Brain-gut photobiomodulation restores cognitive alterations in chronically stressed mice through the regulation of Sirt1 and neuroinflammation.

BACKGROUND: Chronic stress is an important risk factor for the development of major depressive disorder (MDD). Recent studies have shown microbiome dysbiosis as one of the pathogenic mechanisms associated with MDD. Thus, it is important to find novel non-pharmacological therapeutic strategies that can modulate gut microbiota and brain activity. One such strategy is photobiomodulation (PBM), which involves the non-invasive use of light. OBJECTIVE/HYPOTHESIS: Brain-gut PBM could have a synergistic beneficial effect on the alterations induced by chronic stress. METHODS: We employed the chronic unpredictable mild stress (CUMS) protocol to induce a depressive-like state in mice. Subsequently, we administered brain-gut PBM for 6 min per day over a period of 3 weeks. Following PBM treatment, we examined behavioral, structural, molecular, and cellular alterations induced by CUMS. RESULTS: We observed that the CUMS protocol induces profound behavioral alterations and an increase of sirtuin1 (Sirt1) levels in the hippocampus. We then combined the stress protocol with PBM and found that tissue-combined PBM was able to rescue cognitive alterations induced by CUMS. This rescue was accompanied by a restoration of hippocampal Sirt1 levels, prevention of spine density loss in the CA1 of the hippocampus, and the modulation of the gut microbiome. PBM was also effective in reducing neuroinflammation and modulating the morphology of Iba1-positive microglia. LIMITATIONS: The molecular mechanisms behind the beneficial effects of tissue-combined PBM are not fully understood. CONCLUSIONS: Our results suggest that non-invasive photobiomodulation of both the brain and the gut microbiome could be beneficial in the context of stress-induced MDD.

VPS13A knockdown impairs corticostriatal synaptic plasticity and locomotor behavior in a new mouse model of chorea-acanthocytosis.

Chorea-acanthocytosis (ChAc) is an inherited neurodegenerative movement disorder caused by VPS13A gene mutations leading to the absence of protein expression. The striatum is the most affected brain region in ChAc patients. However, the study of the VPS13A function in the brain has been poorly addressed. Here we generated a VPS13A knockdown (KD) model and aimed to elucidate the contribution of VPS13A to synaptic plasticity and neuronal communication in the corticostriatal circuit. First, we infected primary cortical neurons with miR30-shRNA against VPS13A and analyzed its effects on neuronal plasticity. VPS13A-KD neurons showed a higher degree of branching than controls, accompanied by decreased BDNF and PSD-95 levels, indicative of synaptic alterations. We then injected AAV-KD bilaterally in the frontal cortex and two different regions of the striatum of mice and analyzed the effects of VPS13A-KD on animal behavior and synaptic plasticity. VPS13A-KD mice showed modification of the locomotor behavior pattern, with increased exploratory behavior and hyperlocomotion. Corticostriatal dysfunction in VPS13A-KD mice was evidenced by impaired striatal long-term depression (LTD) after stimulation of cortical afferents, which was partially recovered by BDNF administration. VPS13A-KD did not lead to neuronal loss in the cortex or the striatum but induced a decrease in the neuronal release of CX3CL1 and triggered a microglial reaction, especially in the striatum. Notably, CX3CL1 administration partially restored the impaired corticostriatal LTD in VPS13A-KD mice. Our results unveil the involvement of VPS13A in neuronal connectivity modifying BDNF and CX3CL1 release. Moreover, the involvement of VPS13A in synaptic plasticity and motor behavior provides key information to further understand not only ChAc pathophysiology but also other neurological disorders.

Nutrition and Health Programming and Outreach in Grocery Retail Settings: A Community Coalition in Action.

Grocery stores can provide a conducive environment for interventions targeting healthy eating and access to health services, particularly in low-income communities. A wide array of organizations deliver nutrition and related programs in community settings, but rarely in a coordinated fashion. Collaboration of local health promotion organizations with grocery stores could increase consumers' access to and selection of healthy foods and related services. This evaluation of the In-Store Programming and Outreach Coalition (IPOC) uses thematic analysis of first-person accounts from coalition members. To our knowledge, this is the first study of such a coalition. We present perspectives from six stakeholders about the IPOC strengths, challenges, and recommendations for strengthening the delivery of in-store interventions. Themes identified include partnership, increased client reach and cross-referrals, conflicting work schedules, leadership, and recommendations to identify coalition leaders and expand services to other grocery stores. We conclude that grocery stores can offer a suitable setting for programming and community outreach through coalitions.

