Increases in the soil ammonia oxidizing phylotypes and their rechange due to long-term irrigation with wastewater.

Wastewater irrigation is a common practice for agricultural systems in arid and semiarid zones, which can help to overcome water scarcity and contribute with nutrient inputs. Ammonia-oxidizing bacteria (AOB) and archaea (AOA) are key in the transformation of NH4+-N in soil and can be affected by variations in soil pH, EC, N and C content, or accumulation of pollutants, derived from wastewater irrigation. The objective of this study was to determine the changes in the ammonia oxidizing communities in agricultural soils irrigated with wastewater for different periods of time (25, 50, and 100 years), and in rainfed soils (never irrigated). The amoA gene encoding for the catalytic subunit of the ammonia monooxygenase was used as molecular reporter; it was quantified by qPCR and sequenced by high throughput sequencing, and changes in the community composition were associated with the soil physicochemical characteristics. Soils irrigated with wastewater showed up to five times more the abundance of ammonia oxidizers (based on 16S rRNA gene relative abundance and amoA gene copies) than those under rainfed agriculture. While the amoA-AOA: amoA-AOB ratio decreased from 9.8 in rainfed soils to 1.6 in soils irrigated for 100 years, indicating a favoring environment for AOB rather than AOA. Further, the community structure of both AOA and AOB changed during wastewater irrigation compared to rainfed soils, mainly due to the abundance variation of certain phylotypes. Finally, the significant correlation between soil pH and the ammonia oxidizing community structure was confirmed, mainly for AOB; being the main environmental driver of the ammonia oxidizer community. Also, a calculated toxicity index based on metals concentrations showed a correlation with AOB communities, while the content of carbon and nitrogen was more associated with AOA communities. The results indicate that wastewater irrigation influence ammonia oxidizers communities, manly by the changes in the physicochemical environment.

Roles of the α1B-Adrenergic Receptor Phosphorylation Domains in Signaling and Internalization.

The function of the α1B-adrenergic receptor phosphorylation sites previously detected by mass spectrometry was evaluated by employing mutants, substituting them with non-phosphorylatable amino acids. Substitution of the intracellular loop 3 (IL3) sites did not alter baseline or stimulated receptor phosphorylation, whereas substitution of phosphorylation sites in the carboxyl terminus (Ctail) or both domains (IL3/Ctail) markedly decreased receptor phosphorylation. Cells expressing the IL3 or Ctail receptor mutants exhibited a noradrenaline-induced calcium-maximal response similar to those expressing the wild-type receptor, and a shift to the left in the concentration-response curve to noradrenaline was also noticed. Cells expressing the IL3/Ctail mutant exhibited higher apparent potency and increased maximal response to noradrenaline than those expressing the wild-type receptor. Phorbol ester-induced desensitization of the calcium response to noradrenaline was reduced in cells expressing the IL3 mutant and abolished in cells in which the Ctail or the IL3/Ctail were modified. In contrast, desensitization in response to preincubation with noradrenaline was unaffected in cells expressing the distinct receptor mutants. Noradrenaline-induced ERK phosphorylation was surprisingly increased in cells expressing IL3-modified receptors but not in those expressing receptors with the Ctail or IL3/Ctail substitutions. Our data indicate that phosphorylation sites in the IL3 and Ctail domains mediate and regulate α1B-adrenergic receptor function. Phorbol ester-induced desensitization seems to be closely associated with receptor phosphorylation, whereas noradrenaline-induced desensitization likely involves other elements.

Linking the shifts in the metabolically active microbiota in a UASB and hybrid anaerobic-aerobic bioreactor for swine wastewater treatment.

