Biology subjects

Xu, P.

Publications and source records attributed to Xu, P..

At least 145 records · Page 8Linked to original sources

bioRxiv2022

A view of the pan-genome of domesticated cowpea (Vigna unguiculata Walp.)

Cowpea, Vigna unguiculata L. Walp., is a diploid warm-season legume of critical importance as both food and fodder in sub-Saharan Africa. This species is also grown in Northern Africa, Europe, Latin America, North America, and East to Southeast Asia. To capture the genomic diversity of domesticates of this important legume, de novo genome assemblies were produced for representatives of six sub-populations of cultivated cowpea identified previously from genotyping of several hundred diverse accessions. In the most complete assembly (IT97K-499-35), 26,026 core and 4,963 noncore genes were identified, with 35,436 pan genes when considering all seven accessions. GO-terms associated with response to stress and defense response were highly enriched among the noncore genes, while core genes were enriched in terms related to transcription factor activity, and transport and metabolic processes. Over 5 million SNPs relative to each assembly and over 40 structural variants >1 Mb in size were identified by comparing genomes. Vu10 was the chromosome with the highest frequency of SNPs, and Vu04 had the most structural variants. Noncore genes harbor a larger proportion of potentially disruptive variants than core genes, including missense, stop gain, and frameshift mutations; this suggests that noncore genes substantially contribute to diversity within domesticated cowpea. Article SummaryThis study reports annotated genome assemblies of six cowpea accessions. Together with the previously reported annotated genome of IT97K-499-35, these constitute a pan-genome resource representing six subpopulations of domesticated cowpea. Annotations include genes, variant calls for SNPs and short indels, larger presence or absence variants, and inversions. Noncore genes are enriched for loci involved in stress response and harbor many genic variants with potential effects on coding sequence.

genomics↗

bioRxiv2022

Universal mechanism of hormone and allosteric agonist mediated activation of glycoprotein hormone receptors as revealed by structures of follicle stimulating hormone receptor

Follicle stimulating hormone (FSH) is an essential glycoprotein hormone for human reproduction, which functions are mediated by a G protein-coupled receptor, FSHR. Aberrant FSH-FSHR signaling causes infertility and ovarian hyperstimulation syndrome. Here we report cryo-EM structures of FSHR in both inactive and active states, with the active structure bound to FSH and an allosteric agonist compound 21f. The structures of FSHR are similar to other glycoprotein hormone receptors, highlighting a conserved activation mechanism of hormone-induced receptor activation. Compound 21f formed extensive interactions with the TMD to directly activate FSHR. Importantly, the unique residue H6157.42 in FSHR plays an essential role in determining FSHR selectivity for various allosteric agonists. Together, our structures provide a molecular basis of FSH and small allosteric agonist-mediated FSHR activation, which could inspire the design of FSHR-targeted drugs for the treatment of infertility and controlled ovarian stimulation for in vitro fertilization.

cell biology↗

bioRxiv2022

Genetic variation in BnGRP1 contributes to low phosphorus tolerance in Brassica napus

The lack of phosphorus (P) is a major environmental factor affecting rapeseed root growth and development. For breeding purposes, it is crucial to identify the molecular mechanisms of root system architecture (RSA) traits underlying low P tolerance in rapeseed. Using genome-wide association study (GWAS), transcriptome and re-sequencing analyses were done on 400 rapeseed cultivars, the natural variations of glycine-rich protein gene, BnGRP1, in response to low P tolerance. Based on 11 SNP mutations in the BnGRP1 sequence, ten types of haplotypes (Hap) were formed. Compared with the other types, the cultivar of the BnGRP1Hap1 type in the panel demonstrated the longest root length and heaviest root weight. Over-expression of BnGRP1Hap1 in rapeseed depicted the ability to enhance the resistance of rapeseed in its response to low P tolerance. CRISPR/Cas9-derived BnGRP1Hap1 knockout mutations in rapeseed could lead to sensitivity to low P stress. Furthermore, BnGRP1Hap1 influenced the expression of phosphate transporter 1 (PHT1) genes associated with P absorption. Overall, the findings of this study highlight the new mechanisms of GRP1 genes in enhancing the low P tolerance in rapeseed.

