As,Pb, Sb,and Zn transfer from soil to root of wild rosemary: do native symbionts matter?

Background and aims

This is an in natura study aimed to determine the potential of Rosmarinus officinalis for phytostabilization of trace metal and metalloid (TMM)-contaminated soils in the Calanques National Park (Marseille, southeast of France). The link between rosemary tolerance/accumulation of As, Pb, Sb, and Zn and root symbioses with arbuscular mycorrhizal (AM) fungi and/or dark septate endophytes (DSE) was examined.

Methods

Eight sites along a gradient of contamination were selected for soil and root collections. TMM concentrations were analyzed in all the samples and root symbioses were observed. Moreover, in the roots of various diameters collected in the most contaminated site, X-ray microfluorescence methods were used to determine TMM localization in tissues.

Results

Rosemary accumulated, in its roots, the most labile TMM fraction in the soil. The positive linear correlation between TMM concentrations in soil and endophyte root colonization rates suggests the involvement of AM fungi and DSE in rosemary tolerance to TMM. Moreover, a typical TMM localization in root peripheral tissues of thin roots containing endophytes forming AM and DSE development was observed using X-ray microfluorescence.

Conclusions

Rosemary and its root symbioses appeared as a potential candidate for a phytostabilization process of metal-contaminated soils in Mediterranean area.     

Def1 Promotes the Degradation of Pol3 for Polymerase Exchange to Occur During DNA-Damage–Induced Mutagenesis in Saccharomyces cerevisiae

DNA damages hinder the advance of replication forks because of the inability of the replicative polymerases to synthesize across most DNA lesions. Because stalled replication forks are prone to undergo DNA breakage and recombination that can lead to chromosomal rearrangements and cell death, cells possess different mechanisms to ensure the continuity of replication on damaged templates. Specialized, translesion synthesis (TLS) polymerases can take over synthesis at DNA damage sites. TLS polymerases synthesize DNA with a high error rate and are responsible for damage-induced mutagenesis, so their activity must be strictly regulated. However, the mechanism that allows their replacement of the replicative polymerase is unknown. Here, using protein complex purification and yeast genetic tools, we identify Def1 as a key factor for damage-induced mutagenesis in yeast. In in vivo experiments we demonstrate that upon DNA damage, Def1 promotes the ubiquitylation and subsequent proteasomal degradation of Pol3, the catalytic subunit of the replicative polymerase δ, whereas Pol31 and Pol32, the other two subunits of polymerase δ, are not affected. We also show that purified Pol31 and Pol32 can form a complex with the TLS polymerase Rev1. Our results imply that TLS polymerases carry out DNA lesion bypass only after the Def1-assisted removal of Pol3 from the stalled replication fork.     

The interaction between the cholinergic and dopaminergic system in learning and memory process in rats.

In normal rats, muscarinic acetylcholine receptors (mAChRs) have a facilitating role on both short-term and long-term memory tested by Y-maze task and multi-trial passive avoidance test, respectively, since scopolamine, a specific mAChRs antagonist, impairs both types of memory. A low dose of nicotine (0.3 mg/kg b.w., i.p.), a specific nicotinic acetylcholine receptors (nAChRs) agonist, administered once caused a significant facilitating effect on short-term memory. A higher dose of nicotine (3 mg/kg b.w., i.p.) administered 5 consecutively days had about the same facilitating effect on short- and long-term memory without affecting information acquisition. In rats, having mAChRs and nAChRs blocked by means of scopolamine and chlorisondamine respectively, a low dose of nicotine administered once caused a significant improvement of long-term memory deficits without affecting significantly short-term memory. A higher dose of nicotine administered 5 consecutive days in rats with a double blockade of cholinergic receptors had the same ameliorating effect on long-term memory deficits as low dose. Our data suggest that the antiamnesic effect of nicotine can result from an action at nicotinic receptors subtypes not blocked by chlorisondamine or at nonnicotinic receptors.     

Ten simple rules for creating a brand-new virtual academic meeting (even amid a pandemic)

The increased democratization of the creation, implementation, and attendance of academic conferences has been a serendipitous benefit of the movement toward virtual meetings. The Coronavirus Disease 2019 (COVID-19) pandemic has accelerated the transition to online conferences and, in parallel, their democratization, by necessity. This manifests not just in the mitigation of barriers to attending traditional physical conferences but also in the presentation of new, and more importantly attainable, opportunities for young scientists to carve out a niche in the landscape of academic meetings. Here, we describe an early “proof of principle” of this democratizing power via our experience organizing the Canadian Computational Neuroscience Spotlight (CCNS; crowdcast.io/e/CCNS), a free 2-day virtual meeting that was built entirely amid the pandemic using only virtual tools. While our experience was unique considering the obstacles faced in creating a conference during a pandemic, this was not the only factor differentiating both our experience and the resulting meeting from other contemporary online conferences. Specifically, CCNS was crafted entirely by early career researchers (ECRs) without any sponsors or partners, advertised primarily using social media and “word of mouth,” and designed specifically to highlight and engage trainees. From this experience, we have distilled “10 simple rules” as a blueprint for the design of new virtual academic meetings, especially in the absence of institutional support or partnerships, in this unprecedented environment. By highlighting the lessons learned in implementing our meeting under these arduous circumstances, we hope to encourage other young scientists to embrace this challenge, which would serve as a critical next step in further democratizing academic meetings.     