Due to the high concentration of pollutants, swine wastewater needs to be treated prior to disposal. The combination of anaerobic and aerobic technologies in one hybrid system allows to obtain higher removal efficiencies compared to those achieved via conventional biological treatment, and the performance of a hybrid system depends on the microbial community in the bioreactor. Here, we evaluated the community assembly of an anaerobic-aerobic hybrid reactor for swine wastewater treatment. Sequencing of partial 16S rRNA coding genes was performed using Illumina from DNA and retrotranscribed RNA templates (cDNA) extracted from samples from both sections of the hybrid system and from a UASB bioreactor fed with the same swine wastewater influent. Proteobacteria and Firmicutes were the dominant phyla and play a key role in anaerobic fermentation, followed by Methanosaeta and Methanobacterium. Several differences were found in the relative abundances of some genera between the DNA and cDNA samples, indicating an increase in the diversity of the metabolically active community, highlighting Chlorobaculum, Cladimonas, Turicibacter and Clostridium senso stricto. Nitrifying bacteria were more abundant in the hybrid bioreactor. Beta diversity analysis revealed that the microbial community structure significantly differed among the samples (p < 0.05) and between both anaerobic treatments. The main predicted metabolic pathways were the biosynthesis of amino acids and the formation of antibiotics. Also, the metabolism of C5-branched dibasic acid, Vit B5 and CoA, exhibited an important relationship with the main nitrogen-removing microorganisms. The anaerobic-aerobic hybrid bioreactor showed a higher ammonia removal rate compared to the conventional UASB system. However, further research and adjustments are needed to completely remove nitrogen from wastewater.

G protein-coupled receptors as therapeutic target.

Receptors are proteins coded by DNA, some of which have already been crystalized, thus allowing the details of their structure at the atomic level and some aspects of their function to be known. This review focuses on the most diverse and abundant family of receptors, G protein-coupled receptors. This family of receptors recognizes and mediates the action of several endogenous ligands (hormones, neurotransmitters, growth factors and local hormones) and also intervenes in the pathogenesis of various diseases, which is why they are targeted by approximately 30 to 40% of medications that are used in daily clinical practice and of various illegal drugs as well. X-ray crystallography is one of the essential tools that has allowed to observe the structure of these receptors in the amino acids that participate in this interaction, which allows to know the binding site of the endogenous ligand and of synthetic molecules that act on them to modulate their action. Molecular modeling or "docking" is also a computational bioinformatics tool that supports research on receptor-ligand binding, which allows the design and development of increasingly specific drugs. These developments have brought along significant changes in fundamental pharmacodynamic concepts.

Los receptores son proteínas codificadas por el ADN, algunos de los cuales ya han sido cristalizados, lo que permite conocer los detalles de su estructura a nivel atómico y algunos aspectos de su función. Esta revisión se enfoca en los más diversos y abundantes, los receptores acoplados a la proteína G. Esta familia de receptores reconoce y media la acción de varios ligandos endógenos (hormonas, neurotransmisores, factores de crecimiento y hormonas locales) y también interviene en la patogenia de diversas enfermedades, por lo que son el blanco terapéutico de aproximadamente 30 a 40 % de los medicamentos que se emplean en la práctica clínica cotidiana y de diversas drogas ilegales. La cristalografía de rayos X es una de las herramientas clave que ha permitido observar la estructura de estos receptores en los aminoácidos que participan en esta interacción, lo que posibilita conocer el sitio de unión del ligando endógeno y de moléculas sintéticas que actúan sobre ellos para modular su acción. El modelado molecular es también una herramienta bioinformática computacional que apoya la investigación sobre la unión receptor-ligando, que hace posible el diseño y desarrollo de fármacos cada vez más específicos. A estos desarrollos se suman importantes cambios en los conceptos farmacodinámicos fundamentales.

Changes in the prokaryotic diversity in response to hydrochemical variations during an acid mine drainage passive treatment.