molecular biology↗

bioRxiv2022

Oscillatory mechanisms of intrinsic brain networks

Neuroimaging studies of hemodynamic fluctuations have shown specific network-based organization of the brain at rest, yet the neurophysiological underpinning of these networks in human brain remain unclear. Here, we recorded resting-state activities of neuronal populations in the key regions of default mode network (DMN, posterior cingulate cortex and medial prefrontal cortex), frontoparietal network (FPN, dorsolateral prefrontal cortex and inferior parietal lobule), and salience network (SN, anterior insula and dorsal anterior cingulate cortex) from 42 human participants using intracranial electroencephalogram (iEEG). We observed stronger within-network connectivity of the DMN, FPN and SN in broadband iEEG power, stronger phase synchronization within the DMN across theta and alpha bands, and weaker phase synchronization within the FPN in delta, theta and alpha band. We also found positive power correlations in high frequency band (70-170Hz) and negative power correlations in alpha and beta band for FPN-DMN and FPN-SN. Robust negative correlations in DMN-SN were found in alpha, beta and gamma band. These findings provide intracranial electrophysiological evidence in support of the network model for intrinsic organization of human brain and shed light on the way how the brain networks communicate at rest.

neuroscience↗

bioRxiv2022

Neurocomputational mechanisms underlying fear-biased adaptation learning in changing environments

Humans are able to adapt to the fast-changing world by estimating statistical regularities of the environment. Although fear can profoundly impact adaptive behaviors, the neural mechanisms underlying this phenomenon remain elusive. Here, we conducted a behavioral experiment (n = 21) and a functional magnetic resonance imaging experiment (n = 37) with a novel cue-biased adaptation learning task, during which we simultaneously manipulated emotional valence (fearful/neutral expressions of the cue) and environmental volatility (frequent/infrequent reversals of reward probabilities). Across two experiments, computational modelling consistently revealed a higher learning rate for the environment with frequent versus infrequent reversals following neutral cues. In contrast, this flexible adjustment was absent in the environment with fearful cues, suggesting a suppressive role of fear in adaptation to environmental volatility. This suppressive effect was underpinned by activity of the posterior parietal cortex, ventral striatum, hippocampus and dorsal anterior cingulate cortex (dACC) as well as increased functional connectivity between the dACC and temporal-parietal junction (TPJ) for fear with environmental volatility. Dynamic causal modelling identified that the driving effect was located in the TPJ and was associated with dACC activation, suggesting that the suppression of fear on adaptive behaviors occurs at the early stage of bottom-up processing. These findings provide a neuro-computational account of how fear interferes with adaptation to volatility during dynamic environments.

neuroscience↗

bioRxiv2022

Coopting T cell proximal signaling molecules enables Boolean logic-gated CAR T cell control

Introductory paragraphWhile CAR T cells have altered the treatment landscape for B cell malignancies, the risk of on-target, off-tumor toxicity has hampered their development for solid tumors because most target antigens are shared with normal cells1,2. Researchers have attempted to apply Boolean logic gating to CAR T cells to prevent on-target, off-tumor toxicity3-7; however, a truly safe and effective logic-gated CAR has remained elusive8. Here, we describe a novel approach to CAR engineering in which we replace traditional ITAM-containing CD3{zeta} domains with intracellular proximal T cell signaling molecules. We demonstrate that certain proximal signaling CARs, such as a ZAP-70 CAR, can activate T cells and eradicate tumors in vivo while bypassing upstream signaling proteins such as CD3{zeta}. The primary role of ZAP-70 is to phosphorylate LAT and SLP-76, which form a scaffold for the propagation of T cell signaling. We leveraged the cooperative role of LAT and SLP-76 to engineer Logic-gated Intracellular NetworK (LINK) CAR, a rapid and reversible Boolean-logic AND-gated CAR T cell platform that outperforms other systems in both efficacy and the prevention of on-target, off-tumor toxicity. LINK CAR will dramatically expand the number and types of molecules that can be targeted with CAR T cells, enabling the deployment of these powerful therapeutics for solid tumors and diverse diseases such as autoimmunity9 and fibrosis10. In addition, this work demonstrates that the internal signaling machinery of cells can be repurposed into surface receptors, a finding that could have broad implications for new avenues of cellular engineering.

