From the top down: towards a predictive biology of signalling networks

True quantitative and predictive biology requires the ability to interpret increasingly complex datasets in new ways to reveal underlying functional interactions. This has come a step closer with two recent articles that describe a 'top-down' modelling approach to reconstructing functional networks from microarray data.     

Cadmium: tissue distribution and binding protein induction in the painted turtle, Chrysemys picta

The freshwater painted turtle, Chrysemys picta, was used to investigate (a) the distribution of an injected dose of ¹⁰⁹Cd in tissues over a period of 192 h (8 days) and (b) the effect of non-isotopic cadmium injection on tissue metal-binding protein levels. Cadmium is cleared from the blood with 9% remaining in the circulation at 192 h. ¹⁰⁹Cd is found in all tissues, but is accumulated preferentially in liver, kidney, pancreas, and gastrointestinal tract. The liver is the primary site of Cd accumulation, accounting for 46.4% of the injected dose by 192 h and the highest Cd concentration (cpm/mg tissue). Steroidogenic tissues and the oviduct accumulate significant amounts of ¹⁰⁹Cd and the isotope is present in yolk. An increase in tissue metal-binding protein level after non-isotopic CdCl₂ injection is consistent with ¹⁰⁹Cd distribution, in that metal-binding protein concentration after CdCl₂ injection is highest in liver, followed by pancreas and kidney with low, but with significant levels of cadmium-binding protein in gonads and steroid target organs. We conclude that the liver is the major site of storage after a single injection of isotopic cadmium and induction of a metal-binding protein may be an adaptive response to exposure to cadmium.     

Changes in Nuclear Receptor and Vitellogenin Gene Expression in Response to Steroids and Heavy Metal in Caenorhabditis elegans

To gain basic understanding of the reproductive and developmental effects of endocrine disrupting chemicals in invertebrates, we have used C. elegans as an animal model. The completion of the C. elegans genome sequence brings to bear microarray analysis as a tool for these studies. We previously showed that the C. elegans genome was responsive to vertebrate steroid hormones, and changes in gene expression of traditional biomarkers used in environmental studies were detected; i.e., vitellogenin (vtg), cytochrome P450 (cyp450), glutathione-S-transferase (gst) and heat shock proteins (hsp). The data were interpreted to suggest that exogenous lipophilic compounds can be metabolized via cytochrome P450 proteins, and that the resulting metabolites can bind to members of the Nuclear Receptor (NR) class of proteins and regulate gene expression. In the present study, using DNA microarrays, we examined the pattern of gene expression after progesterone (10(-5), 10(-7) M), estradiol (10(-5) M), cholesterol (10(-9) M) and cadmium (0.1, 1 and 10 μM) exposure, with special attention to the members of NRs. Of approximately 284 NRs in C. elegans, expression of 25 NR genes (representing 9% of the total NRs in C. elegans) was altered after exposure to steroids. Of note, each steroid activated or inhibited different subsets of NR genes, and only estradiol regulated NR genes implicated in neurogenesis. These results suggest that NRs respond to a variety of exogenous steroids, which regulate important metabolic and developmental pathways. The response of the C. elegans genome to cholesterol and cadmium was analyzed in more detail. Cholesterol is a probable precursor to signaling molecules that may interact with NRs and we focused on expression of genes related to lipid metabolism (cyp450), transport and storage (i.e., vitellogenin). Worms exposed to cadmium respond principally by activating the expression of genes encoding stress-responsive proteins, such as mtl-2 and cdr-1, and no significant changes in expression of NRs or vtg genes were observed. The possible implications of these results with regard to the evolution of steroid receptors, endocrine disruption and the role of vitellogenin as a lipid transporter are discussed.     

Comparing antigen-independent mechanisms of T cell regulation

Key features of the kinetics of T lymphocyte proliferative responses are remarkably insensitive to the nature of the antigenic stimulus. This consistency suggests the presence of an antigen-independent mechanism regulating T cell clonal expansion. Knowledge of such a mechanism could allow us to modulate T helper cell (CD4+) and cytotoxic T cell (CD8+) responses more effectively. Using a simple mathematical model of T cell proliferation and death, we investigate a variety of plausible mechanisms and compare the model predictions to experimental data from the literature. We find that irrespective of the details of the mechanism, rates of apoptosis must progressively increase to control a T cell response. If apoptosis is mediated by cell-cell contact this alone is sufficient to regulate both (CD4+) and (CD8+) T cell responses. Proliferation of both T cell subsets can also be regulated by an internal programme, by cytokine signalling, or by an APC-mediated route. To regulate (CD8+) T cells these mechanisms must change both apoptosis and division rates, and this change must occur with time not division number.     

Circadian rhythms in human muscular efficiency: continuous physical exercise versus continuous rest. A crossover study

This study deals with the influence of time of day on neuromuscular efficiency in competitive cyclists during continuous exercise versus continuous rest. Knee extension torque was measured in ultradistance cyclists over a 24h period (13:00 to 13:00 the next day) in the laboratory. The subjects were requested to maintain a constant speed (set at 70% of their maximal aerobic speed obtained during a preliminary test) on their own bicycles, which were equipped with cyclosimulators. Every 4h, torque developed and myoelectric activity were estimated during maximal isometric voluntary contractions of knee extensors using an isokinetic dynamometer. Mesenteric temperature was monitored by telemetry. The same measures were also recorded while the subjects were resting awake until 13:00 the next day. During activity, torque changed within the 24h period (p < .005), with an acrophase at 19:10 and an amplitude of 7.8% around the mean of 70.7%. At rest, a circadian rhythm was observed in knee extensor torque (p < .05), with an acrophase at 19:30 and an amplitude of 6% around the mean of 92.3%. Despite the standardized conditions, the results showed that isometric maximal strength varied with time of day during both a submaximal exercise and at rest without prior exercise. The sine waves representing these two rhythms were correlated significantly. Although at rest the diurnal rhythm followed muscular activity (i.e., neurophysiological factors), during exercise, this rhythm was thought to stem more from fluctuations in the contractile state of muscle.     

A transformation procedure for recalcitrant tomato by addressing transgenic plant-recovery limiting factors

Agrobacterium tumefaciens technology is the battle horse for tomato genetic transformation. However, tomato varieties with low regeneration capacity are very difficult to transform. In the past, tomato transformation through Agrobacterium infection was focused on varieties capable of high regeneration yield, while successful transformation of low regenerable cultivars has not been reported. The genotype response to tissue culture conditions is believed to drive the frequency of regeneration of transgenic plant, whereas the capacity for cell proliferation could determine the transformation efficiency through this technology. The Campbell-28 cultivar is an example of constraints arising from a high morphogenetic potential with low conversion compared to normal plants. In the present work the roles that contribute to improved transgenic plant recovery from this recalcitrant variety were explored for factors like Agrobacterium concentration and antibiotics for bacterial removal and transformant selection. Analysis of the efficiency from independent transformation experiments revealed a more than twofold increase of transformant regeneration after selection on ammonium glufosinate compared to kanamycin selection, showing a transformation efficiency of 21.5%.