Acid mine drainage (AMD) causes major environmental problems and consequently, several treatments are proposed, favoring the passive systems because of their many advantages. The main goal of these procedures is the neutralization and removal of potentially toxic elements (PTE), yet little is known about the changes in the microbial assemblages in response to the hydrochemical variations during the treatments. Therefore, the main objective of this research was to determine the changes in the diversity and structure of the prokaryotic assemblages in a hybrid abiotic and biological (wetland) passive treatment system. The 16S rRNA gene survey showed that the AMD coming from the mine (pH 2.6) was mainly composed of acidophilic genera such as Acidithiobacillus, Leptospirillum, Ferritrophicum, and Cuniculiplasma (up to 76 % relative abundance). In the abiotic treatment, Acidiphilium was dominant in the sections with limestone filters (pH 2.2-4.8), followed by Limnobacter in the subsequent dolomite/limestone and phosphoric rock filters (pH 5.2-5.8). In these abiotic passive treatment sections, the microbial assemblage showed a limited diversity and richness. However, when the treated AMD reached the two final wetlands (pH ~6.8), the microbial diversity and richness increased, suggesting that further bioattenuation mechanisms might be occurring. Limnobacter and Novosphingobium were the main bacterial genera in the water samples of the wetland sections (Arundo donax). These changes in the composition of the microbial assemblages were highly correlated with the pH and Eh values during the treatment (p-value <0.001); however, the concentration of metal(loid)s such as Al, Cd, Fe, Mn, Ni, and Zn were also significantly related (p-value <0.05). In conclusion, the studied passive AMD treatment system enhanced the chemical quality of the treated AMD, showing high removal efficiencies for Al and Fe (> 99 %), and increasing the microbial diversity and richness in the effluent.

Los receptores acoplados a proteínas G como blanco terapéutico / G protein-coupled receptors as therapeutic target

Resumen Los receptores son proteínas codificadas por el ADN, algunos de los cuales ya han sido cristalizados, lo que permite conocer los detalles de su estructura a nivel atómico y algunos aspectos de su función. Esta revisión se enfoca en los más diversos y abundantes, los receptores acoplados a la proteína G. Esta familia de receptores reconoce y media la acción de varios ligandos endógenos (hormonas, neurotransmisores, factores de crecimiento y hormonas locales) y también interviene en la patogenia de diversas enfermedades, por lo que son el blanco terapéutico de aproximadamente 30 a 40 % de los medicamentos que se emplean en la práctica clínica cotidiana y de diversas drogas ilegales. La cristalografía de rayos X es una de las herramientas clave que ha permitido observar la estructura de estos receptores en los aminoácidos que participan en esta interacción, lo que posibilita conocer el sitio de unión del ligando endógeno y de moléculas sintéticas que actúan sobre ellos para modular su acción. El modelado molecular es también una herramienta bioinformática computacional que apoya la investigación sobre la unión receptor-ligando, que hace posible el diseño y desarrollo de fármacos cada vez más específicos. A estos desarrollos se suman importantes cambios en los conceptos farmacodinámicos fundamentales.

Abstract Receptors are proteins coded by DNA, some of which have already been crystalized, thus allowing the details of their structure at the atomic level and some aspects of their function to be known. This review focuses on the most diverse and abundant family of receptors, G protein-coupled receptors. This family of receptors recognizes and mediates the action of several endogenous ligands (hormones, neurotransmitters, growth factors and local hormones) and also intervenes in the pathogenesis of various diseases, which is why they are targeted by approximately 30 to 40% of medications that are used in daily clinical practice and of various illegal drugs as well. X-ray crystallography is one of the essential tools that has allowed to observe the structure of these receptors in the amino acids that participate in this interaction, which allows to know the binding site of the endogenous ligand and of synthetic molecules that act on them to modulate their action. Molecular modeling or "docking" is also a computational bioinformatics tool that supports research on receptor-ligand binding, which allows the design and development of increasingly specific drugs. These developments have brought along significant changes in fundamental pharmacodynamic concepts.

Prokaryotic diversity across a pH gradient in the "El Chichón" crater-lake: a naturally thermo-acidic environment.

The "El Chichón" crater-lake in Mexico is a thermo-acidic environment whose microorganisms have been scarcely studied. In this study, we surveyed the prokaryotic communities by amplicon sequencing of the 16S rRNA gene considering samples of sediment and water collected within a pH/temperature gradient (pH 1.9-5.1, 38-89 °C). Further, we interpreted these results within a physicochemical context. The composition of the microbial assemblage differed significantly between the sediments and water. Sediment communities were different in the site with the highest temperature and lower pH value compared to the other ones sampled, while those in the water were relatively similar at all points. Most of the genera found were related to Alicyclobacillus, Acinetobacter, Bacillus, Mesoaciditoga, Methanothermobacter, Desulfitobacterium, Therminicanus, Kyrpidia, Paenibacillus, Thermoanaerobacterium, and Gelria. Some of these genera are known by their thermo-acidic tolerant capacities with flexible metabolisms to use diverse electron donor/acceptors (S or Fe), while others are thermo(acid)philes that mainly occur in the most extreme sites of the lake. These results show the presence of a microbial community adapted to the changing conditions of a very dynamic crater-lake, that include chemoorganotrophs and chemolithotrophs.

