Biology subjects

Song, W.

Publications and source records attributed to Song, W..

At least 163 records · Page 9Linked to original sources

bioRxiv2020

Multiple approaches for massively parallel sequencing of HCoV-19 genomes directly from clinical samples

COVID-19 has caused a major epidemic worldwide, however, much is yet to be known about the epidemiology and evolution of the virus. One reason is that the challenges underneath sequencing HCoV-19 directly from clinical samples have not been completely tackled. Here we illustrate the application of amplicon and hybrid capture (capture)-based sequencing, as well as ultra-high-throughput metatranscriptomic (meta) sequencing in retrieving complete genomes, inter-individual and intra-individual variations of HCoV-19 from clinical samples covering a range of sample types and viral load. We also examine and compare the bias, sensitivity, accuracy, and other characteristics of these approaches in a comprehensive manner. This is, to date, the first work systematically implements amplicon and capture approaches in sequencing HCoV-19, as well as the first comparative study across methods. Our work offers practical solutions for genome sequencing and analyses of HCoV-19 and other emerging viruses.

genomics↗

bioRxiv2020

The Glycosphingolipid GM3 Modulates Conformational Dynamics of the Glucagon Receptor

The extracellular domain (ECD) of Class B1 G-protein coupled receptors (GPCRs) plays a central role in signal transduction and is uniquely positioned to sense both the extracellular and membrane environments. Whilst recent studies suggest a role for membrane lipids in the modulation of Class A and Class F GPCR signalling properties, little is known about the effect of lipids on Class B1 receptors. In this study, we employed multiscale molecular dynamics (MD) simulations to access the dynamics of the glucagon receptor (GCGR) ECD in the presence of native-like membrane bilayers. Simulations showed that the ECD could move about a hinge region formed by residues Q122-E126 to adopt both closed and open conformations relative to the TMD. ECD movements were modulated by binding of the glycosphingolipid GM3. These large-scale fluctuations in ECD conformation that may affect the ligand binding and receptor activation properties. We also identify a unique PIP2 interaction profile near ICL2/TM3 at the G-protein coupling interface, suggesting a mechanism of engaging G-proteins which may have a distinct dependence on PIP2 compared to Class A GPCRs. Given the structural conservation of Class B1 GPCRs, the modulatory effects of GM3 and PIP2 on GCGR may be conserved across these receptors, offering new insights into potential therapeutic targeting. Statement of SignificanceThe role of lipids in regulation of Class B GPCRs remains elusive, despite recent structural advances. In this study, multi-scale molecular dynamics simulations are used to evaluate lipid interactions with the glucagon receptor, a Class B1 GPCR. We find that the glycosphingolipid GM3 binds to the glucagon receptor extracellular domain (ECD), modulating the dynamics of the ECD and promoting movement away from the transmembrane domain. We also identify a unique PIP2 interaction fingerprint in a region known to be important for bridging G-protein coupling in Class A GPCRs. Thus, this study provides molecular insight into the behaviour of the glucagon receptor in a complex lipid bilayer environment which may aid understanding of glucagon receptor signalling properties.

biophysics↗

bioRxiv2020

Sapphire Nanopores for Low-Noise DNA Sensing

Silicon-supported (SiS) solid-state nanopores have broad applications in single-molecule biosensing and diagnostics, but their high capacitive noise has seriously limited both their sensing accuracy and recording speed. Nanopores on insulating glass have demonstrated reduced capacitance and noise, but it remains challenging to bulk-etch amorphous glass to create membranes reproducibly and uniformly. Here a new approach is reported to form triangular sapphire-supported (SaS) nanopore membranes by batch-processing-compatible anisotropic wet etching of sapphire, with membrane dimension demonstrated from ~200 μm to 5 μm. A SaS nanopore in 68 μm-wide silicon nitride membrane has 130 times smaller capacitance (10 pF) compared to a SiS nanopore (~4 μm SiN membrane, ~1.3 nF), despite a 100 times larger membrane. It has a current noise of 18 pA over 100 kHz bandwidth, much smaller than that from our SiS nanopore (46 pA) and comparable with the best reported low-noise nanopores. Further, the SaS nanopore displays a higher signal-to-noise ratio (SNR, 21 versus 11 for SiS nanopore) in DNA sensing, although the SNR can be further improved using thinner membranes and smaller pores. The SaS nanopore presents a simple platform in both fabrication and structure that is particularly suitable for low-noise and high-speed molecular diagnostics.Competing Interest StatementThe authors have declared no competing interest.View Full Text

bioengineering↗

bioRxiv2020

A simple magnetic nanoparticles-based viral RNA extraction method for efficient detection of SARS-CoV-2

