Tyrosine nitration as mediator of cell death

Nitrotyrosine is used as a marker for the production of peroxynitrite and other reactive nitrogen species. For over 20 years the presence of nitrotyrosine was associated with cell death in multiple pathologies. Filling the gap between correlation and causality has proven to be a difficult task. Here, we discuss the evidence supporting tyrosine nitration as a specific posttranslational modification participating in the induction of cell death signaling pathways.     

Extracellular proteins as enterobacterial thermometers

Biological thermometers are cellular components or structures which sense increasing temperatures, interaction of the thermometer and the thermal stress bringing about the switching-on of inducible responses, with gradually enhanced levels of response induction following gradually increasing temperatures. In enterobacteria, for studies of such thermometers, generally induction of heat shock protein (HSP) synthesis has been examined, with experimental studies aiming to establish (often indirectly) how the temperature changes which initiate HSP synthesis are sensed; numerous other processes and responses show graded induction as temperature is increased, and how the temperature changes which induce these are sensed is also of interest. Several classes of intracellular component and structure have been proposed as enterobacterial thermometers, with the ribosome and the DnaK chaperone being the most favoured, although for many of the proposed intracellular thermometers, most of the evidence for their functioning in this way is indirect. In contrast to the above, the studies reviewed here firmly establish that for four distinct stress responses, which are switched-on gradually as temperature increases, temperature changes are sensed by extracellular components (extracellular sensing components, ESCs) i.e. there is firm and direct evidence for the occurrence of extracellular thermometers. All four thermometers described here are proteins, which appear to be distinct and different from each other, and on sensing thermal stress are activated by it to four distinct extracellular induction components (EICs), which interact with receptors on the surface of organisms to induce the appropriate responses. It is predicted that many other temperature-induced processes, including the synthesis of HSPs, will be switched-on following the activation of similar extracellular thermometers by thermal stimuli.     

G proteins as drug targets

The structure and function of heterotrimeric G protein subunits is known in considerable detail. Upon stimulation of a heptahelical receptor by the appropriate agonists, the cognate G proteins undergo a cycle of activation and deactivation; the α-subunits and the βγ-dimers interact sequentially with several reaction partners (receptor, guanine nucleotides and effectors as well as regulatory proteins) by exposing appropriate binding sites. For most of these domains, low molecular weight ligands have been identified that either activate or inhibit signal transduction. These ligands include short peptides derived from receptors, G protein subunits and effectors, mastoparan and related insect venoms, modified guanine nucleotides, suramin analogues and amphiphilic cations. Because compounds that act on G proteins may be endowed with new forms of selectivity, we propose that G protein subunits may therefore be considered as potential drug targets. Received 18 September 1998; received after revision 6 November 1998; accepted 11 November 1998     

Cisplatin-induced genes as potential markers for thyroid cancer

Despite the uncontested role of p53 in cycle arrest/cell death after cisplatin treatment, to date the question whether wild-type p53 confers a resistant or sensitive status on the cell is still a matter of debate. Isogenic and isophenotypic human thyroid papillary carcinoma cell line variants for p53 differently expressed cycle genes after cisplatin treatment. Seven genes (CDC6-related protein, CCNC, GAS1, TFDP2, MAPK10/JNK3, WEE1, RPA1) selected after expression on an Atlas human cell cycle array were analyzed by quantitative real-time PCR. While cisplatin treatment increased their expression in p53 wild-type cells it decreased it in cells with inactivated p53 and had no or less effect on cells with mutated p53. These results show that in a well-defined system, different alterations of p53 can lead to a different regulation of genes and hence to either resistance or sensitivity to cisplatin. Moreover for the first time, MAPK10/JNK3 was identified in human thyroid cells and tissue. Four of the genes (CDC6-related protein, CCNC, GAS1 and TFDP2) were decreased in human papillary carcinoma tissues. Relevance of these genes (especially a decrease in GAS1 in thyroid papillary carcinoma) in various malignant pathologies has already been shown. These genes may be explored as new markers in advanced thyroid cancer such as metastatic and anaplastic forms displaying p53 alterations.Received 26 July 2004; received after revision 14 September 2004; accepted 26 October 2004     

Functions of disordered regions in mammalian early base excision repair proteins

Reactive oxygen species, generated endogenously and induced as a toxic response, produce several dozen oxidized or modified bases and/or single-strand breaks in mammalian and other genomes. These lesions are predominantly repaired via the conserved base excision repair (BER) pathway. BER is initiated with excision of oxidized or modified bases by DNA glycosylases leading to formation of abasic (AP) site or strand break at the lesion site. Structural analysis by experimental and modeling approaches shows the presence of a disordered segment commonly localized at the N- or C-terminus as a characteristic signature of mammalian DNA glycosylases which is absent in their bacterial prototypes. Recent studies on unstructured regions in DNA metabolizing proteins have indicated their essential role in interaction with other proteins and target DNA recognition. In this review, we have discussed the unique presence of disordered segments in human DNA glycosylases, and AP endonuclease involved in the processing of glycosylase products, and their critical role in regulating repair functions. These disordered segments also include sites for posttranslational modifications and nuclear localization signal. The teleological basis for their structural flexibility is discussed.     

