Mind the gap: keeping UV lesions in check

Cells respond to genotoxic insults by triggering a DNA damage checkpoint surveillance mechanism and by activating repair pathways. Recent findings indicate that the two processes are more related than originally thought. Here we discuss the mechanisms involved in responding to UV-induced lesions in different phases of the cell cycle and summarize the most recent data in a model where Nucleotide Excision Repair (NER) and exonucleolytic activities act in sequence leading to checkpoint activation in non replicating cells. The critical trigger is likely represented by problematic intermediates that cannot be completely or efficiently repaired by NER. In S phase cells, on the other hand, the replicative polymerases, blocked by bulky UV lesions, re-initiate DNA synthesis downstream of the lesions, leaving behind a ssDNA tract. If these gaps are not rapidly refilled, checkpoint kinases will be activated.     

Calcium gradients in conifer pollen tubes; dynamic properties differ from those seen in angiosperms

Pollen tubes are an established model system for examining polarized cell growth. The focus here is on pollen tubes of the conifer Norway spruce (Picea abies, Pinaceae); examining the relationship between cytosolic free Ca2+, tip elongation, and intracellular motility. Conifer pollen tubes show important differences from their angiosperm counterparts; they grow more slowly and their organelles move in an unusual fountain pattern, as opposed to reverse fountain, in the tip. Ratiometric ion imaging of growing pollen tubes, microinjected with fura-2-dextran, reveals a tip-focused [Ca2+]i gradient extending from 450 nM at the extreme apex to 225 nM at the base of the tip clear zone. Injection of 5,5' dibromo-BAPTA does not dissipate the apical gradient, but stops cell elongation and uniquely causes rapid, transient increases of apical free Ca2+. The [Ca2+]i gradient is, however, dissipated by reversible perfusion of extracellular caffeine. When the basal cytosolic free Ca2+ concentration falls below 150 nM, again a large increase in apical [Ca2+]i occurs. An external source of calcium is not required for germination but significantly enhances elongation. However, both germination and elongation are significantly inhibited by the inclusion of calcium channels blockers, including lanthanum, gadolinium, or verapamil. Modulation of intracellular calcium also affects organelle position and motility. Extracellular perfusion of lanthanides reversibly depletes the apical [Ca2+]i gradient, altering organelle positioning in the tip. Later, during recovery from lanthanide perfusion, organelle motility switches direction to a reverse fountain. When taken together these data show a unique interplay in Picea abies pollen tubes between intracellular calcium and the motile processes controlling cellular organization.     

DNA sequence polymorphism and divergence at the erect wing and suppressor of sable loci of Drosophila melanogaster and D. simulans

Several evolutionary models of linked selection (e.g., genetic hitchhiking, background selection, and random environment) predict a reduction in polymorphism relative to divergence in genomic regions where the rate of crossing over per physical distance is restricted. We tested this prediction near the telomere of the Drosophila melanogaster and D. simulans X chromosome at two loci, erect wing (ewg) and suppressor of sable [su(s)]. Consistent with this prediction, polymorphism is reduced at both loci, while divergence is normal. The reduction is greater at ewg, the more distal of the two regions. Two models can be discriminated by comparing the observed site frequency spectra with those predicted by the models. The hitchhiking model predicts a skew toward rare variants in a sample, while the spectra under the background-selection model are similar to those of the neutral model of molecular evolution. Statistical tests of the fit to the predictions of these models require many sampled alleles and segregating sites. Thus we used SSCP and stratified DNA sequencing to cover a large number of randomly sampled alleles (approximately 50) from each of three populations. The result is a clear trend toward negative values of Tajima's D, indicating an excess of rare variants at ewg, the more distal of the two loci. One fixed difference among the populations and high FST values indicate strong population subdivision among the three populations at ewg. These results indicate genetic hitchhiking at ewg, in particular, geographically localized hitchhiking events within Africa. The reduction of polymorphism at su(s) combined with the excess of high-frequency variants in D. simulans is inconsistent with the hitchhiking and background-selection models.     

Elevated polymorphism and divergence in the class C scavenger receptors of Drosophila melanogaster and D. simulans

Scavenger receptor proteins are involved in the cellular internalization of a broad variety of foreign material, including pathogenic bacteria during phagocytosis. I find here that nonsynonymous divergence in three class C scavenger receptors (Sr-C's) between Drosophila melanogaster and D. simulans and between each of these species and D. yakuba is approximately four times the typical genome average. These genes also exhibit unusually high levels of segregating nonsynonymous polymorphism in D. melanogaster and D. simulans populations. A fourth Sr-C is comparatively conserved. McDonald-Kreitman tests reveal a significant excess of replacement fixations between D. melanogaster and D. simulans in the Sr-C's, but tests of polymorphic site frequency spectra do not support models of directional selection. It is possible that the molecular functions of SR-C proteins are sufficiently robust to allow exceptionally high amino acid substitution rates without compromising organismal fitness. Alternatively, SR-Cs may evolve under diversifying selection, perhaps as a result of pressure from pathogens. Interestingly, Sr-CIII and Sr-CIV are polymorphic for premature stop codons. Sr-CIV is also polymorphic for an in-frame 101-codon deletion and for the absence of one intron.     