synthetic biology↗

bioRxiv2022

Basis for antibody- and hormone-mediated activation of TSHR in Graves' disease

Thyroid stimulating hormone (TSH), through activation of its G protein-coupled receptor TSHR, controls the synthesis of thyroid hormone (TH), an essential metabolic hormone. Aberrant signaling of TSHR by autoantibodies causes Graves disease and hypothyroidism that affect millions of patients worldwide. Here we report the active structures of TSHR with TSH and an activating autoantibody M22, both bound to an allosteric agonist ML-109, as well as an inactive TSHR structure with inhibitory antibody K1-70. Both TSH and M22 push the extracellular domain (ECD) of TSHR into the upright active conformation. In contrast, K1-70 blocks TSH binding and is incapable of pushing the ECD to the upright conformation. Comparisons of the active and inactive structures of TSHR with those of the luteinizing hormone-choriogonadotropin receptor (LHCGR) reveal a universal activation mechanism of glycoprotein hormone receptors, in which a conserved 10-residue fragment (P10) from the hinge C-terminal loop mediated interactions from the receptor ECD to its transmembrane domain. One surprisingly feature is that there are over 15 cholesterols surrounding TSHR, supporting its preferential location in lipid rafts. These structures also highlight a common mechanism for TSH and autoantibody M22 to activate TSHR, thus providing the molecular basis for Graves disease.

molecular biology↗

bioRxiv2022

Strain dependent differences in coordination of yeast signaling networks

The yeast mitogen activated protein kinase pathways serve as a model system for understanding how network interactions affect the way in which cells coordinate the response to multiple signals. We have quantitatively compared two yeast strain backgrounds YPH499 and {Sigma}1278b (both of which have previously been used to study these pathways) and found several important differences in how they coordinate the interaction between the high osmolarity glycerol (HOG) and mating pathways. In the {Sigma}1278b background, in response to simultaneous stimulus, mating pathway activation is dampened and delayed in a dose dependent manner. In the YPH499 background, only dampening is dose dependent. Further, leakage from the HOG pathway into the mating pathway (crosstalk) occurs during osmostress alone in the {Sigma}1278b background only. The mitogen activated protein kinase Hog1p suppresses crosstalk late in an induction time course in both strains but does not affect the early crosstalk seen in the {Sigma}1278b background. Finally, the kinase Rck2p plays a greater role suppressing late crosstalk in the {Sigma}1278b background than in the YPH499 background. Our results demonstrate that comparisons between laboratory yeast strains provide an important resource for understanding how signaling network interactions are tuned by genetic variation without significant alteration to network structure.

molecular biology↗

bioRxiv2022

An intelligent synthetic bacterium with sound-integrated ability for chronological toxicant detection, degradation, and lethality

Modules, toolboxes, and systems of synthetic biology are being designed to solve environmental problems. However, weak and decentralized functional modules require complicated controls. To address this issue, we investigated an integrated system that can complete detection, degradation, and lethality, in chronological order without exogenous inducers. Biosensors were optimized by regulating expression of receptor and reporter to get higher sensitivity and output signal. Several stationary-phase promoters were selected and compared, while promoter Pfic was chosen to express the degradation enzyme. We created two concepts of lethal circuits by testing various toxic proteins, with a toxin/antitoxin circuit showing a potent lethal effect. Three modules were coupled, step-by-step. Detection, degradation, and lethality were sequentially completed, and the modules had partial attenuation compared to pre-integration, except for degradation. Our study provides a novel concept for integrating and controlling functional modules, which can accelerate the transition of synthetic biology from a concept to practical applications. TeaserWe provide new ideas for integration and chronological control of multiple modules in synthetic biology.

