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L Shapiro

Publications and source records attributed to L Shapiro.

At least 37 records · Page 2Linked to original sources
PubMed1999

Predominant expression of the mitochondrial dicarboxylate carrier in white adipose tissue.

We report the identification of a novel mouse protein closely related to the family of mitochondrial uncoupling proteins and the oxoglutarate carrier. The cDNA encodes a protein of 287 amino acids that shares all the hallmark features of the mitochondrial transporter superfamily, including six predicted transmembrane domains. It is nearly identical to the sequence recently reported for the rat mitochondrial dicarboxylate carrier (DIC). We find that murine DIC (mDIC) is expressed at very high levels in mitochondria of white adipocytes and is strongly induced in the course of 3T3-L1 adipogenesis. To determine the consequences of the presence of mDIC on the mitochondrial membrane potential, we transiently expressed mDIC in 293-T cells. Overexpression of mDIC leads to significant mitochondrial hyperpolarization. In addition, exposure to cold down-regulates mDIC levels in vivo. In contrast, free fatty acids lead to an up-regulation of mDIC protein in 3T3-L1 adipocytes. This is the first report demonstrating preferential expression in white adipose tissue of any mitochondrial transporter. However, it remains to be determined which metabolic pathways most critically depend on high level expression of mDIC in the adipocyte.

3T3 Cells↗
PubMed1999

The Caulobacter crescentus smc gene is required for cell cycle progression and chromosome segregation.

The highly conserved SMC (Structural Maintenance of Chromosomes) proteins function in chromosome condensation, segregation, and other aspects of chromosome dynamics in both eukaryotes and prokaryotes. A null mutation in the Caulobacter crescentus smc gene is conditionally lethal and causes a cell cycle arrest at the predivisional cell stage. Chromosome segregation in wild-type and smc null mutant cells was examined by monitoring the intracellular localization of the replication origin and terminus by using fluorescence in situ hybridization. In wild-type cells, the origin is located at the flagellated pole of swarmer cells and, immediately after the initiation of DNA replication in stalked cells, one of the origins moves to the opposite pole, giving a bipolar localization of the origins. The terminus moves from the end of the swarmer cell opposite the origin to midcell. A subpopulation of the smc null mutant cells had mislocalized origins or termini, showing that the smc null mutation gives DNA segregation defects. Nucleoid morphology was also abnormal. Thus, we propose that the Caulobacter chromosomal origins have specific cellular addresses and that the SMC protein plays important roles in maintaining chromosome structure and in partitioning. The specific cell cycle arrest in the smc null mutant indicates the presence of a cell cycle checkpoint that senses perturbations in chromosome organization or segregation.

Bacterial Proteins↗
PubMed1999

Feedback control of a master bacterial cell-cycle regulator.

The transcriptional regulator CtrA controls several key cell-cycle events in Caulobacter crescentus, including the initiation of DNA replication, DNA methylation, cell division, and flagellar biogenesis. CtrA is a member of the response regulator family of two component signal transduction systems. Caulobacter goes to great lengths to control the time and place of the activity of this critical regulatory factor during the cell cycle. These controls include temporally regulated transcription and phosphorylation and spatially restricted proteolysis. We report here that ctrA expression is under the control of two promoters: a promoter (P1) that is active only in the early predivisional cell and a stronger promoter (P2) that is active in the late predivisional cell. Both promoters exhibit CtrA-mediated feedback regulation: the early P1 promoter is negatively controlled by CtrA, and the late P2 promoter is under positive feedback control. The CtrA protein footprints conserved binding sites within the P1 and P2 promoters. We propose that the P1 promoter is activated after the initiation of DNA replication in the early predivisional cell. The ensuing accumulation of CtrA results in the activation of the P2 promoter and the repression of the P1 promoter late in the cell cycle. Thus, two transcriptional feedback loops coupled to cell cycle-regulated proteolysis and phosphorylation of the CtrA protein result in the pattern of CtrA activity required for the temporal and spatial control of multiple cell-cycle events.

Bacterial Proteins↗
PubMed1999

Cell cycle-dependent polar localization of an essential bacterial histidine kinase that controls DNA replication and cell division.

The master CtrA response regulator functions in Caulobacter to repress replication initiation in different phases of the cell cycle. Here, we identify an essential histidine kinase, CckA, that is responsible for CtrA activation by phosphorylation. Although CckA is present throughout the cell cycle, it moves to a cell pole in S phase, and upon cell division it disperses. Removal of the membrane-spanning region of CckA results in loss of polar localization and cell death. We propose that polar CckA functions to activate CtrA just after the initiation of DNA replication, thereby preventing premature reinitiations of chromosome replication. Thus, dynamic changes in cellular location of critical signal proteins provide a novel mechanism for the control of the prokaryote cell cycle.

