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Nonbioluminescent strains of Photobacterium phosphoreum produce the cell-to-cell communication signal N-(3-Hydroxyoctanoyl)homoserine lactone.

Bioluminescence is a common phenotype in marine bacteria, such as Vibrio and Photobacterium species, and can be quorum regulated by N-acylated homoserine lactones (AHLs). We extracted a molecule that induced a bacterial AHL monitor (Agrobacterium tumefaciens NT1 [pZLR4]) from packed cod fillets, which spoil due to growth of Photobacterium phosphoreum. Interestingly, AHLs were produced by 13 nonbioluminescent strains of P. phosphoreum isolated from the product. Of 177 strains of P. phosphoreum (including 18 isolates from this study), none of 74 bioluminescent strains elicited a reaction in the AHL monitor, whereas 48 of 103 nonbioluminescent strains did produce AHLs. AHLs were also detected in Aeromonas spp., but not in Shewanella strains. Thin-layer chromatographic profiles of cod extracts and P. phosphoreum culture supernatants identified a molecule similar in relative mobility (Rf value) and shape to N-(3-hydroxyoctanoyl)homoserine lactone, and the presence of this molecule in culture supernatants from a nonbioluminescent strain of P. phosphoreum was confirmed by high-performance liquid chromatography-positive electrospray high-resolution mass spectrometry. Bioluminescence (in a non-AHL-producing strain of P. phosphoreum) was strongly up-regulated during growth, whereas AHL production in a nonbioluminescent strain of P. phosphoreum appeared constitutive. AHLs apparently did not influence bioluminescence, as the addition of neither synthetic AHLs nor supernatants delayed or reduced this phenotype in luminescent strains of P. phosphoreum. The phenotypes of nonbioluminescent P. phosphoreum strains regulated by AHLs remains to be elucidated.

4-Butyrolactone↗

Identification of genes related to mechanical stress in human periodontal ligament cells using microarray analysis.

BACKGROUND AND OBJECTIVE: Differential expression of genes in human periodontal ligament (PDL) under mechanical stress, such as orthodontic force, is thought to be involved in the remodeling of PDL cells and periodontal tissues. However, little is known about the genes expressed in PDL cells under mechanical stress. MATERIAL AND METHODS: We employed microarray analysis to assess, in a comprehensive manner, the gene expression profiles in PDL cells compressed by a static force using an in vitro three-dimensional culture system. Six genes were selected and validated by quantitative real-time polymerase chain reaction analysis, consistent with the microarray data. RESULTS: The microarray data revealed that 108 of 30,000 genes tested were differentially expressed by mechanical force loading. Among them, 85 genes were up-regulated by mechanical stress, while 23 genes were down-regulated, judging by the thresholds of a two-fold increase/decrease compared with the controls. Thirty-two of the up-regulated and eight of the down-regulated genes, well-characterized in protein function, were involved in numerous biological processes including cell communication, cell signaling, cell cycle, stress response, and calcium release. However, several genes differentially expressed in our microarray data have not been well defined as stress-response molecules. CONCLUSION: Our microarray is the first to show the gene profile in PDL cells caused by mechanical stress; however, further studies to clarify the physiological function of these molecules in PDL cells are required.

Calcium↗

Expression and functional characterization of the serine protease inhibitor neuroserpin in endocrine cells.

Serine proteases play essential roles in a wide variety of cellular processes in endocrine cells. There is a growing interest in the roles of serine protease inhibitors, or serpins, as key regulators of their activity. We have cloned two neuroserpin cDNAs from a rat pituitary cDNA library and confirmed tissue plasminogen activator as a potential target for this inhibitor. We show that neuroserpin transcripts are expressed by endocrine cells in the adrenal and pituitary glands and that immunoreactive neuroserpin is stored in densely cored secretory granules in these cells. Overexpression of neuroserpin in an anterior pituitary corticotroph cell line results in the extension of neurite-like processes, suggesting that neuroserpin may play a role in cell communication, cell adhesion, and/or cell migration.

