sulfate and dermatan sulfate are synthesized as galactosaminoglycan polymers containing N ‐acetylgalactosmine alternating with glucuronic acid. The sugar residues are sulfated to varying degrees and positions depending upon the tissue sources and varying conditions of formation. Epimerization of any of the glucuronic acid residues to iduronic acid at the polymer level constitutes the formation of dermatan sulfate. Chondroitin/dermatan glycosaminoglycans are covalently attached by a common tetrasaccharide sequence to the serine residues of core proteins while they are adherent to the inner surface of endoplasmic reticulum/Golgi vesicles. Addition of the first sugar residue, xylose, to core proteins begins in the endoplasmic reticulum, followed by the addition of two galactose residues by two distinct glycosyl transferases in the early cis/medial regions of the Golgi. The linkage tetrasaccharide is completed in the medial/trans Golgi by the addition of the first glucuronic acid residue, followed by transfer of N ‐acetylgalactosamine to initiate the formation of a galactosaminoglycan rather than a glucosaminoglycan. This specific N ‐acetylgalactosaminyl transferase is different from the chondroitin synthase involved in generation of the repeating disaccharide units to form the chondroitin polymer. Sulfation of the chondroitin polymer by specific sulfotransferases occurs as the polymer is being formed. All the enzymes in the pathway for synthesis have been cloned, with the exception of the glucuronyl to iduronyl epimerase involved in the formation of dermatan residues.
Functional Characteristics and Catalytic Mechanisms of the Bacterial Hyaluronan Synthases
Authors: Weigel Paul H.
Published: September 2003

The first gene for a glycosaminoglycan synthase to be cloned was the hyaluronan (HA) synthase from S. pyogenes, which we reported in 1993. Since then, at least 20 bacterial, viral, or eukaryotic HA synthase gene or cDNA sequences and two bacterial chondroitin synthase genes have been reported. During the last decade a great deal has been elucidated about the structure, function, and mechanisms of action of the bacterial HA synthases, which are the focus of this review. Very rapid progress has been made in elucidating the mechanism of HA synthesis by the HA synthase from Pasteurella multocida . Although little of this information is applicable to understanding the mechanism of action of streptococcal HA synthases, good progress has also been made in understanding how these latter enzymes work.
Keratan Sulfate Biosynthesis
Authors: Funderburgh James L.
Published: September 2003

Keratan sulfate was originally identified as the major glycosaminoglycan of cornea but is now known to modify at least a dozen different proteins in a wide variety of tissues. Despite a large body of research documenting keratan sulfate structure, and an increasing interest in the biological functions of keratan sulfate, until recently little was known of the specific enzymes involved in keratan sulfate biosynthesis or of the molecular mechanisms that control keratan sulfate expression. In the last 2 years, however, marked progress has been achieved in identification of genes involved in keratan sulfate biosynthesis and in development of experimental conditions to study keratan sulfate secretion and control in vitro. This review summarizes current understanding of keratan sulfate structure and recent developments in understanding keratan sulfate biosynthesis.
Novel Mechanisms for Feedback Regulation of Phospholipase C‐β Activity
Authors: Litosch Irene
Published: September 2003

The receptor‐regulated phospholipase C‐ β(PLC‐ β) signaling pathway is an important component in a network of signaling cascades that regulate cell function. PLC‐ βsignaling has been implicated in the regulation of cardiovascular function and neuronal plasticity. The G q family of G proteins mediate receptor stimulation of PLC‐ βactivity at the plasma membrane. Mitogens stimulate the activity of a nuclear pool of PLC‐ β. Stimulation of PLC‐ βactivity results in the rapid hydrolysis of phosphatidylinositol‐4,5‐bisphosphate, with production of inositol‐1,4,5‐trisphosphate and diacylglycerol, intracellular mediators that increase intracellular Ca 2+ levels and activate protein kinase C activity, respectively. Diacylglycerol kinase converts diacylglycerol to phosphatidic acid, a newly emerging intracellular mediator of hormone action that targets a number of signaling proteins. Activation of the G q linked PLC‐ βsignaling pathway can also generate additional signaling lipids, including phosphatidylinositol‐3‐phosphate and phosphatidylinositol‐3,4,5‐trisphosphate, which regulate the activity and/or localization of a number of proteins. Novel feedback mechanisms, directed at the level of G q and PLC‐ β, have been identified. PLC‐ βand regulators of G protein signaling (RGS) function as GTPase‐activating proteins on G q to control the amplitude and duration of stimulation. Protein kinases phosphorylate and regulate the activation of specific PLC‐ βisoforms. Phosphatidic acid regulates PLC‐ β1 activity and stimulation of PLC‐ β1 activity by G proteins. These feedback mechanisms coordinate receptor signaling and cell activation. Feedback mechanisms constitute possible targets for pharmacological intervention in the treatment of disease.
p38 Kinase and c‐Jun N‐Terminal Kinase Oppositely Regulates Tumor Necrosis Factor α‐Induced Vascular Cell Adhesion Molecule‐1 Expression and Cell Adhesion in Chondrosarcoma Cells
Authors: Ju Jung‐Won , Kim Song‐Ja , Jun Chang‐Duk , Chun Jang‐Soo
Published: September 2003