Clathrin is required for postmitotic Golgi reassembly

During the G2-M transition, the highly organized Golgi apparatus undergoes reversible fragmentation through unstacking of the cisternal ribbon and disassembly into radially dispersed vesicles and tubules. These Golgi-derived fragments redistribute randomly within the cytoplasm, partition stochastically, and in telophase coalesce to generate a functionally and structurally intact Golgi complex. Here we identified a novel step in postmitotic Golgi reassembly that requires the clathrin heavy chain (CHC). We used siRNA-mediated CHC knockdown, biochemistry, and morphological analysis and showed that the spindle- and spindle pole-associated clathrin pools are membrane-bound and required for postmitotic Golgi reassembly. The results presented here show that clathrin remains associated with the spindle poles throughout mitosis and that this clathrin pool is distinct from the previously characterized spindle-associated population. We suggest that clathrin may provide a template for postmitotic Golgi reassembly and cisternal remodeling. In absence of the CHC, the Golgi apparatus remained disconnected and disordered and failed to regain its characteristic perinuclear, lace-like morphology. Our findings build on previous independent reports that clathrin is required for Golgi reassembly following disruption with pharmacological agents and for mitotic chromosome congression.     

Outcome of treatment of human HeLa cervical cancer cells with roscovitine strongly depends on the dosage and cell cycle status prior to the treatment

Exposure of asynchronously growing human HeLa cervical carcinoma cells to roscovitine (ROSC), a selective cyclin‐dependent kinases (CDKs) inhibitor, arrests their progression at the transition between G₂/M and/or induces apoptosis. The outcome depends on the ROSC concentration. At higher dose ROSC represses HPV‐encoded E7 oncoprotein and initiates caspase‐dependent apoptosis. Inhibition of the site‐specific phosphorylation of survivin and Bad, occurring at high‐dose ROSC treatment, precedes the onset of apoptosis and seems to be a prerequisite for cell death. Considering the fact that in HeLa cells the G₁/S restriction checkpoint is abolished by E7, we addressed the question whether the inhibition of CDKs by pharmacological inhibitors in synchronized cells would be able to block the cell‐cycle in G₁ phase. For this purpose, we attempted to synchronize cells by serum withdrawal or by blocking of the mitotic apparatus using nocodazole. Unlike human MCF‐7 cells, HeLa cells do not undergo G₁ block after serum starvation, but respond with a slight increase of the ratio of G₁ population. Exposure of G₁‐enriched HeLa cells to ROSC after re‐feeding does not block their cell‐cycle progression at G₁‐phase, but increases the ratio of S‐ and G₂‐phase, thereby mimicking the effect on asynchronously growing cells. A quite different impact is observed after treatment of HeLa cells released from mitotic block. ROSC prevents their cell cycle progression and cells transiently accumulate in G₁‐phase. These results show that inhibition of CDKs by ROSC in cells lacking the G₁/S restriction checkpoint has different outcomes depending on the cell‐cycle status prior to the onset of treatment. J. Cell. Biochem. 106: 937–955, 2009. © 2009 Wiley‐Liss, Inc.     

Apoptotic rate in patients with myelodisplastic syndrome treated with modulatory compounds of pro-apoptotic cytokines

Excessive apoptosis has a central role in ineffective hematopoiesis in myelodysplastic syndrome (MDS). The aim of the study was to quantify apoptosis and Bcl-2 expression in patients with MDS and to use these parameters in the evaluation of treatment efficacy with compounds modulating proapoptotic cytokines. Bone marrow (BM) samples from eight MDS patients were studied: four with refractory anemia and four with refractory anemia with ringed sideroblasts. Two patients with Hodgkin disease without BM determination were studied for control. Therapy consisted in administration of pentoxyphylline, dexamethasone and ciprofloxacin. Biochemical assay of apoptosis and Bcl-2 was performed using annexin V-biotin conjugate antibody and anti-human Bcl-2 antibody respectively, followed by streptavidine-peroxidase conjugate, and peroxidase substrate. Ultrastructural investigation of BM samples was performed with standard electron microscopy techniques. Most of BM hematopoietic cells in the MDS patients had ultrastructural features of various stages of apoptosis including chromatin condensation and margination, cytoplasm condensation and budding of nuclear and plasma membranes to produce apoptotic bodies. Bcl-2 expression showed an inverse correlation with the rate of the apoptotic process. Periodic evaluation of these two parameters has shown an increase of Bcl-2 expression and a decrease of apoptotic rate in patients who had responded to the treatment. Response to the treatment was appreciated in accordance with their transfusion needs. Treatment efficiency diminished in time. The rate of apoptosis was inversely correlated with the level of Bcl-2 expression. These results confirm the importance of the apoptotic process evaluation in monitoring MDS treatment.     

Histamine effects on endothelial cell fibronectin interaction studied by atomic force microscopy

Atomic force microscopy was used to investigate the cellular response to histamine, one of the major inflammatory mediators that cause endothelial hyperpermeability and vascular leakage. AFM probes were labeled with fibronectin and used to measure binding strength between alpha5beta1 integrin and fibronectin by quantifying the force required to break single fibronectin-integrin bonds. The cytoskeletal changes, binding probability, and adhesion force before and after histamine treatment on endothelial cells were monitored. Cell topography measurements indicated that histamine induces cell shrinkage. Local cell stiffness and binding probability increased twofold after histamine treatment. The force necessary to rupture single alpha5beta1-fibronectin bond increased from 34.0 +/- 0.5 pN in control cells to 39 +/- 1 pN after histamine treatment. Experiments were also conducted to confirm the specificity of the alpha5beta1-fibronectin interaction. In the presence of soluble GRGDdSP the probability of adhesion events decreased >50% whereas the adhesion force between alpha5beta1 and fibronectin remained unchanged. These data indicate that extracellular matrix-integrin interactions play an important role in the endothelial cell response to changes of external chemical mediators. These changes can be recorded as direct measurements on live endothelial cells by using atomic force microscopy.