Direct electrical control of IgG conformation and functional activity at surfaces

We have devised a supramolecular edifice involving His-tagged protein A and antibodies to yield surface immobilized, uniformly oriented, IgG-type, antibody layers with Fab fragments exposed off an electrode surface. We demonstrate here that we can affect the conformation of IgGs, likely pushing/pulling electrostatically Fab fragments towards/from the electrode surface. A potential difference between electrode and solution acts on IgGs’ charged aminoacids modulating the accessibility of the specific recognition regions of Fab fragments by antigens in solution. Consequently, antibody-antigen affinity is affected by the sign of the applied potential: a positive potential enables an effective capture of antigens; a negative one pulls the fragments towards the electrode, where steric hindrance caused by neighboring molecules largely hampers the capture of antigens. Different experimental techniques (electrochemical quartz crystal microbalance, electrochemical impedance spectroscopy, fluorescence confocal microscopy and electrochemical atomic force spectroscopy) were used to evaluate binding kinetics, surface coverage, effect of the applied electric field on IgGs, and role of charged residues on the phenomenon described. These findings expand the concept of electrical control of biological reactions and can be used to gate electrically specific recognition reactions with impact in biosensors, bioactuators, smart biodevices, nanomedicine, and fundamental studies related to chemical reaction kinetics

Lmx1a-Dependent Activation of miR-204/211 Controls the Timing of Nurr1-Mediated Dopaminergic Differentiation

The development of midbrain dopaminergic (DA) neurons requires a fine temporal and spatial regulation of a very specific gene expression program. Here, we report that during mouse brain development, the microRNA (miR-) 204/211 is present at a high level in a subset of DA precursors expressing the transcription factor Lmx1a, an early determinant for DA-commitment, but not in more mature neurons expressing Th or Pitx3. By combining different in vitro model systems of DA differentiation, we show that the levels of Lmx1a influence the expression of miR-204/211. Using published transcriptomic data, we found a significant enrichment of miR-204/211 target genes in midbrain dopaminergic neurons where Lmx1a was selectively deleted at embryonic stages. We further demonstrated that miR-204/211 controls the timing of the DA differentiation by directly downregulating the expression of Nurr1, a late DA differentiation master gene. Thus, our data indicate the Lmx1a-miR-204/211-Nurr1 axis as a key component in the cascade of events that ultimately lead to mature midbrain dopaminergic neurons differentiation and point to miR-204/211 as the molecular switch regulating the timing of Nurr1 expression

Relationships linking primary production, sea ice melting, and biogenic aerosol in the Arctic

AbstractThis study examines the relationships linking methanesulfonic acid (MSA, arising from the atmospheric oxidation of the biogenic dimethylsulfide, DMS) in atmospheric aerosol, satellite-derived chlorophyll a (Chl-a), and oceanic primary production (PP), also as a function of sea ice melting (SIM) and extension of the ice free area in the marginal ice zone (IF-MIZ) in the Arctic. MSA was determined in PM10 samples collected over the period 2010–2012 at two Arctic sites, Ny Ålesund (78.9°N, 11.9°E), Svalbard islands, and Thule Air Base (76.5°N, 68.8°W), Greenland. PP is calculated by means of a bio-optical, physiologically based, semi-analytical model in the potential source areas located in the surrounding oceanic regions (Barents and Greenland Seas for Ny Ålesund, and Baffin Bay for Thule). Chl-a peaks in May in the Barents sea and in the Baffin Bay, and has maxima in June in the Greenland sea; PP follows the same seasonal pattern of Chl-a, although the differences in absolute values of PP in the three seas during the blooms are less marked than for Chl-a. MSA shows a better correlation with PP than with Chl-a, besides, the source intensity (expressed by PP) is able to explain more than 30% of the MSA variability at the two sites; the other factors explaining the MSA variability are taxonomic differences in the phytoplanktonic assemblages, and transport processes from the DMS source areas to the sampling sites. The taxonomic differences are also evident from the slopes of the correlation plots between MSA and PP: similar slopes (in the range 34.2–36.2 ng m−3of MSA/(gC m−2 d−1)) are found for the correlation between MSA at Ny Ålesund and PP in Barents Sea, and between MSA at Thule and PP in the Baffin Bay; conversely, the slope of the correlation between MSA at Ny Ålesund and PP in the Greenland Sea in summer is smaller (16.7 ng m−3of MSA/(gC m−2 d−1)). This is due to the fact that DMS emission from the Barents Sea and Baffin Bay is mainly related to the MIZ diatoms, which are prolific DMS producers, whereas in the Greenland Sea the DMS peak is related to an offshore pelagic bloom where low-DMS producer species are present. The sea ice dynamic plays a key role in determining MSA concentration in the Arctic, and a good correlation between MSA and SIM (slope = 39 ng m−3 of MSA/106 km2 SIM) and between MSA and IF-MIZ (slope = 56 ng m−3 of MSA/106 km2 IF-MIZ) is found for the cases attributable to bloomings of diatoms in the MIZ. Such relationships are calculated by combining the data sets from the two sites and suggest that PP is related to sea ice melting and to the extension of marginal sea ice areas, and that these factors are the main drivers for MSA concentrations at the considered Arctic sites