synthetic biology↗

bioRxiv2022

A high confidence Physcomitrium patens plasmodesmata proteome by iterative scoring and validation reveals diversification of cell wall proteins during evolution

Cells of multicellular organisms exchange nutrients, building blocks and information. In animals, this happens via gap junctions, in plants via plasmodesmata (PD). PD have striking properties, translocating a large range of molecules from ions, to metabolites, RNA and proteins up to 40 kDa. PD are hard to characterize due to being deeply embedded into cell walls and the presence of several membranes. While previous studies of protein composition of PD from angiosperms identified large lists of proteins, few were validated. Here, we developed a PD scoring approach in conjunction with systematic localization on a large scale to define a high-confidence PD proteome of Physcomitrium patens. This high confidence PD proteome comprises nearly 300 proteins, which together with the bona fide PD proteins from literature, are made available in the public PDDB database. Conservation of localization across plant species strengthens the reliability of plant PD proteomes and provides a basis for exploring the evolution of this important organelle. In particular, the P. patens PD proteome was highly enriched in cell wall modifying proteins. Callose-degrading glycolyl hydrolase family 17 (GHL17) proteins are presented as an abundant PD protein family with representatives across an evolutionary scale. Exclusively members of the alpha-clade of the GHL17 family are shown to be PD localized and their orthologs occur only in plant species which have developed PD. Members of the EXORDIUM-family and xyloglucan transglycosylases are additional cell-wall located proteins highly abundant in the P. patens PD proteome also showing evolutionary diversification of PD localized family members from other clade members.

plant biology↗

bioRxiv2022

A super robust and efficient DNA storage architecture based on modulation encoding and decoding

Thanks to its high density and long durability, synthetic DNA has been widely considered as a promising solution to the data explosion problem. However, due to the large amount of random base insertion-deletion-substitution (IDSs) errors from sequencing, reliable data recovery remains a critical challenge, which hinders its large-scale application. Here, we propose a modulation-based DNA storage architecture. Experiments on simulation and real datasets demonstrate that it has two distinct advantages. First, modulation encoding provides a simple way to ensure the encoded DNA sequences comply with biological sequence constraints (i.e., GC balanced and no homopolymers); Second, modulation decoding is highly efficient and extremely robust for the detection of insertions and deletions, which can correct up to ~40% errors. These two advantages pave the way for future high-throughput and low-cost techniques, and will kickstart the actualization of a viable, large-scale system for DNA data storage.

synthetic biology↗

bioRxiv2022

Using NAMs to characterize chemical bioactivity at the transcriptomic, proteomic and phosphoproteomic levels

Omic-based technologies are of particular interest and importance for non-animal chemical hazard and risk characterization based on the premise that any apical endpoint change must be underpinned by some alterations measured at the omic levels. In this work we studied cellular responses to caffeine and coumarin by generating and integrating multi-omic data from transcriptomic, proteomic and phosphoproteomic experiments. We have shown that the methodology presented here is able to capture the complete chain of events from the first compound-induced changes at the phosphoproteome level to changes in gene expression induced by transcription factors and lastly to changes in protein abundance that further influence changes at the cellular level. In HepG2 cells we found the metabolism of lipids and general cellular stress to be dominant biological processes in response to caffeine and coumarin exposure, respectively. The phosphoproteomic changes were detected early in time, at very low concentrations and provided a fast adaptive cellular response to chemical exposure. Changes in protein abundance were found much less frequently than the transcriptomic changes and can be used, together with the transcriptomic changes, to facilitate a more complete understanding of pathway responses to chemical exposure. GRAPHIC ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=169 SRC="FIGDIR/small/492410v1_ufig1.gif" ALT="Figure 1"> View larger version (48K): org.highwire.dtl.DTLVardef@1e348c8org.highwire.dtl.DTLVardef@bf50c5org.highwire.dtl.DTLVardef@4fea36org.highwire.dtl.DTLVardef@998cb8_HPS_FORMAT_FIGEXP M_FIG C_FIG