Interplay of microbial communities with mineral environments in coralline algae.

Coralline algae are worldwide carbonate builders, considered to be foundational species and biodiversity hotspots. Coralline habitats face increasing pressure from human activities and effects related to Global Change, yet their ecological properties and adaptive responses remain poorly understood. The relationships of the algal microbiota with the mineral bioconstructions, as well as plasticity and resilience of coralline holobionts in a changing environment, are of particular interest. In the Gulf of California, Neogoniolithon trichotomum (Rhodophyta) is the main carbonate builder in tidal pools. We performed a multi-disciplinary assessment of the N. trichotomum microstructure using XRD, SEM microscopy and SR-FTIR spectromicroscopy. In the algal perithallus, magnesium-calcite and aragonite were spatially segregated and embedded in a polysaccharide matrix (rich in sulfated polysaccharides). Mg-calcites (18-19 mol% Mg) were the main mineral components of the thallus overall, followed by iron carbonates related to dolomite (ankerite) and siderite. Minerals of late evaporitic sequences (sylvite and bischofite) were also present, suggesting potential halophilic microenvironments within the algal thalli. The diverse set of abundant halophilic, halotolerant and oligotrophic taxa, whose abundance increase in the summer, further suggests this condition. We created an integrated model, based on environmental parameters and the microbiota distribution, that identified temperature and nutrient availability (particularly nitrate and silicate) as the main parameters related to specific taxa patterns. Among these, Hahella, Granulossicoccus, Ferrimonas, Spongiibacteraceae and cyanobacterial Xenococcaceae and Nostocaceae change significantly between seasons. These bacterial components might play relevant roles in algal plasticity and adaptive responses to a changing environment. This study contributes to the understanding of the interplay of the prokaryotic microbiota with the mineral microenvironments of coralline algae. Because of their carbonates with potential resistance to dissolution in a higher pCO2 world and their seasonally dynamic bacteria, coralline algae are relevant targets to study coastal resilience and carbonated systems responses to changing environments.

Assessing the Diversity of Benthic Sulfate-Reducing Microorganisms in Northwestern Gulf of Mexico by Illumina Sequencing of dsrB Gene.

This study investigates the community composition, structure, and abundance of sulfate-reducing microorganisms (SRM) in surficial sediments of the Northwestern Gulf of Mexico (NWGoM) along a bathymetric gradient. For these purposes, Illumina sequencing and quantitative PCR (qPCR) of the dissimilatory sulfite reductase gene beta subunit (dsrB gene) were performed. Bioinformatic analyses indicated that SRM community was predominantly composed by members of Proteobacteria and Firmicutes across all the samples. However, Actinobacteria, Thermodesulfobacteria, and Chlorobi were also detected. Phylogenetic analysis indicated that unassigned dsrB sequences were related to Deltaproteobacteria and Nitrospirota superclusters, Euryarchaeota, and to environmental clusters. PCoA ordination revealed that samples clustered in three different groups. PERMANOVA indicated that water depth, temperature, redox, and nickel and cadmium content were the main environmental drivers for the SRM communities in the studied sites. Alpha diversity and abundance of SRM were lower for deeper sites, suggesting decreasing sulfate reduction activity with respect to water depth. This study contributes with the understanding of distribution and composition of dsrAB-containing microorganisms involved in sulfur transformations that may contribute to the resilience and stability of the benthic microbial communities facing metal and hydrocarbon pollution in the NWGoM, a region of recent development for oil and gas drilling.

Temporal analysis of the microbial communities in a nitrate-contaminated aquifer and the co-occurrence of anammox, n-damo and nitrous-oxide reducing bacteria.