Expression of Circulating MicroRNAs Associated with Obesity and their Relationships with Biochemical Parameters and Health-related Physical Fitness in Children 6 to 10 Years Old in Cali, Colombia.

BACKGROUND: Obesity is a public health problem worldwide; it has reached pandemic proportions in the last 40 years. Its prevalence in children and adolescents increased from 0.7% to 7.8% between 1975 and 2016. Recently, microRNAs (miRNAs) have been reported as regulatory factors related to molecular functions under different conditions. These can be used as biomarkers of a disease to estimate risks in the early stages. OBJECTIVE: This study aimed to determine the expression levels of miRNAs associated with childhood obesity and their relationships with biochemical parameters and Health-related Physical Fitness (HRPF). METHODS: This was a descriptive cross-sectional study in which a population of 40 children between 6 and 10 years of age of both sexes from Cali, Colombia, was evaluated; the children were classified as 20 normal-weight and 20 obese. Blood biochemistry, HRPF, and miRNA expression levels were determined (hsa-miR-122-5p, hsa-miR-15b-5p, hsa-miR-191-5p, hsa-miR-486-3p, hsa-miR-222-3p. Comparisons were made between the groups, miRNA associations between the studied variables, and linear regression analysis. RESULTS: Twenty normal-weight and 20 obese patients were evaluated. Both groups had an average age of eight years old. The miRNA hsa-miR-122-5p (p < 0.05) was overexpressed in the obese group. According to the linear regression analysis, the amount of adipose tissue may be associated with the production of miRNAs (hsa-miR-15b-5p, hsa-miR-222-3p, hsa-miR-122-5p, and hsamiR- 191-5p). CONCLUSION: Four miRNAs (hsa-miR-15b-5p, hsa-miR-222-3p, hsa-miR-122-5p, and hsa-miR- 191-5p) are associated with modifications in biochemical variables of HRPF in this group. Adipose tissue mass could be associated with the production of these miRNAs, thus making them biomarkers of childhood obesity risk.

GPC3-Unc5 receptor complex structure and role in cell migration.

Neural migration is a critical step during brain development that requires the interactions of cell-surface guidance receptors. Cancer cells often hijack these mechanisms to disseminate. Here, we reveal crystal structures of Uncoordinated-5 receptor D (Unc5D) in complex with morphogen receptor glypican-3 (GPC3), forming an octameric glycoprotein complex. In the complex, four Unc5D molecules pack into an antiparallel bundle, flanked by four GPC3 molecules. Central glycan-glycan interactions are formed by N-linked glycans emanating from GPC3 (N241 in human) and C-mannosylated tryptophans of the Unc5D thrombospondin-like domains. MD simulations, mass spectrometry and structure-based mutants validate the crystallographic data. Anti-GPC3 nanobodies enhance or weaken Unc5-GPC3 binding and, together with mutant proteins, show that Unc5/GPC3 guide migrating pyramidal neurons in the mouse cortex, and cancer cells in an embryonic xenograft neuroblastoma model. The results demonstrate a conserved structural mechanism of cell guidance, where finely balanced Unc5-GPC3 interactions regulate cell migration.

Postnatal Foxp2 regulates early psychiatric-like phenotypes and associated molecular alterations in the R6/1 transgenic mouse model of Huntington's disease.

Huntington's Disease (HD) is a devastating disorder characterized by a triad of motor, psychiatric and cognitive manifestations. Psychiatric and emotional symptoms appear at early stages of the disease which are consistently described by patients and caregivers among the most disabling. Here, we show for the first time that Foxp2 is strongly associated with some psychiatric-like disturbances in the R6/1 mouse model of HD. First, 4-week-old (juvenile) R6/1 mice behavioral phenotype was characterized by an increased impulsive-like behavior and less aggressive-like behavior. In this line, we identified an early striatal downregulation of Foxp2 protein starting as soon as at postnatal day 15 that could explain such deficiencies. Interestingly, the rescue of striatal Foxp2 levels from postnatal stages completely reverted the impulsivity-phenotype and partially the social impairments concomitant with a rescue of dendritic spine pathology. A mass spectrometry study indicated that the rescue of spine loss was associated with an improvement of several altered proteins related with cytoskeleton dynamics. Finally, we reproduced and mimicked the impulsivity and social deficits in wild type mice by reducing their striatal Foxp2 expression from postnatal stages. Overall, these results imply that early postnatal reduction of Foxp2 might contribute to the appearance of some of the early psychiatric symptoms in HD.