Adrenal Glands↗

New placental factors: Between implantation and inflammatory reaction.

The trophoblast invasion is a dynamic changes in cell-cell and cell matrix interaction and it create in the endometrio a reaction similar to the inflammatory reaction. In the recent past most of the investigator in this field had focus on the mediator of this process especially on cytokine and on vasoactive agent. We have studied the inducible isoform of nitric oxide synthase (iNOS), adrenomedullin (AM), fatty acid synthase (FAS), and S-100 protein. Several evidences had lead our choice: the two first factors are essentially local mediator of vasospasm which works in synergy during the inflammatory reaction, but the AM I addition stimulates the DNA synthesis and cell proliferation of Swiss 3T3 fibroblasts, induces the cell cycle progression from G0 to G1 phase and the malignant cell lines express AM mRNA and mRNA for its receptor and the pattern of distribution of ir-AM and its mRNA during murine development is similar to that of other growth factors (TGFb isoforms, PDGF, FGF, IGF 1 and 2); Fatty acid synthase (FAS) is a major biosynthetic enzyme of intracellular fatty acid synthesis, its expression and activity is enhanced in proliferative tissues, such as malignant cells; S-100 is an acid calcium binding protein present in central nervous system where it is in mainly concentrated in the glial cells, astrocytes, Schwann cells, and neurons. It regulates several functions such as cell-cell communication, cell growth, cell structure, energy metabolism, contraction, and intracellular signal transduction. Recently the appearance in blood of S100 in preterm infants has been used as a marker of brain damage in perinatal period. The iNOS, AM, S100 and FAS are localized in both decidual and trophoblastic cells in early pregnancy, but we found that changes in iNOS, S100, AM, and FAS are different in spontaneous abortion (SA) versus voluntary pregnancy termination (VPT). Considering the VPT as non pathologic pregnancy, this difference may reflect functional modifications of placental tissues. Our result shown that AM and NO are increased in VPT vs SA showing that when the process of implantation is on course we found an higher expression that we have during the inflammatory reaction. [Table: see text]

Abortion, Induced↗

Protein phosphatases: function and regulation.

Thousands of proteins are expressed in a typical mammalian cell, of which a third are now thought to contain covalently bound phosphate. About 200 protein kinases and 100 protein phosphatases have already been identified. Whereas the classification, properties and regulation of protein kinases are largely studied, the information about the protein phosphatase is far to be completed. Relatively better is studied a group of serine/treonine protein phosphatases. It has been discovered that besides the control of the key enzymes of metabolic pathways, these phosphatases participate in the regulation of gene transcription and cell division in eukariots. In addition, the identification of tyrosine protein phosphatases points to a novel intracellular signalling pathways, controlling cell-cell communication, cell proliferation and signal transduction of receptors for peptide hormones and growth factors. The recent progress in these families of protein phosphatases is the topic of present review.

Animals↗

Involvement of nerve growth factor in the ovulatory cascade: trkA receptor activation inhibits gap junctional communication between thecal cells.

Activation of trkA, the nerve growth factor (NGF) tyrosine kinase receptor, has been recently implicated in the process of mammalian ovulation. During the hour preceding follicular rupture, a marked increase in trkA and NGF gene expression occurs in thecal-interstitial cells of the ovary. Immunoneutralization of NGF actions or pharmacological blockade of trkA transducing activity inhibits ovulation, suggesting that activation of the NGF-trkA complex in nonneural cells of the periovulatory follicle is a physiological component of the ovulatory cascade. As thecal cells of Graafian follicles are functionally coupled by gap junctions, and the ovulatory rupture requires dissociation of thecal cell-cell communication, we sought to determine whether NGF affects the integrity of this communication. We now report that NGF-induced activation of trkA receptors in isolated ovarian thecal cells disrupts cell to cell communication by affecting the functional integrity of gap junctions. Bovine thecal cells expressing trkA receptors, but not cells lacking the receptors, respond to NGF with a reduction in the transfer of calcein, a fluorescent dye that passes through gap junctions. This effect was associated with a rapid (10-30 min) increase in serine phosphorylation of connexin-43, the main protein constituent of gap junctions in the ovary. The reduction in dye transfer was not observed when the cells were exposed to epidermal growth factor or other neurotrophins, including neurotrophin 3, neurotrophin 4, and brain-derived neurotrophic factor. Thus, cell-specific activation of trkA receptors in periovulatory follicles may provide one of the signals involved in inducing the cellular dissociation of the follicular wall that precedes ovulatory rupture.

