Spatial and temporal variation of photosynthesis in intertidal Mazzaella laminarioides (Bory)Fredericq (Rhodophyta, Gigartinales)

The red alga Mazzaella laminarioides is an economically important species with an extended latitudinal distribution along the Chilean coast. Its populations form mid-intertidal stands, several meters wide, and therefore are differentially exposed to environmental variables that result in temporal and spatial variability in productivity. We evaluated the effect of latitude and intertidal height on productivity by in situ measurement of photosynthetic performance. Daily and seasonal variations of O₂-evolution rate and maximal quantum yield (F _v/F _m) were determined in plants from the upper and lower intertidal zone at two localities 1500,km apart. Results suggest that plant responses were mainly affected by irradiation, temperature and desiccation. At local level, upper intertidal plants showed a reduced photosynthetic rate and quantum efficiency as compared to those displayed by plants from the lower intertidal, indicating their higher level of excitation energy acclimation. Stronger acclimation differences between upper and lower intertidal plants were observed in spring and summer. Differences in photosynthetic parameters between reproductive phases were recorded in autumn and winter, regardless of the position of the individuals in the intertidal zone. The effects of tidal elevation on seasonal patterns of photosynthesis were also influenced by latitude. Seasonal variation in photosynthetic efficiency was observed in plants from the northern population at both intertidal elevations, but only at the upper intertidal level in the southern population. This study shows that production variability in M. laminarioides results from differences in the intensity of environmental factors observed seasonally at local (intertidal) and latitudinal scales.     

PP2A regulates BCL-2 phosphorylation and proteasome-mediated degradation at the endoplasmic reticulum

Anti-apoptotic activity of BCL-2 is mediated by phosphorylation at the endoplasmic reticulum (ER), but how this phosphorylation is regulated and the mechanism(s) by which it regulates apoptosis are unknown. We purified macromolecular complexes containing BCL-2 from ER membranes and found that BCL-2 co-purified with the main two subunits of the serine/threonine phosphatase, PP2A. The association of endogenous PP2A and BCL-2 at the ER was verified by co-immunoprecipitation and microcystin affinity purification. Knock down or pharmacological inhibition of PP2A caused degradation of phosphorylated BCL-2 and led to an overall reduction in BCL-2 levels. We found that this degradation was due to the action of the proteasome acting selectively at the ER. Conversely, overexpression of PP2A caused elevation in endogenous BCL-2. Most importantly, we found that PP2A knock down sensitized cells to several classes of death stimuli (including ER stress), but this effect was abolished in a genetic background featuring knock in of a non-phosphorylatable BCL-2 allele. These studies support the hypothesis that PP2A-mediated dephosphorylation of BCL-2 is required to protect BCL-2 from proteasome-dependent degradation, affecting resistance to ER stress.     

Norepinephrine induces calcium spikes and proinflammatory actions in human hepatic stellate cells

Catecholamines participate in the pathogenesis of portal hypertension and liver fibrosis through alpha1-adrenoceptors. However, the underlying cellular and molecular mechanisms are largely unknown. Here, we investigated the effects of norepinephrine (NE) on human hepatic stellate cells (HSC), which exert vasoactive, inflammatory, and fibrogenic actions in the injured liver. Adrenoceptor expression was assessed in human HSC by RT-PCR and immunocytochemistry. Intracellular Ca2+ concentration ([Ca2+]i) was studied in fura-2-loaded cells. Cell contraction was studied by assessing wrinkle formation and myosin light chain II (MLC II) phosphorylation. Cell proliferation and collagen-alpha1(I) expression were assessed by [3H]thymidine incorporation and quantitative PCR, respectively. NF-kappaB activation was assessed by luciferase reporter gene and p65 nuclear translocation. Chemokine secretion was assessed by ELISA. Normal human livers expressed alpha(1A)-adrenoceptors, which were markedly upregulated in livers with advanced fibrosis. Activated human HSC expressed alpha(1A)-adrenoceptors. NE induced multiple rapid [Ca2+]i oscillations (Ca2+ spikes). Prazosin (alpha1-blocker) completely prevented NE-induced Ca2+ spikes, whereas propranolol (nonspecific beta-blocker) partially attenuated this effect. NE caused phosphorylation of MLC II and cell contraction. In contrast, NE did not affect cell proliferation or collagen-alpha1(I) expression. Importantly, NE stimulated the secretion of inflammatory chemokines (RANTES and interleukin-8) in a dose-dependent manner. Prazosin blocked NE-induced chemokine secretion. NE stimulated NF-kappaB activation. BAY 11-7082, a specific NF-kappaB inhibitor, blocked NE-induced chemokine secretion. We conclude that NE stimulates NF-kappaB and induces cell contraction and proinflammatory effects in human HSC. Catecholamines may participate in the pathogenesis of portal hypertension and liver fibrosis by targeting HSC.     

Gene Duplication and Positive Selection Explains Unusual Physiological Roles of the Relaxin Gene in the European Rabbit

The relaxin gene family is a group of genes involved in different physiological roles, most of them related to reproduction. In vertebrates the genes in this family are located in three separate chromosomal locations, and have been called relaxin family locus (RFL) A, B, and C. Among mammals the RFLA and RFLC are the most conserved as no gene copy-number variation has been observed thus far. The RFLB locus is also conserved on most mammals other than primates, where there are several gene gains and losses. Interestingly, the relaxin gene found on the RFLB locus in the European rabbit has acquired a novel role. In addition to the classical reproductive roles, this gene is expressed in tracheobronchial epithelial cells and its expression has been linked to squamous differentiation. We reconstructed the evolutionary history of the European rabbit RFLB locus using the tools of comparative genomics and molecular evolution. We found that the European rabbit possess a RFLB locus which is unique among mammals in that there are five tandemly arranged relaxin gene copies, which contrast with the single relaxin copy gene found in most mammals. In addition we also found that the ancestral pre-duplication gene was subject to the action of positive selection, and several amino acid sites were identified under the action of natural selection including the sites B12 and B13 which are part of the receptor recognition and binding site.     

