Serotonin,behavior, and natural selection in New World monkeys

Traits that undergo massive natural selection pressure, with multiple events of positive selection, are hard to find. Social behaviour, in social animals, is crucial for survival, and genetic networks involved in behaviour, such as those of serotonin (5‐HT) and other neurotransmitters, must be the target of natural selection. Here, we used molecular analyses to search for signals of positive selection in the 5‐HT system and found such signals in the M3‐M4 intracellular domain of the 5‐HT3A serotonin receptor subunit (HTR3A) in primates. We detected four amino acid sites with signs of putatively positive selection (398, 403, 432 and 416); the first three showed indications of being selected in New World monkeys (NWM, Platyrrhini), specifically in the Callitrichinae branch. Additionally, we searched for associations of these amino acid variants with social behavioural traits (i.e. sex‐biased dispersal, dominance and social monogamy) using classical and Bayesian methods, and found statistically significant associations for unbiased sex dispersal (398L and 416S), unbiased sex dominance (416S) and social monogamy (416S), as well as significant positive correlation between female dispersal and 403G. Furthermore, we found putatively functional protein motifs determined by three selected sites, of which we highlight a ligand motif to GSK3 in the 416S variant, appearing only in Platyrrhini. 5‐HT, 5‐HT3A receptor and GSK3 are part of a network that participates in neurodevelopment and regulates behaviour, among other functions. We suggest that these genetic variations, together with those found in other neurotransmitter systems, must contribute to adaptive behaviours and consequently to fitness in NWMs.     

Distinctively variable sequence-based nuclear DNA markers for multilocus phylogeography of the soybean- and rice-infecting fungal pathogen Rhizoctonia solani AG-1 IA

A series of multilocus sequence-based nuclear DNA markers was developed to infer the phylogeographical history of the Basidiomycetous fungal pathogen Rhizoctonia solani AG-1 IA infecting rice and soybean worldwide. The strategy was based on sequencing of cloned genomic DNA fragments (previously used as RFLP probes) and subsequent screening of fungal isolates to detect single nucleotide polymorphisms (SNPs). Ten primer pairs were designed based on these sequences, which resulted in PCR amplification of 200-320 bp size products and polymorphic sequences in all markers analyzed. By direct sequencing we identified both homokaryon and heterokaryon (i.e. dikaryon) isolates at each marker. Cloning the PCR products effectively estimated the allelic phase from heterokaryotic isolates. Information content varied among markers from 0.5 to 5.9 mutations per 100 bp. Thus, the former RFLP codominant probes were successfully converted into six distinctively variable sequence-based nuclear DNA markers. Rather than discarding low polymorphism loci, the combination of these distinctively variable anonymous nuclear markers would constitute an asset for the unbiased estimate of the phylogeographical parameters such as population sizes and divergent times, providing a more reliable species history that shaped the current population structure of R. solani AG-1 IA.     

Dynamic chromosomes

A report on the sixth biannual FASEB conference on Yeast Chromosome Structure, Replication and Segregation held in Snowmass, Colorado, 19-24 August, 2000.     

Physiological behaviour, oxidative damage and antioxidative protection of olive trees grown under different irrigation regimes

Irrigation effects were investigated on an 8-year-old olive (Olea europaea L., cv. Cobrançosa) commercial orchard located in northeast Portugal. Trees were subjected to a rainfed control (T0) and three treatments (T1, T2, T3) that received a seasonal water amount equivalent to 30%, 60% and 100% of the estimated local evaporative demand by a drip irrigation system. Irrigation increases the photosynthetic activity of olive trees, in association with increases in water status, and reduces the midday and afternoon depression in gas exchange. The closely association between photosynthetic rate (A) and stomatal conductance (g _s) revealed that the decline in net photosynthesis over the course of the day was largely a consequence of stomatal limitation. However, the ratio of intercellular to atmospheric CO₂ concentration increased markedly from morning to midday in non-irrigated plants, in spite of lower g _s, suggesting that non-stomatal limitations of photosynthesis also occur when environmental conditions become more stressful. The occurrence of perturbations at chloroplastic level in rainfed plants was demonstrated by a lower maximum photochemical efficiency of photosystem II during the afternoon. Chlorophyll fluorescence measurements also revealed the occurrence of a dynamic photoinhibition in irrigated trees, mainly in T2 and T3, which seemed to be effective in protecting the photosynthetic apparatus from photodamage. Irrigation enhances antioxidant protection and decreases the oxidative damage at leaf level. Leaves grown under rainfed conditions revealed symptoms of oxidative stress, like the reduction (14%) in chlorophyll concentration and the increased levels (57%) of lipid peroxidation. We also found that the scavenging function of superoxide dismutase was impaired in rainfed plants. In contrast, the low thiobarbituric acid reactive substances concentration in T3 indicates that irrigation enhances the repairing mechanisms and decreases the oxidative damage by lipid peroxidation. Accordingly, leaves in T3 treatment had high levels of –SH compounds and the highest antioxidant potential. Meanwhile, the finding that guaiacol peroxidase activity increased in rainfed plants, associated with the appearance of oxidative damage, suggests that this enzyme has no major antioxidative function in olive.     