Groundwater-N pollution derives from agricultural and urban activities, and compromises water quality in shallow aquifers, putting human and environmental health at risk. Nonetheless, subsurface microbiota can transform dissolved inorganic nitrogen into N2. In this study, we surveyed the microbial community of a shallow aquifer by sampling one well, one piezometer and a spring within an agricultural area that receives N-inputs of more than 700 kg/ha per year through irrigation with wastewater. The survey was conducted during a year with a 16S rRNA next-gen approach. In parallel, we quantified the number of gene copies and transcripts related to anaerobic ammonium oxidation (anammox, hzo), nitrite-dependent anaerobic methane oxidation (n-damo, nod and pmoA) and nitrous oxide reduction (last step of denitrification, nosZ), during the dry and rainy seasons. Our results showed that the groundwater samples had 17.7 to 22.5 mg/L of NO3--N. The bacterial and archaeal community structure was distinctive at each site, and it remained relatively stable over time. We verified the co-occurrence of N-transforming bacteria, which was correlated with the concentration of NO2-/NO3- and ORP/DO values (DO: ~3.0 mg/L). Our analyses suggest that these conditions may allow the presence of nitrifying microorganisms which can couple with anammox, n-damo and denitrifying bacteria in interrelated biogeochemical pathways. Gene density (as the number of gene copies per litre) was lower in the rainy season than in the dry season, possibly due to dilution by rainwater infiltration. Yet, the numbers of hzo gene copies here found were similar to those reported in oceanic oxygen minimum zones and in a carbonate-rock aquifer. The transcript sequences showed that Candidatus Brocadia spp. (anammox), Candidatus Methylomirabilis spp. (n-damo) and autotrophic denitrifying Betaproteobacteria coexist in the groundwater environment, with the potential to attenuate the concentration of dissolved inorganic nitrogen by reducing it to N2 rather than N2O; delivering thus, an important ecosystem service to remove contaminants.

Isolation of a freshwater microalgae and its application for the treatment of wastewater and obtaining fatty acids from tilapia cultivation.

In the present study, isolated strains of the microalgae Chlamydomonas sp. (CH) and Chlorella vulgaris (CV) were used to treat aquaculture wastewater and to obtain fatty acids and from a fattening culture of tilapia. The microalgae were cultivated for 11 days in tubular photobioreactors with an operating volume of 2 L, constant aeration and illumination. High removal rates of NO3- and PO43- were achieved for both Chlamydomonas sp. (84.7% and 96%, respectively) and Chlorella vulgaris (94.6 and 97.9%, respectively). The maximum biomass productivity achieved by Chlamydomonas sp. was 0.06 and 0.10 gL-1d-1 for Chlorella vulgaris. Therefore, tilapia wastewater contained the necessary nutrient concentration for algal growth and development. Chlamydomonas sp. biomass lipid content was 69%, while that of Chlorella vulgaris was 40%. The lipid profile of both microalgae was abundant in palmitic acid (78% for Chlamydomonas sp. and 35% for Chlorella vulgaris). This fatty acid is suitable for biodiesel production. Tilapia wastewater is low-cost alternative culture medium as it contains the necessary nutrient concentration for microalgae development and growth.

Glycogen Synthase Kinase-3 modulates α1A-adrenergic receptor action and regulation.

The possibility that glycogen synthase kinase 3 (GSK3) could modulate α1A-adrenergic receptor (α1A-AR) function and regulation was tested employing LNCaP and HEK293 cells transfected to express the enhanced green fluorescent protein-tagged human α1A-AR. Receptor phosphorylation and internalization, intracellular free calcium, α1A-AR-GSK3 colocalization, and coimmunoprecipitation were studied. The effects of the pharmacological GSK3 inhibitor, SB-216763, and the coexpression of a dominant-negative mutant of this kinase, as well as the signaling, desensitization, and internalization of receptors with S229, S258, S352, and S381 substitutions for alanine or aspartate, were also determined. SB-216763 inhibited agonist- and phorbol myristate acetate (PMA)-mediated α1A-AR phosphorylation, reduced oxymetazoline-induced desensitization, and magnified that induced by PMA. Agonists and PMA increased receptor-GSK3 colocalization and coimmunoprecipitation. Expression of a dominant-negative GSK3 mutant reduced agonist- but not PMA-induced receptor internalization. α1A-AR with the GSK3 putative target sites mutated to alanine exhibited reduced phosphorylation and internalization in response to agonists and increased PMA-induced desensitization. Agonist-induced, but not PMA-induced, receptor-ß arrestin intracellular colocalization was diminished in cells expressing the GSK3 putative target sites mutated to alanine. Our data indicated that GSK3 exerts a dual action on α1A-AR participating in agonist-mediated desensitization and internalization and avoiding PMA-induced desensitization.