Generalization of the dynamic clamp concept in neurophysiology and behavior

The idea of closed-loop interaction in in vitro and in vivo electrophysiology has been successfully implemented in the dynamic clamp concept strongly impacting the research of membrane and synaptic properties of neurons. In this paper we show that this concept can be easily generalized to build other kinds of closed-loop protocols beyond (or in addition to) electrical stimulation and recording in neurophysiology and behavioral studies for neuroethology. In particular, we illustrate three different examples of goal-driven real-time closed-loop interactions with drug microinjectors, mechanical devices and video event driven stimulation. Modern activity-dependent stimulation protocols can be used to reveal dynamics (otherwise hidden under traditional stimulation techniques), achieve control of natural and pathological states, induce learning, bridge between disparate levels of analysis and for a further automation of experiments. We argue that closed-loop interaction calls for novel real time analysis, prediction and control tools and a new perspective for designing stimulus-response experiments, which can have a large impact in neuroscience research.     

Gene turnover and differential retention in the relaxin/insulin-like gene family in primates

The relaxin/insulin-like gene family is related to the insulin gene family, and includes two separate types of peptides: relaxins (RLNs) and insulin-like peptides (INSLs) that perform a variety of physiological roles including testicular descent, growth and differentiation of the mammary glands, trophoblast development, and cell differentiation. In vertebrates, these genes are found on three separate genomic loci, and in mammals, variation in the number and nature of genes in this family is mostly restricted to the Relaxin Family Locus B. For example, this locus contains a single copy of RLN in platypus and opossum, whereas it contains copies of the INSL6, INSL4, RLN2 and RLN1 genes in human and chimp. The main objective of this research is to characterize changes in the size and membership composition of the RLN/INSL gene family in primates, reconstruct the history of the RLN/INSL genes of primates, and test competing evolutionary scenarios regarding the origin of INSL4 and of the duplicated copies of the RLN gene of apes. Our results show that the relaxin/INSL-like gene family of primates has had a more dynamic evolutionary history than previously thought, including several examples of gene duplications and losses which are consistent with the predictions of the birth-and-death model of gene family evolution. In particular, we found that the differential retention of relatively old paralogs played a key role in shaping the gene complement of this family in primates. Two examples of this phenomenon are the origin of the INSL4 gene of catarrhines (the group that includes Old World monkeys and apes), and of the duplicate RLN1 and RLN2 paralogs of apes. In the case of INSL4, comparative genomics and phylogenetic analyses indicate that the origin of this gene, which was thought to represent a catarrhine-specific evolutionary innovation, is as old as the split between carnivores and primates, which took place approximately 97 million years ago. In addition, in the case of the RLN1 and RLN2 genes of apes our phylogenetic trees and topology tests indicate that the duplication that gave rise to these two genes maps to the last common ancestor of anthropoid primates. All these genomic changes in gene complement, which are particularly prevalent among anthropoid primates, might be linked to the many physiological and anatomical changes found in this group. Given the various roles of members of the RLN/INSL-like gene family in reproductive biology, it might be that changes in this gene family are associated to changes in reproductive traits.     

A Kinetic Examination of Penicillin Acylase Stability in Water-organic Solvent Systems at Different Temperatures

We have studied the thermal stability of penicillin acylase from Streptomyces lavendulae in water-organic solvent monophasic systems at the range of temperatures between 40 and 60°C. We found a linear correlation between the log λP value of the solvent and the activation free energy for denaturation ( ΔG d ) at all temperatures tested. Thermodynamic analysis of the results indicates that solvents with log λP ≤-2.3 have protective effects, whereas solvents with log λP ≥-1.8 are deleterious for penicillin acylase.     

Recent Male-Mediated Gene Flow over a Linguistic Barrier in Iberia, Suggested by Analysis of a Y-Chromosomal DNA Polymorphism

We have examined the worldwide distribution of a Y-chromosomal base-substitution polymorphism, the T/C transition at SRY-2627, where the T allele defines haplogroup 22; sequencing of primate homologues shows that the ancestral state cannot be determined unambiguously but is probably the C allele. Of 1,191 human Y chromosomes analyzed, 33 belong to haplogroup 22. Twenty-nine come from Iberia, and the highest frequencies are in Basques (11%; n=117) and Catalans (22%; n=32). Microsatellite and minisatellite (MSY1) diversity analysis shows that non-Iberian haplogroup-22 chromosomes are not significantly different from Iberian ones. The simplest interpretation of these data is that haplogroup 22 arose in Iberia and that non-Iberian cases reflect Iberian emigrants. Several different methods were used to date the origin of the polymorphism: microsatellite data gave ages of 1,650, 2,700, 3,100, or 3,450 years, and MSY1 gave ages of 1,000, 2,300, or 2,650 years, although 95% confidence intervals on all of these figures are wide. The age of the split between Basque and Catalan haplogroup-22 chromosomes was calculated as only 20% of the age of the lineage as a whole. This study thus provides evidence for direct or indirect gene flow over the substantial linguistic barrier between the Indo-European and non-Indo-European-speaking populations of the Catalans and the Basques, during the past few thousand years.