1The ongoing outbreak of the novel coronavirus disease 2019 (COVID-19) originating from Wuhan, China, draws worldwide concerns due to its long incubation period and strong infectivity. Although RT-PCR-based molecular diagnosis techniques are being widely applied for clinical diagnosis currently, timely and accurate diagnosis are still limited due to labour intensive and time-consuming operations of these techniques. To address the issue, herein we report the synthesis of poly (amino ester) with carboxyl groups (PC)-coated magnetic nanoparticles (pcMNPs), and the development of pcMNPs-based viral RNA extraction method for the sensitive detection of COVID-19 causing virus, the SARS-CoV-2. This method combines the lysis and binding steps into one step, and the pcMNPs-RNA complexes can be directly introduced into subsequent RT-PCR reactions. The simplified process can purify viral RNA from multiple samples within 20 min using a simple manual method or an automated high-throughput approach. By identifying two different regions (ORFlab and N gene) of viral RNA, a 10-copy sensitivity and a strong linear correlation between 10 and 105 copies of SARS-CoV-2 pseudovirus particles are achieved. Benefitting from the simplicity and excellent performances, this new extraction method can dramatically reduce the turn-around time and operational requirements in current molecular diagnosis of COVID-19, in particular for the early clinical diagnosis.

molecular biology↗

bioRxiv2020

TRIBE editing reveals specific mRNA targets of eIF4E-BP in Drosophila and in mammals

4E-BP (eIF4E-BP) represses translation initiation by binding to the 5cap-binding protein eIF4E and inhibiting its activity. Although 4E-BP has been shown to be important in growth control, stress response, cancer, neuronal activity and mammalian circadian rhythms, it is not understood how it preferentially represses a subset of mRNAs. We successfully used hyperTRIBE (Targets of RNA-binding proteins identified by editing) to identify in vivo 4E-BP mRNA targets in both Drosophila and mammals under conditions known to activate 4E-BP. The protein associates with specific mRNAs, and ribosome profiling data show that mTOR inhibition changes the translational efficiency of 4E-BP TRIBE targets compared to non-targets. In both systems, these targets have specific motifs and are enriched in translation-related pathways, which correlate well with the known activity of 4E-BP and suggest that it modulates the binding specificity of eIF4E and contributes to mTOR translational specificity.

molecular biology↗

bioRxiv2020

Enhancement of Target Specificity of CRISPR-Cas12a by Using a Chimeric DNA-RNA Guide

The CRISPR-Cas9 system is widely used for target-specific genome engineering. Cpf1 is one of the CRISPR effectors that controls target genes by recognizing thymine-rich protospacer adjacent motif (PAM) sequences. Cpf1 has a higher sensitivity to mismatches in the guide RNA than does Cas9; therefore, off-target sequence recognition and cleavage are lower. However, it tolerates mismatches in regions distant from the PAM sequence (TTTN or TTN) in the protospacer, and off-target cleavage issues may become more problematic when Cpf1 activity is improved for therapeutic purposes. In our study, we investigated off-target cleavage by Cpf1 and modified the Cpf1 (cr)RNA to address the off-target cleavage issue. We developed a CRISPR-Cpf1 that can induce mutations in target DNA sequences in a highly specific and effective manner by partially substituting the (cr)RNA with DNA to change the energy potential of base pairing to the target DNA. A model to explain how chimeric (cr)RNA guided CRISPR-Cpf1 and SpCas9 nickase effectively work in the intracellular genome is suggested. In our results, CRISPR-Cpf1 induces less off-target mutations at the cell level, when chimeric DNA-RNA guide was used for genome editing. This study has a potential for therapeutic applications in incurable diseases caused by genetic mutation.

biochemistry↗

bioRxiv2019

Pseudouridine synthases in the RluA family negatively regulate Drosophila nociception.