Transmission of gram-negative antibiotic-resistant bacteria following differing exposure to antibiotic-resistance reservoirs in a rural community: a modelling study for bloodstream infections.

Exposure to community reservoirs of gram-negative antibiotic-resistant bacteria (GN-ARB) genes poses substantial health risks to individuals, complicating potential infections. Transmission networks and population dynamics remain unclear, particularly in resource-poor communities. We use a dynamic compartment model to assess GN-ARB transmission quantitatively, including the susceptible, colonised, infected, and removed populations at the community-hospital interface. We used two side streams to distinguish between individuals at high- and low-risk exposure to community ARB reservoirs. The model was calibrated using data from a cross-sectional cohort study (N = 357) in Chile and supplemented by existing literature. Most individuals acquired ARB from the community reservoirs (98%) rather than the hospital. High exposure to GN-ARB reservoirs was associated with 17% and 16% greater prevalence for GN-ARB carriage in the hospital and community settings, respectively. The higher exposure has led to 16% more infections and attributed mortality. Our results highlight the need for early-stage identification and testing capability of bloodstream infections caused by GN-ARB through a faster response at the community level, where most GN-ARB are likely to be acquired. Increasing treatment rates for individuals colonised or infected by GN-ARB and controlling the exposure to antibiotic consumption and GN-ARB reservoirs, is crucial to curve GN-ABR transmission.

Association between circulating microRNAs and the metabolic syndrome in adult populations: A systematic review.

BACKGROUND: Circulating microRNAs (miRNAs) are one of the most abundant classes of gene regulatory molecules, and had been associated to the metabolic syndrome, higher BMI, dyslipidemia and diabetes mellitus. In this sense, miRNAs could help to understand the mechanism behind the development of metabolic syndrome. OBJECTIVE: To determine the relationship between circulating microRNAs and the metabolic syndrome in adult population. METHODS: We performed a systematic review according to the recommendations of the Cochrane Collaboration and following the PRISMA Statement. The results were grouped for miRNAs levels in MetS and metabolic variables included in MetS and their statistic association with miRNAs levels. RESULTS: We finally included sixteen studies with a total of 7195 individuals. We found 47 miRNAs reported to be related to metabolic syndrome (p < 0,05) and 98 associated with the metabolic alterations included in its diagnostic (p < 0,05). Forty-nine miRNAs levels were described as relate to insulin resistance, 29 with high triglycerides, 35 with hypertension, 28 with obesity, and 16 miRNAs with cholesterol HDL(p < 0,05). Changes in levels of miR-505-5p, miR-148a-3p, miR-19b-3p, miR-320b, miR-342-3p, miR-197-3p, miR-192-5p, miR-122-5p, miR-103, miR-130a, miR-155-5p and miR-375, were reported as significant in more than one study. The results only included a descriptive synthesis, clinical heterogeneity did not allow a meta-analysis. CONCLUSION: The findings on the current systematic review suggests a possible relationship between miRNAs with metabolic syndrome and metabolic traits. This association could help to understand the mechanism behind the develop of the metabolic syndrome. However, more studies are necessary for further validation.

Unraveling the Spatiotemporal Distribution of VPS13A in the Mouse Brain.

Loss-of-function mutations in the human vacuolar protein sorting the 13 homolog A (VPS13A) gene cause Chorea-acanthocytosis (ChAc), with selective degeneration of the striatum as the main neuropathologic feature. Very little is known about the VPS13A expression in the brain. The main objective of this work was to assess, for the first time, the spatiotemporal distribution of VPS13A in the mouse brain. We found VPS13A expression present in neurons already in the embryonic stage, with stable levels until adulthood. VPS13A mRNA and protein distributions were similar in the adult mouse brain. We found a widespread VPS13A distribution, with the strongest expression profiles in the pons, hippocampus, and cerebellum. Interestingly, expression was weak in the basal ganglia. VPS13A staining was positive in glutamatergic, GABAergic, and cholinergic neurons, but rarely in glial cells. At the cellular level, VPS13A was mainly located in the soma and neurites, co-localizing with both the endoplasmic reticulum and mitochondria. However, it was not enriched in dendritic spines or the synaptosomal fraction of cortical neurons. In vivo pharmacological modulation of the glutamatergic, dopaminergic or cholinergic systems did not modulate VPS13A concentration in the hippocampus, cerebral cortex, or striatum. These results indicate that VPS13A has remarkable stability in neuronal cells. Understanding the distinct expression pattern of VPS13A can provide relevant information to unravel pathophysiological hallmarks of ChAc.