Glycine-rich proteins as structural components of plant cell walls

Glycine-rich proteins (GRPs) have been found in the cell walls of many higher plants and form a third group of structural protein components of the wall in addition to extensins and proline-rich proteins. The primary sequences of GRPs contain more than 60% glycine. GRPs are localized mainly in the vascular tissue of the plant, and their coding genes provide an excellent system to analyze the molecular basis of vascular-specific gene expression. In French bean, the major cell wall GRP has been localized at the ultrastructural level in the modified primary cell wall of protoxylem. Immunological studies showed that it forms a major part of these highly extensible and specialized cell walls. Specific digestion of GRP1.8 from bean by collagenase suggests that it shares structural similarities with collagen. The protein is synthesized by living protoxylem cells as well as xylem parenchyma cells. After cell death, GRPs are exported from neighboring xylem parenchyma cells to the protoxylem wall, a rare example of protein transport between cells in plants. We propose that GRPs are part of a repair system of the plant during the stretching phase of protoxylem.     

Role of FGFRL1 and other FGF signaling proteins in early kidney development

The mammalian kidney develops from the ureteric bud and the metanephric mesenchyme. In mice, the ureteric bud invades the metanephric mesenchyme at day E10.5 and begins to branch. The tips of the ureteric bud induce the metanephric mesenchyme to condense and form the cap mesenchyme. Some cells of this cap mesenchyme undergo a mesenchymal-to-epithelial transition and differentiate into renal vesicles, which further develop into nephrons. The developing kidney expresses Fibroblast growth factor (Fgf)1, 7, 8, 9, 10, 12 and 20 and Fgf receptors Fgfr1 and Fgfr2. Fgf7 and Fgf10, mainly secreted by the metanephric mesenchyme, bind to Fgfr2b of the ureteric bud and induce branching. Fgfr1 and Fgfr2c are required for formation of the metanephric mesenchyme, however the two receptors can substitute for one another. Fgf8, secreted by renal vesicles, binds to Fgfr1 and supports survival of cells in the nascent nephrons. Fgf9 and Fgf20, expressed in the metanephric mesenchyme, are necessary to maintain survival of progenitor cells in the cortical region of the kidney. FgfrL1 is a novel member of the Fgfr family that lacks the intracellular tyrosine kinase domain. It is expressed in the ureteric bud and all nephrogenic structures. Targeted deletion of FgfrL1 leads to severe kidney dysgenesis due to the lack of renal vesicles. FgfrL1 is known to interact mainly with Fgf8. It is therefore conceivable that FgfrL1 restricts signaling of Fgf8 to the precise location of the nascent nephrons. It might also promote tight adhesion of cells in the condensed metanephric mesenchyme as required for the mesenchymal-to-epithelial transition.     

Ubiquitin-proteasome pathway as a primary defender against TRAIL-mediated cell death

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces apoptotic cell death as well as expression of proinflammatory genes such as CXCL8 in malignant human astrocytoma cells. However, the molecular mechanisms that determine the fate of cells are not yet understood. The ubiquitin (Ub)-proteasome pathway regulates a wide range of cellular functions through degradation of various regulatory proteins; given this, we hypothesized that this pathway may play a central role in TRAIL-mediated signaling. We demonstrate here that inhibition of the Ub-proteasome pathway enhanced TRAIL-mediated cell death of human astrocytoma CRT-MG cells within hours by blocking degradation of active caspase-8 and -3. Proteasome inhibitors suppressed TRAIL-mediated activation of NF-B; however, inhibition of the NF-B pathway alone was not sufficient to enhance TRAIL-mediated cell death. Collectively, these results suggest that the Ub-proteasome pathway may play an important role as an antiapoptotic surveillance system by eliminating activated caspases as well as mediating NF-B-dependent signals.Received 30 December 2003; received after revision 9 February 2004; accepted 13 February 2004     

Cell division and cell death during regression of the chick embryo Müllerian ducts.

In 9-day-old chick embryos, decreased DNA synthesis and enhanced necrosis were observed in a defined area of the right female Müllerian ducts, supporting the idea of the existence of a regression process in this organ. In the male ducts, decreased DNA synthesis and a low level of necrosis were present all over the studied portion of the organ.     

Bacteria as cell surface markers on normal and malignant mammalian cells

Zusammenfassung Bakterien haften an der Oberfläche kultivierter Säugetierzellen in charakteristischer Weise. An Krebszellen haften sie nicht nur am Vorderteil, sondern auch an der Zelloberfläche und in Kernnähe.

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I thank the Royal Society for some of the equipment used in this research, the Nuffield Foundation for the award of a fellowship, and Prof.I. M. Goodbody for facilities in the Department of Zoology, University of the West Indies, Jamaica, while on sabbatical leave of absence from the University of Glasgow. I am also indebted toJ. B. Cowey, A. S. G. Curtis andD. R. Newth, of Glasgow University for their criticisms and advice, and to Miss E. Cray for assistance in the experiments.