The α1-adrenoceptor-mediated human hyperplastic prostate cells proliferation is impaired by EGF receptor inhibition.

Benign prostatic hyperplasia (BPH) is an aging-related and progressive disease linked to an up-regulation of α1-adrenoceptors. The participation of EGF receptors (EGFR) in the GPCRs' signalosome has been described but so far data about the contribution of these receptors to prostatic stromal hyperplasia are scanty. We isolated and cultured vimentin-positive prostate stromal cells obtained from BPH patients. According to intracellular Ca2+ measurements, cell proliferation and Western blotting assays, these cultured hyperplastic stromal cells express functional α1-adrenoceptors and EGFR, and proliferate in response to the α1-adrenoceptor agonist phenylephrine. Interestingly, in these cells the inhibition of EGFR signaling with GM6001, CRM197, AG1478 or PD98059 was associated with full blockage of α1-adrenoceptor-mediated cell proliferation, while cell treatment with each inhibitor alone did not alter basal cell growth. Moreover, the co-incubation of AG1478 (EGFR inhibitor) with α1A/α1D-adrenoceptor antagonists showed no additive inhibitory effect. These findings highlight a putative role of EGFR signaling to α1-adrenoceptor-mediated human prostate hyperplasia, suggesting that the inhibition of this transactivation cascade could be useful to reduce BPH progression.

Sites phosphorylated in human α1B-adrenoceptors in response to noradrenaline and phorbol myristate acetate.

Phosphorylation of the human α1B-adrenergic receptor (fused with the green fluorescent protein) was studied employing the inducible Flp-ln HEK293 T-Rex system for expression. Serine/alanine substitutions were performed in five sites corresponding to those previously identified as phosphorylation targets in the hamster ortholog. Desensitization was decreased in these mutants but receptor phosphorylation was still clearly detected. The protein phosphorylation of the wild-type receptor (fused to the green fluorescent protein) was studied, using mass spectrometry, under baseline and stimulated conditions (noradrenaline or phorbol myristate acetate). Basal phosphorylation was detected at sites located at the intracellular loop 3 and carboxyl terminus, and the number of sites detected increased under agonist activation and stimulation of protein kinase C. The phosphorylation patterns differed under the distinct conditions. Three of the phosphorylation sites detected in this work corresponded to those observed in the hamster receptor. The phosphorylation sites detected included the following: a) at the intracellular loop 3: serines 246, 248, 257, 267, and 277; and threonines 252, 264, and 268, and b) at the carboxyl terminus: serines 396, 400, 402, 406, 423, 425, 427, 455, and 470, and threonines 387, 392, 420, and 475. Our data indicate that complex phosphorylation patterns exist and suggest the possibility that such differences could be relevant in receptor function and subcellular localization.

Extant microbial communities in the partially desiccated Rincon de Parangueo maar crater lake in Mexico.

Rincon de Parangueo is a maar where a perennial lake was present until the 1980s. A conspicuous feature of the lake's sediments is the presence of bioherms and organo-sedimentary deposits produced by microbial communities. The gradual lake desiccation during the last 40 years has produced dramatic environmental changes inside the maar basin, which resulted in the formation of a highly saline-alkaline system with extant microorganisms. In this paper we succinctly describe the geologic setting where the microbial communities have developed inside of the maar crater and the results obtained from high-throughput sequencing methods to characterize the microbial component (Bacteria, Eukarya and Archaea) in endolithic mats of calcareous sediments, and microbial mats and free-living microorganisms in the soda ponds. The studied sites displayed different microbial communities with a diverse number of phylotypes belonging to Bacteria and Eukarya, contrasting with a much less diverse component in Archaea. The sequences here detected were related to environmental sequences from sites with extreme life conditions such as high alkalinity (alkaliphiles), high salinity (halophiles) and high temperature (thermophiles). Moreover, our results indicate an important unexplored endemic microbial biodiversity in the vestiges of the former lake that need to be studied.