FLRT3 Marks Direction-Selective Retinal Ganglion Cells That Project to the Medial Terminal Nucleus.

The mammalian retina extracts a multitude of diverse features from the visual scene such as color, contrast, and direction of motion. These features are transmitted separately to the brain by more than 40 different retinal ganglion cell (RGC) subtypes. However, so far only a few genetic markers exist to fully characterize the different RGC subtypes. Here, we present a novel genetic Flrt3-CreERT2 knock-in mouse that labels a small subpopulation of RGCs. Using single-cell injection of fluorescent dyes in Flrt3 positive RGCs, we distinguished four morphological RGC subtypes. Anterograde tracings using a fluorescent Cre-dependent Adeno-associated virus (AAV) revealed that a subgroup of Flrt3 positive RGCs specifically project to the medial terminal nucleus (MTN), which is part of the accessory optic system (AOS) and is essential in driving reflex eye movements for retinal image stabilization. Functional characterization using ex vivo patch-clamp recordings showed that the MTN-projecting Flrt3 RGCs preferentially respond to downward motion in an ON-fashion. These neurons distribute in a regular pattern and most of them are bistratified at the level of the ON and OFF bands of cholinergic starburst amacrine cells where they express the known ON-OFF direction-selective RGC marker CART. Together, our results indicate that MTN-projecting Flrt3 RGCs represent a new functionally homogeneous AOS projecting direction-selective RGC subpopulation.

FLRT2 and FLRT3 Cooperate in Maintaining the Tangential Migratory Streams of Cortical Interneurons during Development.

Neuron migration is a hallmark of nervous system development that allows gathering of neurons from different origins for assembling of functional neuronal circuits. Cortical inhibitory interneurons arise in the ventral telencephalon and migrate tangentially forming three transient migratory streams in the cortex before reaching the final laminar destination. Although migration defects lead to the disruption of inhibitory circuits and are linked to aspects of psychiatric disorders such as autism and schizophrenia, the molecular mechanisms controlling cortical interneuron development and final layer positioning are incompletely understood. Here, we show that mouse embryos with a double deletion of FLRT2 and FLRT3 genes encoding cell adhesion molecules exhibit an abnormal distribution of interneurons within the streams during development, which in turn, affect the layering of somatostatin+ interneurons postnatally. Mechanistically, FLRT2 and FLRT3 proteins act in a noncell-autonomous manner, possibly through a repulsive mechanism. In support of such a conclusion, double knockouts deficient in the repulsive receptors for FLRTs, Unc5B and Unc5D, also display interneuron defects during development, similar to the FLRT2/FLRT3 mutants. Moreover, FLRT proteins are chemorepellent ligands for developing interneurons in vitro, an effect that is in part dependent on FLRT-Unc5 interaction. Together, we propose that FLRTs act through Unc5 receptors to control cortical interneuron distribution in a mechanism that involves cell repulsion.SIGNIFICANCE STATEMENT Disruption of inhibitory cortical circuits is responsible for some aspects of psychiatric disorders such as schizophrenia or autism. These defects include interneuron migration during development. A crucial step during this process is the formation of three transient migratory streams within the developing cortex that determine the timing of interneuron final positioning and the formation of functional cortical circuits in the adult. We report that FLRT proteins are required for the proper distribution of interneurons within the cortical migratory streams and for the final laminar allocation in the postnatal cortex. These results expand the multifunctional role of FLRTs during nervous system development in addition to the role of FLRTs in axon guidance and the migration of excitatory cortical neurons.

TUNAR lncRNA Encodes a Microprotein that Regulates Neural Differentiation and Neurite Formation by Modulating Calcium Dynamics.