pharmacology and toxicology↗

bioRxiv2022

Genomic insights into the adaptive and convergent evolution of Leuciscus waleckii inhabiting extremely alkaline environments

Leuciscus waleckii is widely distributed in Northeast Asia and has high economic value. Different from its freshwater counterparts, the population in Lake Dali Nur has a strong alkalinity tolerance and can adapt to extremely alkaline-saline water with bicarbonate over 50 mmol/L (pH 9.6), thus providing an exceptional model with which to explore the mechanisms of adaptive evolution under extreme alkaline environments. Here, we assembled a high quilty chromosome-level reference genome for L. waleckii from Lake Dali Nur, which provides an important genomic resource for the exploitation of alkaline water fishery resources and adaptive evolution research across teleost fish. Notably, we identified significantly expanded long terminal repeats (LTRs) and long interspersed nuclear elements (LINEs) in L. waleckii compared to other Cypriniformes fish, suggesting their more recent insertion into the L. waleckii genome. We also identified expansions in genes encoding gamma-glutamyltransferase, which possibly underlie the adaptation to extreme environmental stress. Based on the resequencing of 85 L.waleckii individuals from divergent populations, the historical population size of L.waleckii in Lake Dali Nur dramatically expanded in a thousand years approximately 13,000 years ago, and experienced a cliff recession in the process of adapting to the alkaline environment of Lake Dali Nur approximately 6,000 years ago. Genome scans further revealed the significant selective sweep regions from Lake Dali Nur, which harbour a set of candidate genes involved in hypoxia tolerance, ion transport, acid-base regulation and nitrogen metabolism. In particular, 5 alkali population specific nonsynonymous mutations were identified in CA15 gene copies. In addition, two sites with convergent amino acid mutation were detected in the RHCG-a gene among several alkali environment adapted Cypriniformes fish, this mutation may increase the NH3 excretion rate of the RHCG channel. Our findings provide comprehensive insight into the genomic mechanisms of L. waleckii and reveal their adaptative evolution under extreme alkaline environments.

evolutionary biology↗

bioRxiv2022

Somatosensory Cortical Signature of Facial Nociception and Vibrotactile Touch Induced Analgesia

Pain relief by vibrotactile touch is a common human experience. Previous neurophysiological investigations in animals focused on spinal mechanisms while human studies suggested the involvement of supraspinal mechanisms. Here we asked whether and how the primary somatosensory cortex (S1) is involved in touch induced analgesia. We discovered that in mice, vibrotactile reafferent signals from self-generated whisking significantly reduce facial nociception, which is abolished by specifically blocking touch transmission from thalamus to the barrel cortex (S1B). The presence of whisking altered nociceptive signal processing in S1B neurons. Intrinsic manifold analysis of S1B population activity revealed that whisking pushes the transition of neural state induced by noxious stimuli towards the state encoding non-nocifensive actions. Thus, S1B integrates facial tactile and noxious signals to enable touch mediated analgesia. TeaserVibrotactile signals modulate barrel cortex population dynamics during touch mediated facial analgesia

neuroscience↗

bioRxiv2022

The phosphoproteome is a first responder in tiered cellular adaptation to chemical stress followed by proteomics and transcriptomics alteration