Bacterial Proteins↗
PubMed1999

Chromosome segregation during the prokaryotic cell division cycle.

Recent work has dramatically changed our view of chromosome segregation in bacteria. Rather than being a passive process, it involves rapid movement of parts of the circular chromosome. Several genes involved in chromosome segregation have been identified, and the analysis of their functions and intracellular localization are beginning to shed light on the mechanisms that ensure efficient chromosome segregation.

Bacillus subtilis↗
PubMed1999

Differential localization of two histidine kinases controlling bacterial cell differentiation.

The bacterium C. crescentus coordinates cellular differentiation and cell cycle progression via a network of signal transduction proteins. Here, we demonstrate that the antagonistic DivJ and PleC histidine kinases that regulate polar differentiation are differentially localized as a function of the cell cycle. The DivJ kinase localizes to the stalked pole in response to a signal at the G1-to-S transition, while the PleC kinase is localized to the flagellar pole in swarmer and predivisional cells but is dispersed throughout the cell in the stalked cell. PleC, which is required for DivJ localization, may provide the cue at the G1-to-S transition that directs the polar positioning of DivJ. The dynamic positioning of signal transduction proteins may contribute to the regulation of polar differentiation at specific times during the bacterial cell cycle.

Bacterial Proteins↗
PubMed1999

The CtrA response regulator mediates temporal control of gene expression during the Caulobacter cell cycle.

In its role as a global response regulator, CtrA controls the transcription of a diverse group of genes at different times in the Caulobacter crescentus cell cycle. To understand the differential regulation of CtrA-controlled genes, we compared the expression of two of these genes, the fliQ flagellar gene and the ccrM DNA methyltransferase gene. Despite their similar promoter architecture, these genes are transcribed at different times in the cell cycle. PfliQ is activated earlier than PccrM. Phosphorylated CtrA (CtrA approximately P) bound to the CtrA recognition sequence in both promoters but had a 10- to 20-fold greater affinity for PfliQ. This difference in affinity correlates with temporal changes in the cellular levels of CtrA. Disrupting a unique inverted repeat element in PccrM significantly reduced promoter activity but not the timing of transcription initiation, suggesting that the inverted repeat does not play a major role in the temporal control of ccrM expression. Our data indicate that differences in the affinity of CtrA approximately P for PfliQ and PccrM regulate, in part, the temporal expression of these genes. However, the timing of fliQ transcription but not of ccrM transcription was altered in cells expressing a stable CtrA derivative, indicating that changes in CtrA approximately P levels alone cannot govern the cell cycle transcription of these genes. We propose that changes in the cellular concentration of CtrA approximately P and its interaction with accessory proteins influence the temporal expression of fliQ, ccrM, and other key cell cycle genes and ultimately the regulation of the cell cycle.

Bacterial Proteins↗
PubMed1999

Loving openness as a meta-world hypothesis: expanding our vision of mind and medicine.

In 1942, Stephen C. Pepper published a seminal book, World Hypothesis, that sought to explain how people create hypotheses about the world. Pepper proposed that there were four basic world hypotheses: (1) formistic (the hypothesis that nature exists as categories); (2) mechanistic (the hypothesis that nature obeys cause-effect relationships); (3) contextual (the hypothesis that processes in nature are relative and context dependent); and (4) organismic (the hypothesis that processes in nature reflect interactive relationships in systems). Most classical and modern theories of science and medicine implicitly adopt one or more of these foundational hypotheses. In 1997, Schwartz and Russek proposed that there were four additional world hypotheses: (5) implicit process (the hypothesis that nature consists of invisible forces and information, such as energy and consciousness); (6) circular causality (the hypothesis that nature consists of circulating interactions that inherently change over time); (7) creative unfolding (the hypothesis that processes in nature reflect flexible designs or plans that have adaptive function); and (8) integrative diversity (the hypothesis that phenomena in nature reflect complex integrations of diverse processes). Theories in postmodern and integrative science implicitly adopt hypotheses 5 through 8. However, underlying the creation of the eight world hypotheses is an implicit meta-world hypothesis which we term loving openness (the hypothesis that phenomena in nature reflect levels of openness and caring). This paper briefly explains the origin and implications of the loving openness meta-world hypothesis for understanding mind-body healing in medicine, the reordering of values in integrative medicine (with a primary focus on caring with humility), and the fostering of a new vision for twenty-first century frontier science, spirituality, and medicine.