Long noncoding RNAs (lncRNAs) are regulatory molecules which have been traditionally considered as "non-coding". Strikingly, recent evidence has demonstrated that many non-coding regions, including lncRNAs, do in fact contain small-open reading frames that code for small proteins that have been called microproteins. Only a few of them have been characterized so far, but they display key functions in a wide variety of cellular processes. Here, we show that TUNAR lncRNA encodes an evolutionarily conserved microprotein expressed in the nervous system that we have named pTUNAR. pTUNAR deficiency in mouse embryonic stem cells improves their differentiation potential towards neural lineage both in vitro and in vivo. Conversely, pTUNAR overexpression impairs neuronal differentiation by reduced neurite formation in different model systems. At the subcellular level, pTUNAR is a transmembrane protein that localizes in the endoplasmic reticulum and interacts with the calcium transporter SERCA2. pTUNAR overexpression reduces cytoplasmatic calcium, consistent with a possible role of pTUNAR as an activator of SERCA2. Altogether, our results suggest that our newly discovered microprotein has an important role in neural differentiation and neurite formation through the regulation of intracellular calcium. From a more general point of view, our results provide a proof of concept of the role of lncRNAs-encoded microproteins in neural differentiation.

FLRTing Neurons in Cortical Migration During Cerebral Cortex Development.

During development, two coordinated events shape the morphology of the mammalian cerebral cortex, leading to the cortex's columnar and layered structure: the proliferation of neuronal progenitors and cortical migration. Pyramidal neurons originating from germinal zones migrate along radial glial fibers to their final position in the cortical plate by both radial migration and tangential dispersion. These processes rely on the delicate balance of intercellular adhesive and repulsive signaling that takes place between neurons interacting with different substrates and guidance cues. Here, we focus on the function of the cell adhesion molecules fibronectin leucine-rich repeat transmembrane proteins (FLRTs) in regulating both the radial migration of neurons, as well as their tangential spread, and the impact these processes have on cortex morphogenesis. In combining structural and functional analysis, recent studies have begun to reveal how FLRT-mediated responses are precisely tuned - from forming different protein complexes to modulate either cell adhesion or repulsion in neurons. These approaches provide a deeper understanding of the context-dependent interactions of FLRTs with multiple receptors involved in axon guidance and synapse formation that contribute to finely regulated neuronal migration.

Structural Basis of Teneurin-Latrophilin Interaction in Repulsive Guidance of Migrating Neurons.

Teneurins are ancient metazoan cell adhesion receptors that control brain development and neuronal wiring in higher animals. The extracellular C terminus binds the adhesion GPCR Latrophilin, forming a trans-cellular complex with synaptogenic functions. However, Teneurins, Latrophilins, and FLRT proteins are also expressed during murine cortical cell migration at earlier developmental stages. Here, we present crystal structures of Teneurin-Latrophilin complexes that reveal how the lectin and olfactomedin domains of Latrophilin bind across a spiraling beta-barrel domain of Teneurin, the YD shell. We couple structure-based protein engineering to biophysical analysis, cell migration assays, and in utero electroporation experiments to probe the importance of the interaction in cortical neuron migration. We show that binding of Latrophilins to Teneurins and FLRTs directs the migration of neurons using a contact repulsion-dependent mechanism. The effect is observed with cell bodies and small neurites rather than their processes. The results exemplify how a structure-encoded synaptogenic protein complex is also used for repulsive cell guidance.

Association Between Drug Burden Index and Functional and Cognitive Function in Patients with Multimorbidity.

BACKGROUND: Anticholinergic and sedative drugs are associated with adverse events such as cognitive and functional impairment in elderly. The Drug Burden Index (DBI) is a measure of an individual's total exposure to anticholinergic and sedative drugs. Objetive: The study aimed to evaluate the association between the total DBI and cognitive and functional impairment in patients with multimorbidity. SETTING: Patients with multimorbidity enrolled in the IMPACTO project. METHODS: Cross-sectional observational study. MAIN OUTCOME MEASURE: The anticholinergic and sedative exposure was calculated using DBI. The Pfeiffer Test (PT) was used for cognitive status and the Barthel Index (BI) for functional status. RESULTS: 336 patients were included (mean age 77.6 ± 8.7 years, 54.2% men and a mean of 11.5 ± 3.7 prescribed drugs). 180 patients (53.6%) exposed to anticholinergic and/or sedative drugs were identified. The median score obtained in PT was slightly higher in exposed patients (1 (IQR 0-2) and 2 (IQR 0-4), p = 0.082 in "non-exposed" and "exposed", respectively). The bivariate analysis showed an association [0.544 (95% CI 0.044-1.063, p = 0.03)]. The median obtained in the BI analysis was 85.0 (IQR 30.0) and 75.5 (IQR 42.5) p = 0.002, in "nonexposed" and "exposed", respectively. After the adjusted analysis, a relationship was obtained between both the variables [-9,558 (95% CI-15,794; -3,321, p = 0.03)]. CONCLUSION: Higher DBI is associated with the impairment of functional status and, slightly to the deterioration of cognitive function in patients with multimorbidity. DBI should be considered in patients with multimorbidity to optimize the pharmacological treatment of a group of special interest due to its vulnerability.