Animals↗

Induction of intercellular communications in epithelial cell cultures.

A popular criterion of cell-cell communication in tissue cultures is dye coupling: the ability of the injected fluorescent dye of low molecular weight to be transferred from one cell to another. We report about a new factor which induces cell-to-cell dye coupling in previously uncoupled epithelial sheets. Paradoxically it is the standard fluorescent microscopy itself (that is, blue light of 320- to 480-nm wavelength) which induces rapid morphological alterations of cell culture followed by the transfer of fluorescent dye from one cell to another. Thus monitoring cell-cell dye coupling by fluorescent microscopy may itself induce the dye coupling in previously uncoupled epithelial cells.

Animals↗

Hoxb-5 expression in the developing mouse lung suggests a role in branching morphogenesis and epithelial cell fate.

Hoxb-5 is one of the few homeobox genes strongly expressed in the developing mouse lung. To explore the hypothesis that Hoxb-5 acts to regulate epithelial cell fate and branching morphogenesis in the developing lung, we studied the temporal, spatial, and cell-specific expression of Hoxb-5 from gestational day (d) 13.5 to postnatal day (P) 2. Immunocytochemistry demonstrated regional localization of Hoxb-5 protein to developing conducting airways and surrounding mesenchyme. The cellular expression pattern changed from diffusely positive nuclei of mesenchymal cells on d13.5 to become more localized to nuclei of subepithelial fibroblasts and some adjacent columnar and cuboidal epithelial cells on d14.5. After d14.5, Hoxb-5 protein expression continued to decrease in mesenchymal cells distal from developing airways, but persisted in fibroblasts underlying conducting airways. Hoxb-5 protein expression persisted in nuclei of columnar and cuboidal epithelial cells on d16.5 and d17.5, with expression in low cuboidal epithelial cells as well from d17.5 to P2. Western blot analysis showed temporal and quantitative changes in Hoxb-5 protein expression with peak expression on d14.5-15.5. We conclude that Hoxb-5 protein is developmentally regulated in a temporal, spatial, and cell-specific manner throughout the pseudoglandular, canalicular, and terminal saccular periods of lung development in the mouse. This localization and expression pattern suggests that Hoxb-5 may influence branching morphogenesis, cell-cell communication, cell fate, and differentiation of conducting airway epithelia.

Animals↗

T-/H-cadherin (CDH13): a new marker for differentiating podocytes.

Cadherin molecules are known to be involved in various biological processes other than cell adhesion such as morphogenesis, cell-cell communication, cell recognition or cell signalling. While the classical cadherin molecule is characterized by an extracellular moiety, a transmembrane region and a variable cytoplasmic domain, T-/H-cadherin differs from this pattern due to the absence of a transmembrane region and a cytoplasmic domain, respectively. Its extracellular moiety is bound to the apical cell membrane by a glycosyl-phosphatidyl-inositol anchor and localized to lipid raft domains. As its molecular function and expression pattern is still not fully understood, we used a newly generated anti-T-/H-cadherin antiserum to study immunohistochemically the expression of T-/H-cadherin during the differentiation of foetal human glomeruli. At the early capillary loop stage a strong apical signal comes up for visceral epithelial cells of Bowman's capsule, which begin to differentiate towards podocytes. At the advanced capillary loop stage, when podocytes have become part of the glomerular filtration barrier, the expression pattern, however, becomes more distinct and most likely restricted to the foot processes of the podocytes. We thus postulate a functional role of T-/H-cadherin for the differentiation of the podocytes and the formation of the glomerular capillary network.