Nociceptive neurons of Drosophila melanogaster larvae are characterized by highly branched dendritic processes whose proper morphogenesis relies on a large number of RNA-binding proteins. Post-transcriptional regulation of RNA in these dendrites has been found to play an important role in their function. Here, we investigate the neuronal functions of two putative RNA modification genes, RluA-1 and RluA-2, which are predicted to encode pseudouridine synthases. RluA-1 is specifically expressed in larval sensory neurons while RluA-2 expression is ubiquitous. Nociceptor-specific RNAi knockdown of RluA-1 caused hypersensitive nociception phenotypes, which were recapitulated with genetic null alleles. These were rescued with genomic duplication and nociceptor-specific expression of UAS-RluA-1-cDNA. As with RluA-1, RluA-2 loss of function mutants also displayed hyperalgesia. Interestingly, nociceptor neuron dendrites showed a hyperbranched morphology in the RluA-1 mutants. The latter may be a cause or a consequence of heightened sensitivity in mutant nociception behaviors. Author SummaryPseudouridine (Psi) is a C5-glycoside isomer of uridine and it is the most common posttranscriptional modification of RNAs, including noncoding tRNAs, rRNAs, snRNAs as well as mRNAs. Although first discovered in the 1950s, the biological functions of Psi in multicellular organisms are not well understood. Interestingly, a marker for sensory neurons in Drosophila encodes for a putative pseudouridine synthase called RluA-1. Here, we report our characterization of nociception phenotypes for larvae with RluA-1 loss of function along with that of a related gene RluA-2. Disrupting either or both RluA-1 and RluA-2 resulted in hypersensitive nociception. In addition, RluA-1 mutants have more highly branched nociceptor neurites that innervate the epidermis. Our studies suggest an important role for the RluA family in nociception. This may be through its action on RNAs that regulate neuronal excitability and/or dendrite morphogenesis.

genetics↗

bioRxiv2019

Volumetric fluorescence imaging in a human eye model by oblique scanning laser ophthalmoscope (oSLO): a feasibility study

Fluorescence retinal imaging, such as fluorescein angiography, indocyanine green angiography, and autofluorescence imaging, are valuable tools in ophthalmology and vision science. However, these clinical imaging modalities provide en face view of the retina, with limited capability to discriminate retinal layers over a large field-of-view (FOV). We recently developed a novel retinal imaging method, oblique scanning laser ophthalmoscopy (oSLO), to provide volumetric retinal fluorescence imaging without any depth sectioning. OSLO breaks the coaxial alignment of the excitation and detection, to produce a cross-sectional view on retina using the natural ocular optics. In this paper, we demonstrated oSLO in a realistic human eye model and showed the feasibility for future in vivo human retinal imaging. A new optical design was implemented to significantly simplify our previous oSLO systems. We overcame the limitation by the small numerical aperture (NA) of the human eye, by integrating a pair of cylindrical lens in the remote focusing system. We experimentally showed that the current setup can achieve a FOV of [~]3x6x0.8 mm3, and the transverse and axial resolutions of 7 and 41 {micro}m, respectively. The capability of volumetric fluorescence imaging over a large FOV in the human retina could lead to new clinical imaging paradigms for retinal diseases.

bioengineering↗

bioRxiv2019

Protein ubiquitylation is essential for the schizont to merozoite transition in Plasmodium falciparum blood-stage development

Ubiquitylation is a common post translational modification of eukaryotic proteins and in the human malaria parasite, Plasmodium falciparum (Pf) overall ubiquitylation increases in the transition from intracellular schizont to extracellular merozoite stages in the asexual blood stage cycle. Here, we identify specific ubiquitylation sites of protein substrates in three intracellular parasite stages and extracellular merozoites; a total of 1464 sites in 546 proteins were identified (data available via ProteomeXchange with identifier PXD014998). 469 ubiquitylated proteins were identified in merozoites compared with only 160 in the preceding intracellular schizont stage, indicating a large increase in protein ubiquitylation associated with merozoite maturation. Following merozoite invasion of erythrocytes, few ubiquitylated proteins were detected in the first intracellular ring stage but as parasites matured through trophozoite to schizont stages the extent of ubiquitylation increased. We identified commonly used ubiquitylation motifs and groups of ubiquitylated proteins in specific areas of cellular function, for example merozoite pellicle proteins involved in erythrocyte invasion, exported proteins, and histones. To investigate the importance of ubiquitylation we screened ubiquitin pathway inhibitors in a parasite growth assay and identified the ubiquitin activating enzyme (UBA1 or E1) inhibitor MLN7243 (TAK-243) to be particularly effective. This small molecule was shown to be a potent inhibitor of recombinant PfUBA1, and a structural homology model of MLN7243 bound to the parasite enzyme highlights avenues for the development of P. falciparum specific inhibitors. We created a genetically modified parasite with a rapamycin-inducible functional deletion of uba1; addition of either MLN7243 or rapamycin to the recombinant parasite line resulted in the same phenotype, with parasite development blocked at the late schizont stage. These results indicate that the intracellular target of MLN7243 is UBA1, and this activity is essential for the final differentiation of schizonts to merozoites. The ubiquitylation of many merozoite proteins and their disappearance in ring stages are consistent with the idea that ubiquitylation leads to their destruction via the proteasome once their function is complete following invasion, which would allow amino acid recycling in the period prior to the parasites elaboration of a new food vacuole.