Regulation of Cerebral Cortex Folding by Controlling Neuronal Migration via FLRT Adhesion Molecules.

The folding of the mammalian cerebral cortex into sulci and gyri is thought to be favored by the amplification of basal progenitor cells and their tangential migration. Here, we provide a molecular mechanism for the role of migration in this process by showing that changes in intercellular adhesion of migrating cortical neurons result in cortical folding. Mice with deletions of FLRT1 and FLRT3 adhesion molecules develop macroscopic sulci with preserved layered organization and radial glial morphology. Cortex folding in these mutants does not require progenitor cell amplification but is dependent on changes in neuron migration. Analyses and simulations suggest that sulcus formation in the absence of FLRT1/3 results from reduced intercellular adhesion, increased neuron migration, and clustering in the cortical plate. Notably, FLRT1/3 expression is low in the human cortex and in future sulcus areas of ferrets, suggesting that intercellular adhesion is a key regulator of cortical folding across species.

Perfil químico y actividad antibacterial de los extractos de Peltigera laciniata (Merrill ex Riddle) Gyeln / Chemical profile and antibacterial activity of extracts from Peltigera laciniata (Merrill ex Riddle) Gyeln

Introducción: los líquenes, al presentar metabolitos secundarios como xantonas, antraquinonas y alcaloides, se han postulado como material con alto potencial biológico (e. g. antibiótico y antiviral), siendo el antibacterial muy promisorio, el cual se determina por medio de antibiogramas por difusión, punto central de esta investigación. Objetivo: evaluar la actividad antibacterial de los extractos de Peltigera laciniata (Merrill ex Riddle) Gyeln. Olmo de hoja cortada. Métodos: el material liquénico se sometió a percolación con etanol 96 por ciento. Al extracto crudo etanólico se le realizó el aislamiento de alcaloides totales y flavonoides totales con adición de HCL 3 por ciento y metanol, respectivamente. Ambas fracciones, fueron monitoreados por cromatografía de capa fina y fraccionados utilizando cromatografía de columna. Los extractos y fracciones se sometieron a bioensayos sobre Escherichia coli y Staphylococcus aureus para la valoración de los halos de inhibición, utilizando como control Sultamicilina. Los ensayos fueron realizados tres veces con 2 réplicas. Resultados: al realizar la separación cromatográfica de los alcaloides, se observó aumento de la inhibición en comparación con la mezcla alcaloidal. La fracción A1 presenta valores de inhibición cercanos al control y presentó los menores valores de inhibición con respecto a los demás tratamientos evaluados. El efecto de la fracción de los flavonoides totales tuvo menor impacto sobre E. coli y S. aureus, sin embargo, es importante destacar la acción antibacterial de los compuestos nitrogenados de tipo alcaloidal sobre microoganismos Gram positivos. Conclusiones: en el perfil químico realizado a partir de los extractos de la especie de estudio se visualizó la presencia de metabolitos secundarios de tipo alcaloide y flavonoide, evidenciando el efecto antimicrobiano de los alcaloides presentes en el extracto y la fracción, lo cual ratifica el potencial farmacológico de tipo antibacterial, atribuido al núcleo Protoberberínico(AU)