We investigated signaling pathways leading to tumor necrosis factor (TNF) α‐induced intercellular adhesion molecule (ICAM)‐1 and vascular cell adhesion molecule (VCAM)‐1 expression in chondrosarcoma cells, and determined the functional significance of their expression by examining Jurkat T cell adhesion. TNF αinduced VCAM‐1 and ICAM‐1 expression and Jurkat T cell binding. Antibody blocking assay indicated that VCAM‐1 mediates TNF α‐induced Jurkat T cell adhesion. TNF αcaused activation of mitogen‐activated protein (MAP) kinase subtypes, extracellular signal‐regulated protein kinase, p38 kinase, and c‐jun N‐terminal kinase (JNK). ICAM‐1 expression was not altered by the inhibition of MAP kinases. However, VCAM‐1 expression and Jurkat T cell adhesion was blocked by the inhibition of p38 kinase, whereas inhibition of JNK enhanced VCAM‐1 expression and cell adhesion without any modulation of NF κB activation. Our results, therefore, indicate that p38 kinase mediates TNF α‐induced VCAM‐1 expression and cell adhesion, whereas JNK suppresses VCAM‐1 expression that is independent to NF κB activation.
The Overexpression Catalase Reduces NO‐Mediated Inhibition of Endothelial NO Synthase
Authors: Brennan Lisa A. , Wedgwood Stephen , Black Stephen M.
Published: September 2003

Previously, we have demonstrated that increased superoxide generation plays a role in the nitric oxide (NO)‐mediated inhibition of endothelial NO synthase (eNOS) in endothelial cells (ECs) and that the overexpression of SOD1 could reduce the inhibitory effect of NO. However, SOD1 overexpression did not completely abolish the inhibition of eNOS by NO, indicating the presence of other inhibitory mechanisms. Because superoxide can be dismutated into hydrogen peroxide (H 2 O 2 ), in this study we determined whether exposure of ECs to NO resulted in increased generation of H 2 O 2 and the potential role of H 2 O 2 in eNOS inhibition. Our results indicated that H 2 O 2 levels were increased in response to NO. Using adenoviral‐mediated infection, we demonstrated that catalase overexpression both increased basal eNOS activity in the absence of NO and provided a significant protective effect on eNOS activity in the presence of NO. This protective effect was associated with a significant decrease in H 2 O 2 levels in the presence of NO. In conclusion, our results indicate that increased levels of H 2 O 2 may be involved in the inhibition of eNOS by NO and that the scavenging of H 2 O 2 may be useful to prevent eNOS inhibition during treatments that involve inhaled NO or NO donors.
Nitric Oxide as a Modulator of Gastric Mucin Synthesis: Role of ERK and p38 Mitogen‐Activated Protein Kinase Activation
Authors: Slomiany Bronislaw L. , Slomiany Amalia
Published: September 2003

Nitric oxide (NO) is an important biological messenger in the regulation of tissue homeostasis and pathophysiological processes. Here, we investigated the effect of NO on gastric mucus glycoprotein (mucin) synthesis, apoptotic processes, and the involvement of extracellular signal‐regulated kinase (ERK) and p38 mitogen‐activated protein kinase (MAPK). Exposure of gastric mucosal cells to NO donor led to a dose‐dependent decrease (up to 48%) in mucin synthesis, accompanied by a marked increase in caspase‐3 activity and apoptosis. Inhibition of ERK with PD98059 accelerated (up to 23.8%) the NO‐induced decrease in mucin synthesis, and cause further enhancement in caspase‐3 activity and apoptosis. Blockade of p38 kinase with SB203580 produced reversal in the NO‐induced reduction in mucin synthesis, and substantially countered the induced increase in caspase‐3 activity and apoptosis. Moreover, caspase‐3 inhibitor not only blocked the NO‐induced increase in caspase‐3 activity but also produced an increase in mucin synthesis. Thus, the detrimental influence of NO on mucin synthesis is closely linked to caspase‐3 activation and apoptosis, and involves ERK and p38 kinase participation. Activation of p38 kinase leads to the upregulation of proapoptotic signal, while ERK activation stimulates the anti‐apoptotic pathway.