Double bond isomers of cloprostenol

The syntheses of five compounds isomeric with the potent luteolytic agent cloprostenol are described. These are the trans-delta 5, cis and trans-delta 4, cis-delta 6 (methyl ester) and cis-delta 13 analogues. The cis-delta 4 isomer is as potent as cloprostenol as a luteolytic agent in hamsters but the others are markedly less so.     

Diatom Phytochromes Reveal the Existence of Far-Red-Light-Based Sensing in the Ocean

The absorption of visible light in aquatic environments has led to the common assumption that aquatic organisms sense and adapt to penetrative blue/green light wavelengths but show little or no response to the more attenuated red/far-red wavelengths. Here, we show that two marine diatom species, Phaeodactylum tricornutum and Thalassiosira pseudonana, possess a bona fide red/far-red light sensing phytochrome (DPH) that uses biliverdin as a chromophore and displays accentuated red-shifted absorbance peaks compared with other characterized plant and algal phytochromes. Exposure to both red and far-red light causes changes in gene expression in P. tricornutum, and the responses to far-red light disappear in DPH knockout cells, demonstrating that P. tricornutum DPH mediates far-red light signaling. The identification of DPH genes in diverse diatom species widely distributed along the water column further emphasizes the ecological significance of far-red light sensing, raising questions about the sources of far-red light. Our analyses indicate that, although far-red wavelengths from sunlight are only detectable at the ocean surface, chlorophyll fluorescence and Raman scattering can generate red/far-red photons in deeper layers. This study opens up novel perspectives on phytochrome-mediated far-red light signaling in the ocean and on the light sensing and adaptive capabilities of marine phototrophs.     

pH dependence of formation of a partially unfolded state of a Lys 73 --> His variant of iso-1-cytochrome c: implications for the alkaline conformational transition of cytochrome c

The alkaline conformational transition of a lysine 73 --> histidine variant of iso-1-cytochrome c has been studied. The transition has been monitored at 695 nm, a band sensitive to the presence of the heme-methionine 80 bond, at the heme Soret band which is sensitive to the nature of the heme ligand, and by NMR methods. The guanidine hydrochloride dependence of the alkaline conformational transition has also been monitored. The histidine 73 protein has an unusual biphasic alkaline conformational transition at both 695 nm and the heme Soret band, consistent with a three-state process. The conformational transition is fully reversible. An equilibrium model has been developed to account for this behavior. With this model, it has been possible to obtain the acid constant for the trigger group, pK(H), of the low-pH phase from the equilibrium data. A pK(H) value of 6.6 +/- 0.1 in H(2)O was obtained, consistent with a histidine acting as the trigger group. The NMR data for the low-pH phase of the alkaline conformational transition are consistent with an imidazole ligand replacing Met 80. For the high-pH phase of the biphasic alkaline transition, the NMR data are consistent with lysine 79 being the heme ligand. Guanidine hydrochloride m values of 1.67 +/- 0.08 and 1.1 +/- 0.2 kcal mol(-1) M(-1) were obtained for the low- and high-pH phases of the biphasic alkaline transition of the histidine 73 protein, respectively, consistent with a greater structural disruption for the low-pH phase of the transition.     

Semi-quantitative RT-PCR analysis of photoregulated gene expression in marine diatoms

The low cell densities of diatoms and other phytoplankton in culture has precluded the use of classical RNA analysis techniques for routine studies of gene expression in large numbers of samples. This has seriously hampered studies of the basic biology of such organisms. To circumvent this problem, we have developed a high-throughput semi-quantitative RT-PCR-based protocol and used it to monitor expression of a gene encoding a fucoxanthin, chlorophyll a/c-binding protein (FCP) in the centric planktonic diatom Thalassiosira weissflogii. Analysis of FCP gene expression in dark-adapted diatom cultures revealed that mRNA levels increase 5- to 6-fold in response to white light irradiation and peak around 6 to 8 h. To determine the photoreceptors involved in this response action spectra of FCP gene expression were determined using the Okazaki large spectrograph. Responses consistent with the presence of cryptochrome-, rhodopsin- and phytochrome-type receptors could be detected. The apparent presence of phytochrome-mediated responses is of particular interest given the low fluences of red and far-red light wavelengths in the marine environment.     

An investigation into the effects of stimulators of fluid production on Locusta Malpighian tubule intracellular elemental composition

The intracellular elemental concentrations of Na, K, P, S, Cl and Mg in the type 1 cells of Malpighian tubules of Locusta migratoria L. have been measured using electron probe X-ray microanalysis. The effects of in vitro stimulation with 1 mM cAMP and corpora cardiaca extract (CC-extract) on the elemental concentrations have been quantified. The distribution of elements, particularly Na, K and Cl is not homogeneous in control cells, and concentration gradients exist within the cytoplasm. Dibutyryl-cAMP (DB-cAMP) caused a decrease in [K]_i without disrupting the gradient which increased from the basal to the apical surface, the apical [Na]_i was increased as was the [Cl]_i. In contrast, in vitro application of CC-extract did not cause changes to the intracellular elemental composition as compared with control cells These data are consistent with the interpretation that exogenous cAMP only partially activated the full stimulatory response of Malpighian tubule cells observed with CC-extract. The changes observed in the density and elemental composition of the `dark bodies' in response to DB-cAMP and CC-extract stimulation suggest that these structures have a role in the ionic economy of Malpighian tubule cells. Accepted: 6 April 1999     

Insect thermal tolerance: what is the role of ontogeny,ageing and senescence?

Temperature has dramatic evolutionary fitness consequences and is therefore a major factor determining the geographic distribution and abundance of ectotherms. However, the role that age might have on insect thermal tolerance is often overlooked in studies of behaviour, ecology, physiology and evolutionary biology. Here, we review the evidence for ontogenetic and ageing effects on traits of high- and low-temperature tolerance in insects and show that these effects are typically pronounced for most taxa in which data are available. We therefore argue that basal thermal tolerance and acclimation responses (i.e. phenotypic plasticity) are strongly influenced by age and/or ontogeny and may confound studies of temperature responses if unaccounted for. We outline three alternative hypotheses which can be distinguished to propose why development affects thermal tolerance in insects. At present no studies have been undertaken to directly address these options. The implications of these age-related changes in thermal biology are discussed and, most significantly, suggest that the temperature tolerance of insects should be defined within the age-demographics of a particular population or species. Although we conclude that age is a source of variation that should be carefully controlled for in thermal biology, we also suggest that it can be used as a valuable tool for testing evolutionary theories of ageing and the cellular and genetic basis of thermal tolerance.