Introduction: Due to their content of secondary metabolites such as xanthones, anthraquinones and alkaloids, lichens have been suggested to be a material of high biological potential (e.g. antibiotic and antiviral). Their very promising antibacterial potential may be determined by diffusion antibiograms, the main concern of the present study. Objective: Evaluate the antibacterial activity of extracts obtained from Peltigera laciniata (Merrill ex Riddle) Gyeln, cutleaf elm. Methods: The lichenic material was percolated with 96 percent ethanol. Total alkaloids and total flavonoids were isolated from the crude ethanolic extract by adding 3 percent HCL and methanol, respectively. Both fractions were monitored by thin-layer chromatography and fractioned by column chromatography. Extracts and fractions were subjected to bioassays against Escherichia coli and Staphylococcus aureus for inhibition haloes, using sultamicillin as control. The assays were conducted 3 times with 2 replications. Results: Upon chromatographic separation of the alkaloids, an increase was observed in inhibition when compared with the alkaloidal mixture. Fraction A1 displayed inhibition values close to the control. Fraction FT showed lower inhibition values than the other treatments evaluated. The fraction of total flavonoids had a lesser impact on E. coli and S. aureus, but alkaloidal nitrogenated compounds had significant antibacterial activity against Gram-positive microorganisms. Conclusions: The chemical profile of extracts from the study species revealed the presence of alkaloidal and flavonoidal secondary metabolites, as well as the antimicrobial effect of the alkaloids contained in the extract and the fraction. This confirms the antibacterial pharmacological potential attributed to the protoberberine core(AU)

Super-complexes of adhesion GPCRs and neural guidance receptors.

Latrophilin adhesion-GPCRs (Lphn1-3 or ADGRL1-3) and Unc5 cell guidance receptors (Unc5A-D) interact with FLRT proteins (FLRT1-3), thereby promoting cell adhesion and repulsion, respectively. How the three proteins interact and function simultaneously is poorly understood. We show that Unc5D interacts with FLRT2 in cis, controlling cell adhesion in response to externally presented Lphn3. The ectodomains of the three proteins bind cooperatively. Crystal structures of the ternary complex formed by the extracellular domains reveal that Lphn3 dimerizes when bound to FLRT2:Unc5, resulting in a stoichiometry of 1:1:2 (FLRT2:Unc5D:Lphn3). This 1:1:2 complex further dimerizes to form a larger 'super-complex' (2:2:4), using a previously undescribed binding motif in the Unc5D TSP1 domain. Molecular dynamics simulations, point-directed mutagenesis and mass spectrometry demonstrate the stability and molecular properties of these complexes. Our data exemplify how receptors increase their functional repertoire by forming different context-dependent higher-order complexes.

Structural basis of latrophilin-FLRT interaction.

Latrophilins, receptors for spider venom α-latrotoxin, are adhesion type G-protein-coupled receptors with emerging functions in synapse development. The N-terminal region binds the endogenous cell adhesion molecule FLRT, a major regulator of cortical and synapse development. We present crystallographic data for the mouse Latrophilin3 lectin and olfactomedin-like (Olf) domains, thereby revealing the Olf ß-propeller fold and conserved calcium-binding site. We locate the FLRT-Latrophilin binding surfaces by a combination of sequence conservation analysis, point mutagenesis, and surface plasmon resonance experiments. In stripe assays, we show that wild-type Latrophilin3 and its high-affinity interactor FLRT2, but not the binding-impaired mutants we generated, promote HeLa cell adhesion. In contrast, cortical neurons expressing endogenous FLRTs are repelled by wild-type Latrophilin3 and not by the binding-impaired mutant. Taken together, we present molecular level insights into Latrophilin structure, its FLRT-binding mechanism, and a role for Latrophilin and FLRT that goes beyond a simply adhesive interaction.

FLRT structure: balancing repulsion and cell adhesion in cortical and vascular development.

FLRTs are broadly expressed proteins with the unique property of acting as homophilic cell adhesion molecules and as heterophilic repulsive ligands of Unc5/Netrin receptors. How these functions direct cell behavior and the molecular mechanisms involved remain largely unclear. Here we use X-ray crystallography to reveal the distinct structural bases for FLRT-mediated cell adhesion and repulsion in neurons. We apply this knowledge to elucidate FLRT functions during cortical development. We show that FLRTs regulate both the radial migration of pyramidal neurons, as well as their tangential spread. Mechanistically, radial migration is controlled by repulsive FLRT2-Unc5D interactions, while spatial organization in the tangential axis involves adhesive FLRT-FLRT interactions. Further, we show that the fundamental mechanisms of FLRT adhesion and repulsion are conserved between neurons and vascular endothelial cells. Our results reveal FLRTs as powerful guidance factors with structurally encoded repulsive and adhesive surfaces.