The role of ß-arrestins in G protein-coupled receptor heterologous desensitization: A brief story.

G protein-coupled receptors (GPCRs) are transmembrane proteins that have an important impact in a myriad of cellular functions. Posttranslational modifications on GPCRs are a key processes that allow these proteins to recruit other intracellular molecules. Among these modifications, phosphorylation is the most important way of desensitization of these receptors. Several research groups have described two different desensitization mechanisms: heterologous and homologous desensitization. The first one involves the phosphorylation of the receptors by protein kinases, such as PKC, following the desensitization and internalization of the receptor, while the second one involves the phosphorylation of the receptors by GRKs, allowing for the receptor to recruit ß-arrestins to be desensitized and internalized. Interestingly, a few number of studies have described the participation of ß-arrestins during the heterologous desensitization process. Hence, the aim of this review is to briefly explore the role that ß-arrestins play during the heterologous desensitization of several GPCRs.

Distinct phosphorylation sites/clusters in the carboxyl terminus regulate α1D-adrenergic receptor subcellular localization and signaling.

The human α1D-adrenergic receptor is a seven transmembrane-domain protein that mediates many of the physiological actions of adrenaline and noradrenaline and participates in the development of hypertension and benign prostatic hyperplasia. We recently reported that different phosphorylation patterns control α1D-adrenergic receptor desensitization. However, to our knowledge, there is no data regarding the role(s) of this receptor's specific phosphorylation residues in its subcellular localization and signaling. In order to address this issue, we mutated the identified phosphorylated residues located on the third intracellular loop and carboxyl tail. In this way, we experimentally confirmed α1D-AR phosphorylation sites and identified, in the carboxyl tail, two groups of residues in close proximity to each other, as well as two individual residues in the proximal (T442) and distal (S543) regions. Our results indicate that phosphorylation of the distal cluster (T507, S515, S516 and S518) favors α1D-AR localization at the plasma membrane, i. e., substitution of these residues for non-phosphorylatable amino acids results in the intracellular localization of the receptors, whereas phospho-mimetic substitution allows plasma membrane localization. Moreover, we found that T442 phosphorylation is necessary for agonist- and phorbol ester-induced receptor colocalization with ß-arrestins. Additionally, we observed that substitution of intracellular loop 3 phosphorylation sites for non-phosphorylatable amino acids resulted in sustained ERK1/2 activation; additional mutations in the phosphorylated residues in the carboxyl tail did not alter this pattern. In contrast, mobilization of intracellular calcium and receptor internalization appear to be controlled by the phosphorylation of both third-intracellular-loop and carboxyl terminus-domain residues. In summary, our data indicate that a) both the phosphorylation sites present in the third intracellular loop and in the carboxyl terminus participate in triggering calcium signaling and in turning-off α1D-AR-induced ERK activation; b) phosphorylation of the distal cluster appears to play a role in receptor's plasma membrane localization; and c) T442 appears to play a critical role in receptor phosphorylation and receptor-ß-arrestin colocalization.

Microbial distribution and turnover in Antarctic microbial mats highlight the relevance of heterotrophic bacteria in low-nutrient environments.