Lmx1a-Dependent Activation of miR-204/211 Controls the Timing of Nurr1-Mediated Dopaminergic Differentiation

The development of midbrain dopaminergic (DA) neurons requires a fine temporal and spatial regulation of a very specific gene expression program. Here, we report that during mouse brain development, the microRNA (miR-) 204/211 is present at a high level in a subset of DA precursors expressing the transcription factor Lmx1a, an early determinant for DA-commitment, but not in more mature neurons expressing Th or Pitx3. By combining different in vitro model systems of DA differentiation, we show that the levels of Lmx1a influence the expression of miR-204/211. Using published transcriptomic data, we found a significant enrichment of miR-204/211 target genes in midbrain dopaminergic neurons where Lmx1a was selectively deleted at embryonic stages. We further demonstrated that miR-204/211 controls the timing of the DA differentiation by directly downregulating the expression of Nurr1, a late DA differentiation master gene. Thus, our data indicate the Lmx1a-miR-204/211-Nurr1 axis as a key component in the cascade of events that ultimately lead to mature midbrain dopaminergic neurons differentiation and point to miR-204/211 as the molecular switch regulating the timing of Nurr1 expression

miR-34b/c Regulates Wnt1 and Enhances Mesencephalic Dopaminergic Neuron Differentiation

The differentiation of dopaminergic neurons requires concerted action of morphogens and transcription factors acting in a precise and well-defined time window. Very little is known about the potential role of microRNA in these events. By performing a microRNA-mRNA paired microarray screening, we identified miR-34b/c among the most upregulated microRNAs during dopaminergic differentiation. Interestingly, miR-34b/c modulates Wnt1 expression, promotes cell cycle exit, and induces dopaminergic differentiation. When combined with transcription factors ASCL1 and NURR1, miR-34b/c doubled the yield of transdifferentiated fibroblasts into dopaminergic neurons. Induced dopaminergic (iDA) cells synthesize dopamine and show spontaneous electrical activity, reversibly blocked by tetrodotoxin, consistent with the electrophysiological properties featured by brain dopaminergic neurons. Our findings point to a role for miR-34b/c in neuronal commitment and highlight the potential of exploiting its synergy with key transcription factors in enhancing in vitro generation of dopaminergic neurons.Peer reviewe

miR-218 promotes dopaminergic differentiation and controls neuron excitability and neurotransmitter release through the regulation of a synaptic-related genes network

In the brain microRNAs (miRNAs) are believed to play a role in orchestrating synaptic plasticity at a higher-level by acting as an additional mechanism of translational regulation, alongside the mRNAs/polysome system. Despite extensive research, our understanding of the specific contribution of individual miRNA to the function of dopaminergic neurons (DAn) remains limited. By performing a dopaminergic-specific miRNA screening, we have identified miR-218 as a critical regulator of DAn activity in male and female mice. We have found that miR-218 is specifically expressed in mesencephalic DAn and is able to promote dopaminergic differentiation of embryonic stem cells and functional maturation of transdifferentiated induced DA neurons. Midbrain-specific deletion of both genes encoding for miR-218 (referred to as miR-218-1 and mir218-2) affects the expression of a cluster of synaptic-related mRNAs and alters the intrinsic excitability of DAn, as it increases instantaneous frequencies of evoked action potentials, reduces rheobase current, affects the ionic current underlying the action potential after hyperpolarization phase and reduces dopamine efflux in response to a single electrical stimulus. Our findings provide a comprehensive understanding of the involvement of miR-218 in the dopaminergic system and highlight its role as a modulator of dopaminergic transmission