Next-generation risk assessment for environmental chemicals and ingredients in consumer products involves a weight of evidence (WoE) framework integrating a suite of new approach methodologies (NAMs) based on points of departure (PoD) obtained from in vitro assays. Omics techniques provide broad coverages of the molecular toxicity pathway space. Transcriptomics assays especially play a leading role by providing relatively conservative PoDs in comparison with apical endpoints. However, it is unclear whether and how parameters measured using other omics technicquesparticipate in the cellular response to chemical perturbations, especially at exposure levels below the transcriptomically defined PoD. Multi-omics coverage may provide additional sensitive or confirmative biomarkers to complement and reduce the uncertainty in safety decisions made using targeted and transcriptomics assays. In the present study, we compared changes in transcriptomics, proteomics and phosphoproteomics with two prototype compounds, coumarin, as a main study and doxorubicin, as a complementary study to understand the sensitivity of the three omics techniques in response to chemically-induced changes in HepG2 and AC16 cells. With measurements obtained for multiple chemical concentrations and time points, we demonstrated that, compared with proteomics and transcriptomics, phosphoproteomics alterations occur not only earlier in time as expected, but also at much lower chemical concentrations and hence are proximal to the very early effects induced by chemical exposure. The phosphoproteomics changes appear to approach maximum when the transcriptomics alterations begin to be initiated. The results are consistent with a tiered framework of cellular response to chemical insults, where posttranslational modification of preexisting proteins is first seen before transcriptomics induction is engaged to launch a more energy-expensive defense that defines a useful PoD. We conclude that as the cost becomes more affordable, proteomics covering posttranslational modifications can be utilized to provide a more complete coverage of chemical-induced cellular alteration and supplement transcriptomics-based health safety decision making.

pharmacology and toxicology↗

bioRxiv2022

A CRISPR-based instant DNA repositioning system and the early intranuclear life of HSV-1

The intranuclear localization of viral DNA genomes in relation to the intranuclear environment plays critical roles in determining virus fate. Recent advances in the application of chromosome conformation capture-based sequencing analysis (3C technologies) have revealed valuable aspects of the spatiotemporal interplay of viral genomes with host chromosomes. However, to elucidate the causal relationship between the subnuclear localization of viral genomes and the pathogenic outcome of the infection, manipulative tools are needed. Instant repositioning of viral DNAs to specific subnuclear compartments amid infection is a powerful approach to synchronize and interrogate this dynamically changing process in space and time. Herein, we report an inducible CRISPR-based two-component platform that relocates extrachromosomal DNA pieces (5 kb to 170 kb) to the nuclear periphery instantly (CRISPR-nuPin). Based on this system, investigations of herpes simplex virus 1 (HSV-1), a prototype member of the human herpesvirus family, revealed unprecedently reported insights into the early intranuclear life of the pathogen: I) Viral genomes tethered to the nuclear periphery upon entry, compared with those in the nuclear interior, were wrapped around histones with increased suppressive modifications and subjected to stronger transcriptional silencing and prominent inhibition. II) Relocating HSV-1 genomes at 1 hour post infection significantly promoted transcription of viral {beta} and {gamma} genes, termed an "Escaping" effect. III) Early accumulation of ICP0 was a sufficient but not necessary condition mediating "Escaping". IV) Subnuclear localization was only critical during early infection. Importantly, the CRISPR-nuPin tactic should be widely applicable to many DNA viruses. SummaryThe intranuclear localization of viral DNA genomes plays a critical role in determining virus fate. To elucidate the causal relationship between subnuclear localization and the pathogenic outcome of DNA viruses, manipulative tools are needed. Herein, we report an inducible CRISPR-based two-component platform that relocates DNA pieces (5 kb to 170 kb) to the nuclear periphery instantly (CRISPR-nuPin). Utilizing this tactic, we interrogated the early intranuclear life of herpes simplex virus 1 (HSV-1), a prototype of human herpesviruses, in space and time and revealed that I) viral genomes tethered to the nuclear edge upon entry were prone to suppressive histone packaging and severe inhibition. II) Relocating HSV-1 genomes to the nuclear fringe at 1 hour post infection promoted transcription of viral genes ("Escaping"). III) Early accumulation of ICP0 was a sufficient but not necessary condition mediating "Escaping". IV) Subnuclear localization was a critical factor only during early infection. HighlightsO_LICRISPR-nuPin is an inducible two-component DNA repositioning system C_LIO_LIIt mediates instant nuclear edging of viral DNA during infection C_LIO_LIA powerful approach to interrogate DNA viruses in space and time C_LIO_LIViral DNA at the nuclear periphery upon entry is strongly silenced C_LI In briefAn inducible two-component CRISPR-based platform that instantly repositions HSV-1 genomes to the nuclear edge unveils intranuclear space heterogeneity for the incoming viral genomes and dynamic stages of the host-virus interplay during early infection of the pathogen.