Maritime Antarctica has shown the highest increase in temperature in the Southern Hemisphere. Under this scenario, biogeochemical cycles may be altered, resulting in rapid environmental change for Antarctic biota. Microbes that drive biogeochemical cycles often form biofilms or microbial mats in continental meltwater environments. Limnetic microbial mats from the Fildes Peninsula were studied using high-throughput 16S rRNA gene sequencing. Mat samples were collected from 15 meltwater stream sites, comprising a natural gradient from ultraoligotrophic glacier flows to meltwater streams exposed to anthropogenic activities. Our analyses show that microbial community structure differences between mats are explained by environmental NH4+, NO3-, DIN, soluble reactive silicon and conductivity. Microbial mats living under ultraoligotrophic meltwater conditions did not exhibit a dominance of cyanobacterial photoautotrophs, as has been documented for other Antarctic limnetic microbial mats. Instead, ultraoligotrophic mat communities were characterized by the presence of microbes recognized as heterotrophs and photoheterotrophs. This suggests that microbial capabilities for recycling organic matter may be a key factor to dwell in ultra-low nutrient conditions. Our analyses show that phylotype level assemblages exhibit coupled distribution patterns in environmental oligotrophic inland waters. The evaluation of these microbes suggests the relevance of reproductive and structural strategies to pioneer these psychrophilic ultraoligotrophic environments.

Adrenergic signaling molecular complexes. / Complejos moleculares de la señalización adrenérgica.

Adrenaline and noradrenaline bind to membrane receptors of the superfamily of G protein-coupled receptors (GPCR) in target cells, where they modulate physiological responses such as metabolism, vasoconstriction, vasodilation and proliferation. Alteration in their function is associated with conditions such as hypertension, benign prostatic hyperplasia and cardiac hypertrophy. In response to adrenaline, receptors form signaling complexes, which enables adrenergic action to be specific, rapid and efficient. These signaling complexes or signalosomes are composed of kinases, phosphatases, and adapter and scaffold proteins, which together modulate the receptor function. Manipulation of each protein-protein interaction of the adrenergic signaling complex emerges as a promising therapeutic strategy for the design of drugs that modulate adrenergic action and help to define its pathophysiological significance. An important biological model to perform these investigations is the heart, since it expresses all adrenergic receptors; to date, several heart signalosomes have been described. Mass spectrometry (proteomics), genetic manipulation and biochemical assays, such as two-hybrid and co-immunoprecipitation assays, are tools that are used in these studies.

La adrenalina y la noradrenalina se unen a receptores membranales de la superfamilia de receptores acoplados a proteínas G (GPCR) en las células blanco, donde modulan respuestas fisiológicas tales como el metabolismo, vasoconstricción, vasodilatación y proliferación. La alteración en su función está asociada con hipertensión, hiperplasia prostática benigna e hipertrofia cardiaca. En respuesta a la adrenalina, los receptores forman complejos de señalización, lo que permite que la acción adrenérgica sea específica, rápida y eficiente. Estos complejos de señalización o signalosomas están integrados por cinasas, fosfatasas, proteínas adaptadoras y de andamio, que en conjunto modulan la función del receptor. La manipulación de cada interacción proteína-proteína del complejo de señalización adrenérgico emerge como una estrategia terapéutica prometedora para el diseño de fármacos que modulen la acción adrenérgica y ayuden a definir su significado fisiopatológico. Un modelo biológico importante para realizar estos estudios es el corazón, ya que expresa todos los receptores adrenérgicos; en la actualidad se han descrito varios signalosomas cardiacos. La espectrometría de masas (proteómica), manipulación genética y ensayos bioquímicos como el doble híbrido o la coinmunoprecipitación son herramientas que se emplean en estos estudios.

Dissecting the signaling features of the multi-protein complex GPCR/ß-arrestin/ERK1/2.

G protein-coupled receptors (GPCRs) have emerged as key biological entities that regulate a plethora of physiological processes and participate in the onset and development of many diseases. Moreover, these receptors are important targets of almost 25% of the current therapeutic drugs in the market. Upon agonist binding, GPCRs activate a great number of signaling pathways, resulting in important cellular events like gene transcription, survival, proliferation and differentiation. In order to activate such events, GPCRs interact with a variety of scaffold and molecular entities, particularly with G proteins, but also with ß-arrestins and the extracellular signal-regulated kinases 1 and 2 (ERK1/2) pathway, forming unique signaling modules. The aim of this review is to analyze the signaling features of the multi-protein complex GPCR-ß-arrestin-ERK1/2, a unique signaling module that has received considerable attention from different research groups due to its molecular and physiological roles in diverse cellular contexts.