Cloning, expression, and structural analysis of recombinant BJcuL, a c-type lectin from the Bothrops jararacussu snake venom

The lactose-binding lectin from Bothrops jararacussu venom (BJcuL) is a homodimer belonging to group VII of the c-type animal lectins. BJcuL has also been shown to serve as an interesting tool for combating tumor progression by inhibiting cancer and endothelial cell growth. However, detailed structural studies of BJcuL and its biological mechanisms of cytotoxicity are yet to be reported, perhaps because of the non-availability of recombinant proteins in necessary quantities. Intending to increase the present information about structural and consequently the understating of biological studies, the cDNA coding for BJcuL from a venom gland has been cloned and sequenced. The mature protein-coding region was amplified by PCR with specific oligonucleotides, and subcloned into the pET-15b vector to express the recombinant BJcuL in Escherichia coli BL21 (DE3). The deduced amino acid sequence exhibits a high degree of sequence identity with c-type lectins (CTLs) and c-type lectin-like domains (CTLDs). An insoluble and inactive 18.5-kDa protein was overexpressed after 1.0mM IPTG induction. The recombinant BJcuL was recovered and denatured in a buffer with 6M urea and purified on a nickel-affinity column. Protein refolding was carried out on this column, during procedure purification, followed by dialysis against CTBS and then by gel filtration for separation of the active dimmer. The refolding process of rBJcuL and the analysis of its structure were confirmed by biological assay, circular dichroism, and MALDI-TOF.     

Mechanisms of glutamate release from astrocytes: gap junction "hemichannels", purinergic receptors and exocytotic release

Neuronal exocytotic release of glutamate at synapses involves a highly specialized vesicular apparatus, consisting of a variety of proteins connected to the vesicles or required for vesicular fusion to the presynaptic membrane. Astrocytes also release glutamate, and recent evidence indicates that this release can modify neuronal function. Several mechanisms have been proposed for astrocytic release of glutamate under pathological conditions, such as reversal of glutamate transporters and opening of volume sensitive ion channels. In this review we limit our discussion to findings supporting the exocytotic release of glutamate, as well as two new pathways implicated in this release, the ionotropic (P2X) purinergic receptors and gap junction hemichannels.     

Characterization and recruitment of plasmacytoid dendritic cells in synovial fluid and tissue of patients with chronic inflammatory arthritis

Dendritic cells (DCs) are thought to play a key role in driving the immunopathogenic response underlying chronic inflammatory arthritis. In this study, we have examined the presence and phenotype of plasmacytoid DCs (pDCs) in the synovial fluids (SF) of patients with rheumatoid arthritis (RA), psoriatic arthritis (PA), and osteoarthritis (OA) and determined the chemotactic properties of SF from these patients toward pDCs. Flow cytometry analysis showed that the percentage of pDCs, identified as a population of Lin(-)CD123(++) cells, is 4- to 5-fold higher in RA SF and PA SF than in OA SF. The morphological and immunophenotypic characterization of pDCs isolated from PA and RA SF indicates that they are in an immature state, most likely due to inhibitory factors present in RA SF, but are still able to undergo maturation when exposed ex vivo to viral agent or unmethylated DNA. CD123(+) and BDCA2(+) pDCs were detected by immunohistochemistry in RA synovial tissue in which expression of the IFN-alpha-inducible protein MxA was also found, suggesting production of type I IFN by maturing pDCs. We also show that CXCR3 and CXCR4 are expressed by both blood-derived pDCs and pDCs isolated from RA and PA SF and that CXCL-10, CXCL-11, and CXCL-12 present in RA and PA SF stimulate chemotaxis of blood-derived pDCs. Altogether, these findings suggest that chemokine-driven recruitment of pDCs from the blood to the inflamed synovium could be important in the regulation of the immune response in chronic inflammatory arthritis.