microbiology↗

bioRxiv2019

Diagnostic value of galactomannan in bronchoalveolar lavage fluid for chronic respiratory disease with pulmonary aspergillosis

ObjectiveTo explore the diagnostic value of the bronchoalveolar lavage fluid galactomannan (BALF GM) test for chronic respiratory disease with pulmonary aspergillosis, and establish the optimal cut-off value.\n\nMethodsA total of 180 chronic respiratory disease patients seen at the Respiratory Medicine Department of Peking University Shenzhen Hospital from September 2017 to September 2018 were analyzed. According to the diagnostic criteria, we divided the patients into the case group (n = 70, comprising 5, 20, and 45 proven, probable, and possible cases, respectively) and the control group (n = 110). Bronchoalveolar lavage fluid was collected, and the BALF GM test results were analyzed.\n\nResultsA non-parametric rank-sum test showed that the mean rank of the case group was 140.80, which was higher than that of the control group (58.49). The Z-value was 10.335 (P = 0.000), indicating that the general distribution of BALF GM differed between the two groups. A BALF GM cut-off value of 0.485 showed the highest diagnostic efficacy for pulmonary aspergillosis. The sensitivity, specificity, positive predictive value, and negative predictive value were 92.9%, 100%, 92.8%, and 100%, respectively. As the cut-off value increased, the specificity and sensitivity of the BALF GM test increased and decreased, respectively.\n\nConclusionsThe BALF GM test can be used confirm the diagnosis of patients with pulmonary aspergillosis and chronic respiratory disease. The optimum BALF GM cut-off value was 0.485. Antifungal therapy is important for treating pulmonary aspergillus infection in patients with chronic respiratory disease.

microbiology↗

bioRxiv2019

PsyMuKB: A De Novo Variant Knowledge Base Integrating Transcriptional and Translational Information to Identify Isoform-specific Mutations in Developmental Disorders

De novo variants (DNVs) are one of the most significant contributors to severe early-onset genetic disorders such as autism spectrum disorder, intellectual disability, and other developmental and neuropsychiatric (DNP) disorders. Currently, a plethora of DNVs have being identified through the use of next-generation sequencing and much effort has been made to understand their impact at the gene level; however, there has been little exploration of the impact at the isoform level. The brain contains a high level of alternative splicing and regulation, and exhibits a more divergent splicing program than other tissues; therefore, it is crucial to explore variants at the transcriptional regulation level to better interpret the mechanisms underlying DNP disorders. To facilitate better usage and improve the isoform-level interpretation of variants, we developed the PsyMuKB (NeuroPsychiatric Mutation Knowledge Base), a knowledge base containing a comprehensive, carefully curated list of DNVs with transcriptional and translational annotations to enable identification of isoform-specific mutations. PsyMuKB allows a flexible search of genes or variants and provides both table-based descriptions and associated visualizations, such as expression, transcript genomic structures, protein interactions, and the mutation sites mapped on the protein structures. It also provides an easy-to-use web interface, allowing users to rapidly visualize the locations and characteristics of mutations and the expression patterns of the impacted genes and isoforms. PsyMuKB thus constitutes a valuable resource for identifying tissue-specific de novo mutations for further functional studies of related disorders. PsyMuKB is freely accessible at http://psymukb.net.

