The G-protein–gated K+ channel,IKACh, is required for regulation of pacemaker activity and recovery of resting heart rate after sympathetic stimulation

Parasympathetic regulation of sinoatrial node (SAN) pacemaker activity modulates multiple ion channels to temper heart rate. The functional role of the G-protein–activated K⁺ current (I_KACh) in the control of SAN pacemaking and heart rate is not completely understood. We have investigated the functional consequences of loss of I_KACh in cholinergic regulation of pacemaker activity of SAN cells and in heart rate control under physiological situations mimicking the fight or flight response. We used knockout mice with loss of function of the Girk4 (Kir3.4) gene (Girk4^−/− mice), which codes for an integral subunit of the cardiac I_KACh channel. SAN pacemaker cells from Girk4^−/− mice completely lacked I_KACh. Loss of I_KACh strongly reduced cholinergic regulation of pacemaker activity of SAN cells and isolated intact hearts. Telemetric recordings of electrocardiograms of freely moving mice showed that heart rate measured over a 24-h recording period was moderately increased (10%) in Girk4^−/− animals. Although the relative extent of heart rate regulation of Girk4^−/− mice was similar to that of wild-type animals, recovery of resting heart rate after stress, physical exercise, or pharmacological β-adrenergic stimulation of SAN pacemaking was significantly delayed in Girk4^−/− animals. We conclude that I_KACh plays a critical role in the kinetics of heart rate recovery to resting levels after sympathetic stimulation or after direct β-adrenergic stimulation of pacemaker activity. Our study thus uncovers a novel role for I_KACh in SAN physiology and heart rate regulation.     

ABCA1-dependent serum cholesterol efflux capacity inversely correlates with pulse wave velocity in healthy subjects

The capacity of HDL to induce cell cholesterol efflux is considered one of its main antiatherogenic properties. Little is known about the impact of such HDL function on vascular physiology. We investigated the relationship between ABCA1-dependent serum cholesterol efflux capacity (CEC), an HDL functionality indicator, and pulse wave velocity (PWV), an indicator of arterial stiffness. Serum of 167 healthy subjects was used to conduct CEC measurement, and carotid-femoral PWV was measured with a high-fidelity tonometer. J774 macrophages, labeled with [³H]cholesterol and stimulated to express ABCA1, were exposed to sera; the difference between cholesterol efflux from stimulated and unstimulated cells provided specific ABCA1-mediated CEC. PWV is inversely correlated with ABCA1-dependent CEC (r = −0.183; P = 0.018). Moreover, controlling for age, sex, body mass index, mean arterial pressure, serum LDL, HDL-cholesterol, and fasting plasma glucose, PWV displays a significant negative regression on ABCA1-dependent CEC (β = −0.204; 95% confidence interval, −0.371 to −0.037). The finding that ABCA1-dependent CEC, but not serum HDL cholesterol level (r = −0.002; P = 0.985), is a significant predictor of PWV in healthy subjects points to the relevance of HDL function in vascular physiology and arterial stiffness prevention.     

Fatal Measles Virus Infection Prevented by Brain-Penetrant Fusion Inhibitors

Measles virus (MV) infection causes an acute childhood disease that can include infection of the central nervous system and can rarely progress to severe neurological disease for which there is no specific treatment. We generated potent antiviral peptide inhibitors of MV entry and spreading and MV-induced cell fusion. Dimers of MV-specific peptides derived from the C-terminal heptad repeat region of the MV fusion protein, conjugated to cholesterol, efficiently protect SLAM transgenic mice from fatal MV infection. Fusion inhibitors hold promise for the prophylaxis of MV infection in unvaccinated and immunocompromised people, as well as potential for the treatment of grave neurological complications of measles.     

HnRNP A1 controls a splicing regulatory circuit promoting mesenchymal-to-epithelial transition

Epithelial-to-mesenchymal transition (EMT) is an embryonic program used by cancer cells to acquire invasive capabilities becoming metastatic. ΔRon, a constitutively active isoform of the Ron tyrosine kinase receptor, arises from skipping of Ron exon 11 and provided the first example of an alternative splicing variant causatively linked to the activation of tumor EMT. Splicing of exon 11 is controlled by two adjacent regulatory elements, a silencer and an enhancer of splicing located in exon 12. The alternative splicing factor and oncoprotein SRSF1 directly binds to the enhancer, induces the production of ΔRon and activates EMT leading to cell locomotion. Interestingly, we now find an important role for hnRNP A1 in controlling the activity of the Ron silencer. HnRNP A1 is able to antagonize the binding of SRSF1 and prevent exon skipping. Notably, hnRNP A1, by inhibiting the production of ΔRon, activates the reversal program, namely the mesenchymal-to-epithelial transition, which instead occurs at the final metastasis sites. Also, hnRNP A1 affects Ron splicing by regulating the expression level of hnRNP A2/B1, which similarly to SRSF1 can promote ΔRon production. These results shed light on how splicing regulation contributes to the tumor progression and provide potential targets to develop anticancer therapies.     