microbiology↗

bioRxiv2022

Harnessing cellular perception apparatus for smart metabolic reprogramming

Metabolic reprogramming (MRP) is a fundamental approach in synthetic biology that involves redirecting metabolic flux and remodeling metabolic networks. However, only few approaches have been made in effective metabolic operations, especially at global level of metabolic networks. Naturally existing cellular perception apparatuses (CPAs), such as histidine kinases (HKs), are considered to sit on sensitive nodes of the metabolic network, which can trigger natural MRP upon perceiving environmental fluctuations. We develop a plateform for global MRP by natural environmental stimulation based on the combinational interference of CPAs. The plateform consists of a CRISPRi-mediated dual-gene combinational knockdown (CDCK) strategy and survivorship-based metabolic interaction analysis (SMIA). A total of 35 histidine kinase (HK) genes and 24 glycine metabolism genes were selected as targets to determine effectiveness of our approach for fast-growing chassis Vibrio FA2. Combined knockdown of several genes of HKs and glycine metabolism increased the glycine production. Other other hand, effects of CDCK on bacterial antibiotic resistance were assessed by targeting HKs. Many HKs were identified to be associated with antibiotic resistance in Vibrio FA2, of which combinational knockdown of two HK genes sasA_8 and 04288 reduced the ampicillin resistance. This MRP strategy is powerful and cost-effective, and can be considered as a smart strategy capable of operating a broad range of metabolic networks in microorganisms.

synthetic biology↗

bioRxiv2022

Two mosquito odorant receptors with reciprocal specificity mediated by a single amino acid residue

The southern house mosquito, Culex quinquefasciatus, utilizes two odorant receptors, CquiOR10 and CquiOR2, narrowly tuned to oviposition attractants and well conserved among mosquito species. They detect skatole and indole, respectively, with reciprocal specificity. We swapped the seven transmembrane (TM) domains of CquiOR10 and CquiOR2 and identified TM2 as a specificity determinant. With additional mutations, we showed that CquiOR10A73L behaved like CquiOR2. Conversely, CquiOR2L74A recapitulated CquiOR10 specificity. Next, we generated structural models of CquiOR10 and CquiOR10A73L using RoseTTAFold and AlphaFold and docked skatole and indole using RosettaLigand. These modeling studies suggested space-filling constraints around A73. Consistent with this hypothesis, CquiOR10 mutants with a bulkier residue (Ile, Val) were insensitive to skatole and indole, whereas CquiOR10A73G retained the specificity to skatole and showed a more robust response than the wildtype receptor CquiOR10. On the other hand, Leu to Gly mutation of the indole receptor CquiOR2 reverted the specificity to skatole. Lastly, CquiOR10A73L, CquiOR2, and CquiOR2L74I were insensitive to 3-ethylindole, whereas CquiOR2L74A and CquiOR2L74G gained activity. Additionally, CquiOR10A73G gave more robust responses to 3-ethylindole than CquiOR10. Thus, we suggest the specificity of these receptors is mediated by a single amino acid substitution, leading to finely tuned volumetric space to accommodate specific oviposition attractants.

neuroscience↗