bioinformatics↗

bioRxiv2019

Engineering an Enzyme for Direct Electrical Monitoring of Activity

Proteins have been shown to be electrically-conductive if tethered to an electrode by means of a specific binding agent, allowing single molecules to be wired into an electrical sensing circuit. This development opens the possibility of exploiting the remarkable chemical versatility of enzymes as sensors, detectors and sequencing devices. We have engineered contact points into a {Phi}29 polymerase by introducing biotinylatable peptide sequences. The modified enzyme was bound to electrodes functionalized with streptavidin. {Phi}29 connected by one biotinylated contact and a second non-specific contact showed rapid small fluctuations in current when activated. Signals were greatly enhanced with two specific contacts. Features in the distributions of DC conductance increased by a factor 2 or more over the open-to closed conformational transition of the polymerase. Polymerase activity is manifested by rapid (millisecond) large (25% of background) current fluctuations imposed on the DC conductance.

bioengineering↗

bioRxiv2019

High-speed volumetric fluorescein angiography (vFA) by oblique scanning laser ophthalmoscopy in mouse retina

Oblique scanning laser ophthalmoscopy (oSLO) is a recently developed technique to provide three-dimensional volumetric fluorescence imaging in retina over a large field of view, without the need for depth sectioning. Here in the paper, we present high-speed volumetric fluorescein angiography (vFA) in mouse retina in vivo by oSLO. By simply using a low-cost industrial CMOS camera, we improved the imaging speed by ~10 times comparing to our previous results, achieving vFA at 2 volumes per second. Enabled by high-speed vFA, we visualized hemodynamics at single capillary level in 3D and provided methods to quantify capillary hematocrit, absolute capillary blood flow speed, and detection of capillary flow stagnancy and stalling. The quantitative metrics for capillary hemodynamics at 3D retinal capillary network can offer valuable insight in vision science and retinal pathologies.

bioengineering↗

bioRxiv2019

N-WASP regulates the mobility of the B cell receptor and co-receptors during signaling activation

Regulation of membrane receptor mobility is important in tuning the cells response to external signals. This is particularly relevant in the context of immune receptor signaling. The binding of B cell receptors (BCR) to antigen induces B cell receptor activation. While actin dynamics and BCR signaling are known to be linked, the role of actin dynamics in modulating receptor mobility is not well understood. Here, we use single molecule imaging to examine BCR movement during signaling activation and examine the role of actin dynamics on BCR mobility. We use a novel machine learning based method to classify BCR trajectories into distinct diffusive states and show that the actin regulatory protein N-WASP regulates receptor mobility. Constitutive loss or acute inhibition of N-WASP, which is associated with enhanced signaling, leads to a predominance of BCR trajectories with lower diffusivity and is correlated with a decrease in actin dynamics. Furthermore, loss of N-WASP reduces diffusivity of CD19, a stimulatory co-receptor of the BCR but not that of unstimulated Fc{gamma}RIIB, an inhibitory co-receptor. The effect of N-WASP is mirrored by inhibition of the Arp2/3 complex and formins. Our results implicate the dynamic actin network in fine-tuning receptor mobility and receptor-ligand interactions, thereby modulating B cell signaling.

biophysics↗

bioRxiv2019

A chromosome-level genome of black rockfish, Sebastes schlegelii, provides insights into the evolution of live birth

Black rockfish (Sebastes schlegelii) is a teleost species where eggs are fertilized internally and retained in the maternal reproductive system, where they undergo development until live birth (termed viviparity). In the present study, we report a chromosome-level black rockfish genome assembly. High-throughput transcriptome analysis (RNA-seq and ATAC-seq), coupled with in situ hybridization (ISH) and immunofluorescence, identify several candidate genes for maternal preparation, sperm storage and release, and hatching. We propose that zona pellucida (ZP) genes retain sperm at the oocyte envelope, while genes in two distinct astacin metalloproteinase subfamilies serve to release sperm from the ZP and free the embryo from chorion at pre-hatching stage. Finally, we present a model of black rockfish reproduction, and propose that the rockfish ovarian wall has a similar function to uterus of mammals. Taken together, these genomic data reveal unprecedented insights into the evolution of an unusual teleost life history strategy, and provide a sound foundation for studying viviparity in non-mammalian vertebrates and an invaluable resource for rockfish ecology and evolution research.

genomics↗

bioRxiv2018

Early detection of retinal alteration by visible and near-infrared optical coherence tomography (vnOCT) in a dexamethasone-induced ocular hypertension mouse model