New information on Bobosaurus forojuliensis (Reptilia: Sauropterygia): implications for plesiosaurian evolution

Preparation of the holotype specimen of Bobosaurus forojuliensis, a large sauropterygian from the lower Carnian of northeastern Italy, revealed new morphological data relevant in establishing its phylogenetic affinities among pistosauroid taxa and its relationships with plesiosaurians. Inclusion of B. forojuliensis in two phylogenetic analyses focusing, respectively, on sauropterygians and pistosauroids agreed in placing the Italian taxon as closer to plesiosaurians than to other pistosauroids. The phylogenetic interpretation of Bobosaurus was not biased by assumptions on character weighting, is consistent with its relatively younger age compared to most pistosauroids, extends the fossil record of the plesiosaurian basal lineage back to the Carnian and supports the earliest diversification of the clade during the Late Triassic in agreement with the record of several distinct lineages of rhomaleosaurids, plesiosauroids and pliosauroids in the lowermost Jurassic. Bobosaurus shows that the evolution of the plesiosaurian body plan from the ancestral pistosauroid grade was a step-wise process, and that some of the vertebral and appendicular specialisations of Jurassic and Cretaceous plesiosaurians had already developed in the earliest Late Triassic.     

Plant hormone signaling and modulation of DNA repair under stressful conditions

The role played by phytohormone signaling in the modulation of DNA repair gene and the resulting effects on plant adaptation to genotoxic stress are poorly investigated. Information has been gathered using the Arabidopsis ABA (abscisic acid) overly sensitive mutant abo4-1, defective in the DNA polymerase ε function that is required for DNA repair and recombination. Similarly, phytohormone-mediated regulation of the Ku genes, encoding the Ku heterodimer protein involved in DNA repair, cell cycle control and telomere homeostasis has been demonstrated, highlighting a scenario in which hormones might affect genome stability by modulating the frequency of homologous recombination, favoring plant adaptation to genotoxic stress. Within this context, the characterisation of Arabidopsis AtKu mutants allowed disclosing novel connections between DNA repair and phytohormone networks. Another intriguing aspect deals with the emerging correlation between plant defense response and the mechanisms responsible for genome stability. There is increasing evidence that systemic acquired resistance (SAR) and homologous recombination share common elements represented by proteins involved in DNA repair and chromatin remodeling. This hypothesis is supported by the finding that volatile compounds, such as methyl salicylate (MeSA) and methyl jasmonate (MeJA), participating in the plant-to-plant communication can trigger genome instability in response to genotoxic stress agents. Phytohormone-mediated control of genome stability involves also chromatin remodeling, thus expanding the range of molecular targets. The present review describes the most significant advances in this specific research field, in the attempt to provide a better comprehension of how plant hormones modulate DNA repair proteins as a function of stress.     

Bendamustine overcomes resistance to melphalan in myeloma cell lines by inducing cell death through mitotic catastrophe

Melphalan has been a mainstay of multiple myeloma (MM) therapy for many years. However, following treatment with this alkylator, malignant plasma cells usually escape both apoptosis and cell cycle control, and acquire drug-resistance resulting in tumor progression. Bendamustine is being used in MM patients refractory to conventional DNA-damaging agents, although the mechanisms driving this lack of cross-resistance are still undefined. Here, we investigated the molecular pathway of bendamustine-induced cell death in melphalan-sensitive and melphalan-resistant MM cell lines. Bendamustine affected cell survival resulting in secondary necrosis, and prompted cell death primarily through caspase-2 activation. Also, bendamustine blocked the cell cycle in the G2/M phase and induced micronucleation, erratic chromosome spreading and mitotic spindle perturbations in melphalan-resistant MM cells. In these cells, both Aurora kinase A (AURKA) and Polo-like kinase-1 (PLK-1), key components of the spindle-assembly checkpoint, were down-regulated following incubation with bendamustine, whereas levels of Cyclin B1 increased as a consequence of the prolonged mitotic arrest induced by the drug. These findings indicate that, at least in vitro, bendamustine drives cell death by promoting mitotic catastrophe in melphalan-resistant MM cells. Hence, activation of this alternative pathway of cell death may be a novel approach to the treatment of apoptosis-resistant myelomas.     

The DivJ,CbrA and PleC system controls DivK phosphorylation and symbiosis in Sinorhizobium meliloti

Sinorhizobium meliloti is a soil bacterium that invades the root nodules it induces on Medicago sativa, whereupon it undergoes an alteration of its cell cycle and differentiates into nitrogen‐fixing, elongated and polyploid bacteroid with higher membrane permeability. In Caulobacter crescentus, a related alphaproteobacterium, the principal cell cycle regulator, CtrA, is inhibited by the phosphorylated response regulator DivK. The phosphorylation of DivK depends on the histidine kinase DivJ, while PleC is the principal phosphatase for DivK. Despite the importance of the DivJ in C. crescentus, the mechanistic role of this kinase has never been elucidated in other Alphaproteobacteria. We show here that the histidine kinases DivJ together with CbrA and PleC participate in a complex phosphorylation system of the essential response regulator DivK in S. meliloti. In particular, DivJ and CbrA are involved in DivK phosphorylation and in turn CtrA inactivation, thereby controlling correct cell cycle progression and the integrity of the cell envelope. In contrast, the essential PleC presumably acts as a phosphatase of DivK. Interestingly, we found that a DivJ mutant is able to elicit nodules and enter plant cells, but fails to establish an effective symbiosis suggesting that proper envelope and/or low CtrA levels are required for symbiosis.