Bacterial community structure and activity in different Cd-treated forest soils

In this study we compared indicators of Cd bioavailability (water extracts, Lakanen extracts, free ions) and ecotoxicity in forest soils with contrasting physico-chemical characteristics. Soil samples were treated with CdCl(2) solutions (0, 0.1, 1, 10 and 100 mM) and incubated for 30 days. Microbial activity indexes (acid phosphatase, beta-glucosidase, basal respiration) and changes in bacterial community structure using terminal restriction fragment length polymorphism (T-RFLP) fingerprinting were investigated. The Cd concentrations measured ranged from 1% to 37% of the total additions in water extracts, to higher levels in Lakanen extracts. Effects of Cd were observed at bioavailable concentrations exceeding United Nations/European Economic Commission UN/ECE guidelines for total Cd in the soil solution. Basal respiration was the most affected index, while enzymatic activities showed variable responses to the Cd treatments. We also noticed that soils with pH higher than 6.7 and clay content higher than 50% showed inhibition of basal respiration but no marked shift in bacterial community structure. Soils with lower pH (pH <5.8) with less clay content (<50%) showed in addition strong changes in the bacterial community structure. Our results provide evidence for the importance of relating the effects of Cd on the soil communities to soil properties and to bioavailability.     

Synaptic Plasticity in Neurodegenerative Diseases Evaluated and Modulated by In Vivo Neurophysiological Techniques

Several studies demonstrated in experimental models and in humans synaptic plasticity impairment in some neurodegenerative and neuropsychiatric diseases such as Parkinson's disease, Alzheimer's disease, Huntington's disease, and schizophrenia. Recently new neurophysiological tools, such as repetitive transcranial magnetic stimulation and transcranial direct current stimulation, have been introduced in experimental and clinical settings for studying physiology of the brain and modulating cortical activity. These techniques use noninvasive transcranial electrical or magnetic stimulation to modulate neurons activity in the human brain. Cortical stimulation might enhance or inhibit the activity of cortico?Csubcortical networks, depending on stimulus frequency and intensity, current polarity, and other stimulation parameters such as the configuration of the induced electric field and stimulation protocols. On this basis, in the last two decades, these techniques have rapidly become valuable tools to investigate physiology of the human brain and have been applied to treat drug-resistant neurological and psychiatric diseases. Here we describe these techniques and discuss the mechanisms that may explain these effects.     

Female Drosophila melanogaster suffer reduced defense against infection due to seminal fluid components

Reduced defense against infection is commonly observed as a consequence of reproductive activity, but little is known about how post-mating immunosuppression occurs. In this work, we use Drosophila melanogaster as a model to test the role of seminal fluid components and egg production in suppressing post-mating immune defense. We also evaluate whether systemic immune system activity is altered during infection in mated females. We find that post-mating reduction in female defense depends critically on male transfer of sperm and seminal fluid proteins, including the accessory gland protein known as "sex peptide." However, the effect of these male factors is dependent on the presence of the female germline. We find that mated females have lower antimicrobial peptide gene expression than virgin females in response to systemic infection, and that this lower expression correlates with higher systemic bacterial loads. We conclude that, upon receipt of sperm and seminal fluid proteins, females experience a germline-dependent physiological shift that directly or indirectly reduces their overall ability to defend against infection, at least in part through alteration of humoral immune system activity.     

Light- and singlet oxygen-mediated antifungal activity of phenylphenalenone phytoalexins.

The light-induced singlet oxygen production and antifungal activity of phenylphenalenone phytoalexins isolated from infected banana plants (Musa acuminata) are reported. Upon absorption of light energy all studied phenylphenalenones sensitise the production of singlet oxygen in polar and non-polar media. Antifungal activity of these compounds towards Fusarium oxysporum is enhanced in the presence of light. These results, together with the correlation of IC50 values under illumination with the quantum yield of singlet oxygen production and the enhancing effect of D2O on the antifungal activity, suggest the intermediacy of singlet oxygen produced by electronic excitation of the phenylphenalenone phytoalexins.