Adult↗

The S100 protein family: history, function, and expression.

The S100 family of calcium binding proteins contains approximately 16 members each of which exhibits a unique pattern of tissue/cell type specific expression. Although the distribution of these proteins is not restricted to the nervous system, the implication of several members of this family in nervous system development, function, and disease has sparked new interest in these proteins. We now know that the original two members of this family, S100A1 and S100B, can regulate a diverse group of cellular functions including cell-cell communication, cell growth, cell structure, energy metabolism, contraction and intracellular signal transduction. Although some members of the family may function extracellularly, most appear to function as intracellular calcium-modulated proteins and couple extracellular stimuli to cellular responses via interaction with other cellular proteins called target proteins. Interaction of these proteins with target proteins appear to involve cysteine residues (one in S100A1 and two in S100B), as well as a stretch of 13 amino acids, in the middle of the molecule called the linker region, which connects the two EF-hand calcium binding domains. In addition to the amino acid sequence and secondary structures of these proteins, the structures of the genes encoding these proteins are highly conserved. Studies on the expression of these proteins have demonstrated that a complex mixture of transcriptional and postranscriptional mechanisms regulate S100 expression. Further analysis of the function and expression of these proteins in both nervous and nonnervous tissues will provide important information regarding the role of altered S100 expression in nervous system development, function and disease.

Amino Acid Sequence↗

Application of a methylation gene panel by quantitative PCR for lung cancers.

Detection of lung cancer at early stages could potentially increase survival rates. One promising approach is the application of suitable lung cancer-specific biomarkers to specimens obtained by non-invasive methods. Thus far, clinically useful biomarkers that have high sensitivity have proven elusive. Certain genes, which are involved in cellular pathways such as signal transduction, apoptosis, cell to cell communication, cell cycles and cytokine signaling are down-regulated in cancers and may be considered as potential tumor suppressor genes. Aberrant promoter hypermethylation is a major mechanism for silencing tumor suppressor genes in many kinds of human cancers. Using quantitative real time PCR, we tested 11 genes (3-OST-2, RASSF1A, DcR1, DcR2, P16, DAPK, APC, ECAD, HCAD, SOCS1, SOCS3) for levels of methylation within their promoter sequences in non-small cell lung cancers (NSCLC), adjacent non-malignant lung tissues, in peripheral blood mononuclear cells (PBMC) from cancer free patients, in sputum of cancer patients and controls. Of all the 11 genes tested 3-OST-2 showed the highest levels of promoter methylation in tumors combined with lowest levels of promoter methylation in control tissues. 3-OST-2 followed by, RASSF1A showed increased levels of methylation with advanced tumor stage (P<0.05). Thus, quantitative analysis of 3-OST-2 and RASSF1A methylation appears to be a promising biomarker assay for NSCLC and should be further explored in a clinical study. Our preliminary data on the analysis of sputum DNA specimens from cancer patients further support these observations.

Adenocarcinoma↗

A neurogenomics approach to gene expression analysis in the developing brain.

Secreted and transmembrane proteins provide critical functions in the signaling networks essential for neurogenesis. We used a genetic signal sequence gene trap approach to isolate 189 genes expressed during development in e16.5 whole head, e16.5 hippocampus and e14.5 cerebellum. Gene ontology programs were used to classify the genes into respective biological processes. Four major classes of biological processes known to be important during development were identified: cell communication, cell physiology processes, metabolism and morphogenesis. We used in situ hybridization to determine the temporal and spatial patterns of gene expression in the developing brain using this set of probes. The results demonstrate that gene expression patterns can highlight potential gene functions in specific brain regions. We propose that combining bioinformatics with the gene expression pattern is an effective strategy to identify genes that may play critical roles during brain development.

Animals↗