PurposeTo apply a novel visible and near-infrared optical coherence tomography (vnOCT) in the dexa-methasone-induced ocular hypertension mouse model, and test the capability of four optical markers, peri-papillary retinal nerve fiber layer (RNFL) thickness, total retinal blood flow, VN ratio and hemoglobin oxygen saturation (sO2), in detecting retinal ganglion cell (RGC) damage in association with ocular hyper-tension.\n\nMethodsTwelve mice (C57BL/6J) were separated into a control (n=6) and a dexamethasone group (n=6) receiving twice daily saline or dexamethasone eye drops, respectively, for 7 weeks. Intraocular pressure (IOP) measurements were taken at baseline and weekly. Optical measurements by vnOCT were longitudinally taken at baseline, 4 weeks and 7 weeks. Following week 7, ex vivo RGC counting was performed by immunostaining.\n\nResultsThe dexamethasone group showed a measurable rise in IOP by week 2. Despite the IOP differences between the dexamethasone and control groups, there was not a statistical difference in RNFL thickness or total blood flow over 7 weeks. The dexamethasone group did show an increase in retinal arteriovenous sO2 difference (A-V sO2) that was significant at week 4 and 7. The RNFL VN ratio showed a significant decrease at week 4 and 7 in dexamethasone group associated with a decreased RGC count.\n\nConclusionsRNFL VN ratio and A-V sO2 are capable of detecting early retinal alterations in the dexamethasone-induced ocular hypertension mouse model. Data analysis suggests VN ratio and A-V sO2 are corralated with RGC loss secondary to ocular hypertension, while being independent of IOP.

bioengineering↗

bioRxiv2018

Dynamic plant height QTL revealed in maize through remote sensing phenotyping using a high-throughput unmanned aerial vehicle (UAV)

Plant height is the key factor for plant architecture, biomass and yield in maize (Zea mays). In this study, plant height was investigated using unmanned aerial vehicle high-throughput phenotypic platforms (UAV-HTPPs) for maize diversity inbred lines at four important growth stages. Using an automated pipeline, we extracted accurate plant heights. We found that in temperate regions, from sowing to the jointing period, the growth rate for temperate maize was faster than tropical maize. However, from jointing to flowering stage, tropical maize maintained a vigorous growth state, and finally resulted in a taller plant than temperate lines. Genome-wide association study for temperate, tropical and both groups identified a total of 238 quantitative trait locus (QTLs) for the 16 plant height related traits over four growth periods. And, we found that plant height at different stages were controlled by different genes, for example, PIN1 controlled plant height at the early stage and PIN11 at the flowering stages. In this study, the plant height data collected by the UAV-HTTPs were credible and the genetic mapping power is high, indicating that the application of this UAV-HTTPs into the study of plant height will have great prospects.\n\nHighlightWe used UAV-based sensing platform to investigate plant height over 4 growth stages for different maize populations, and detected numbers of reliable QTLs using GWAS.

genetics↗

bioRxiv2018

State-Dependent Lipid Interactions with the A2a Receptor Revealed by MD Simulations Using In Vivo-Mimetic Membranes

G protein-coupled receptors (GPCRs) are the largest family of integral membrane proteins and a major class of drug targets. Membranes are known to have modulatory effects on GPCRs via specific lipid interactions. However, the mechanisms of such modulations in cell membranes and how they influence GPCR functions remain unclear. Here we report coarse-grained MD simulations on the Adenosine A2a receptor embedded in an in vivo mimetic membrane model comprised of 10 different lipid species. Three conformational states of the receptor, i.e. the inactive state, the active state, and the active state with a mini-GS protein bound were simulated to study the impact of protein-lipid interactions on the receptor activation. The simulations revealed three specific lipids (GM3, cholesterol and PIP2) that form stable and preferential interactions with the receptor, differentiating these from bulk lipids such as PS, PE and PC. In total, nine specific lipid-binding sites were revealed. The strength of lipid interaction with these sites depends on the conformational state of the receptor, suggesting that these lipids may regulate the conformational dynamics of the receptor. In particular, we revealed a dual role of PIP2 in promoting A2aR activation, which involves stabilization of both the characteristic outward tilt of helix TM6 within receptor and also the association of A2aR and mini-Gs when the activated complex forms. Structural comparisons suggested that PIP2 may facilitate G activation. Our results reveal likely allosteric effects of bound lipids in regulating the functional behaviour of GPCRs, providing a springboard for design of allosteric modulators of these biomedically important receptors.

biophysics↗