Attitude↗
PubMed1998

Interleukin 18 stimulates HIV type 1 in monocytic cells.

The cytokine interleukin (IL) 18 (formerly interferon gamma-inducing factor) induces the T helper type 1 response. In the present studies, IL-18 increased HIV type 1 (HIV-1) production from 5- to 30-fold in the chronically infected U1 monocytic cell line. Inhibition of tumor necrosis factor (TNF) activity by the addition of TNF-binding protein reduced IL-18-stimulated HIV-1 production by 48%. In the same cultures, IL-18-induced IL-8 was inhibited by 96%. Also, a neutralizing anti-IL-6 mAb reduced IL-18-induced HIV-1 by 63%. Stimulation of U1 cells with IL-18 resulted in increased production of IL-6, and exogenous IL-6 added to U1 cells increased HIV-1 production 4-fold over control. A specific inhibitor of the p38 mitogen-activated protein kinase reduced IL-18-induced HIV-1 by 73%, and a 50% inhibition was observed at 0.05 microM. In the same cultures, IL-8 was inhibited by 87%. By gel-shift and supershift analyses, increased binding activity of the transcription factor NF-kappaB was measured in nuclear extracts from U1 cells 1 h after exposure to IL-18. These results demonstrate induction of HIV-1 by IL-18 in a monocyte target associated with an intermediate role for TNF and IL-6, activation of p38 mitogen-activated protein kinase, and nuclear translocation of NF-kappaB.

Base Sequence↗
PubMed1998

Role of p38 mitogen-activated protein kinase in HIV type 1 production in vitro.

The proinflammatory cytokines interleukin (IL)-1 and tumor necrosis factor (TNF) promote HIV type 1 viral replication in vitro. In the present studies, HIV production was increased in the macrophagic U1 cell line expressing the HIV genome after exposure to IL-1beta, osmotic stress, or surface adhesion, suggesting a confluence of signaling pathways for proinflammatory cytokines and cell stressors. The p38 mitogen-activated protein kinase (MAPK) mediates both cytokine and stress responses; thus the role of this kinase in HIV production was investigated. HIV production as measured by p24 antigen correlated with changes in the expression of a specific (non-alpha) isoform of p38 MAPK. In the presence of a specific p38 MAPK inhibitor (p38 inh), IL-1beta-induced HIV production was suppressed by more than 90% and IL-1beta-induced IL-8 production was suppressed completely, both with IC50 of 0.01 microM. p38 inhibition blocked cell-associated p24 antigen and secreted virus to a similar extent. The p38 inh also decreased constitutive HIV production in freshly infected peripheral blood mononuclear cells by up to 50% (P < 0.05). Interruption of p38 MAPK activity represents a viable target for inhibition of HIV.

Calcium-Calmodulin-Dependent Protein Kinases↗
PubMed1998

Interleukin-1beta-induced rat pancreatic islet nitric oxide synthesis requires both the p38 and extracellular signal-regulated kinase 1/2 mitogen-activated protein kinases.

Interleukin-1beta (IL-1beta) is cytotoxic to rat pancreatic beta-cells by inhibiting glucose oxidation, causing DNA damage and inducing apoptosis. Nitric oxide (NO) is a necessary but not sufficient mediator of these effects. IL-1beta induced kinase activity toward Elk-1, activation transcription factor 2, c-Jun, and heat shock protein 25 in rat islets. By Western blotting with phosphospecific antibodies and by immunocomplex kinase assay, IL-1beta was shown to activate extracellular signal-regulated kinase (ERK) 1/2 and p38 mitogen-activated protein kinase (p38) in islets and rat insulinoma cells. Specific ERK1/2 and p38 inhibitors individually reduced but in combination blocked IL-1beta-mediated islet NO synthesis, and reverse transcription-polymerase chain reaction of inducible NO synthase mRNA showed that ERK1/2 and p38 controlled IL-1beta-induced islet inducible NO synthase expression at the transcriptional level. Hyperosmolarity caused phosphorylation of Elk-1, activation transcription factor 2, and heat shock protein 25 and activation of ERK1/2 and p38 in islets comparable to that induced by IL-1beta but did not lead to NO synthesis. Inhibition of p38 but not of ERK1/2 attenuated IL-1beta-mediated inhibition of glucose-stimulated insulin release. We conclude that ERK1/2 and p38 activation is necessary but not sufficient for IL-1beta-mediated beta-cell NO synthesis and that p38 is involved in signaling of NO-independent effects of IL-1beta in beta-cells.

Animals↗