Molecular characterization of QTL for grain zinc and iron concentrations in wheat landrace Chinese Spring.

KEY MESSAGE: Three stable QTL for grain zinc concentration were identified in wheat landrace Chinese Spring. Favorable alleles were more frequent in landraces than in modern wheat cultivars. Wheat is a major source of dietary energy for the growing world population. Developing cultivars with enriched zinc and iron can potentially alleviate human micronutrient deficiency. In this study, a recombinant inbred line (RIL) population with 245 lines derived from cross Zhou 8425B/Chinese Spring was used to detect quantitative trait loci (QTL) for grain zinc concentration (GZnC) and grain iron concentration (GFeC) across four environments. Three stable QTL for GZnC with all favorable alleles from Chinese Spring were identified on chromosomes 3BL, 5AL, and 5BL. These QTL explaining maxima of 8.7%, 5.8%, and 7.1% of phenotypic variances were validated in 125 resequenced wheat accessions encompassing both landraces and modern cultivars using six kompetitive allele specific PCR (KASP) assays. The frequencies of favorable alleles for QGZnCzc.caas-3BL, QGZnCzc.caas-5AL and QGZnCzc.caas-5BL were higher in landraces (90.4%, 68.0%, and 100.0%, respectively) compared to modern cultivars (45.9%, 35.4%, and 40.9%), suggesting they were not selected in breeding programs. Candidate gene association studies on GZnC in the cultivar panel further delimited the QTL into 8.5 Mb, 4.1 Mb, and 47.8 Mb regions containing 46, 4, and 199 candidate genes, respectively. The 5BL QTL located in a region where recombination was suppressed. Two stable and three less stable QTL for GFeC with favorable alleles also from Chinese Spring were identified on chromosomes 4BS (Rht-B1a), 4DS (Rht-D1a), 1DS, 3AS, and 6DS. This study sheds light on the genetic basis of GZnC and GFeC in Chinese Spring and provides useful molecular markers for wheat biofortification.

Enhancing Methane Gas Sensing through Defect Engineering in Ag-Ru Co-doped ZnO Nanorods.

Detection of leaks of flammable methane (CH4) gas in a timely manner can mitigate health, safety, and environmental risks. Zinc oxide (ZnO), a polar semiconductor with controllable surface defects, is a promising material for gas sensing. In this study, Ag-Ru co-doped into self-assembled ZnO nanorod arrays (ZnO NRs) was prepared by a one-step hydrothermal method. The Ag-Ru co-doped sample shows a good hydrophobic property as a result of its particular microstructure, which results in high humidity resistance. In addition, oxygen vacancy density significantly increased after Ag-Ru co-doping. Density functional theory (DFT) calculations revealed an exceptionally high charge density accumulated at the Ru sites and the formation of a localized strong electric field, which provides additional energy for the CH4 reaction with •O2- at the surface at room temperature. Optimized AgRu0.025-ZnO demonstrated an outstanding CH4 sensing performance, with a limit of detection (LOD) as low as 2.24 ppm under free-heat and free-light conditions. These findings suggest that introducing defects into the ZnO lattice, such as oxygen vacancies and localized ions, offers a promising approach to improving the gas sensing performance.

Rht12b, a widely used ancient allele of TaGA2oxA13, reduces plant height and enhances yield potential in wheat.

KEY MESSAGE: We identified a new wheat dwarfing allele Rht12b conferring reduced height and higher grain yield, pinpointed its causal variations, developed a breeding-applicable marker, and traced its origin and worldwide distribution. Plant height control is essential to optimize lodging resistance and yield gain in crops. RHT12 is a reduced height (Rht) locus that is identified in a mutationally induced dwarfing mutant and encodes a gibberellin 2-oxidase TaGA2oxA13. However, the artificial dwarfing allele is not used in wheat breeding due to excessive height reduction. Here, we confirmed a stable Rht locus, overlapping with RHT12, in a panel of wheat cultivars and its dwarfing allele reduced plant height by 5.4-8.2 cm, equivalent to Rht12b, a new allele of RHT12. We validated the effect of Rht12b on plant height in a bi-parent mapping population. Importantly, wheat cultivars carrying Rht12b had higher grain yield than those with the contrasting Rht12a allele. Rht12b conferred higher expression level of TaGA2oxA13. Transient activation assays defined SNP-390(C/A) in the promoter of TaGA2oxA13 as the causal variation. An efficient kompetitive allele-specific PCR marker was developed to diagnose Rht12b. Conjoint analysis showed that Rht12b plus the widely used Rht-D1b, Rht8 and Rht24b was the predominant Rht combination and conferred a moderate plant height in tested wheat cultivars. Evolutionary tracking uncovered that RHT12 locus arose from a tandem duplication event with Rht12b firstly appearing in wild emmer. The frequency of Rht12b was approximately 70% (700/1005) in a worldwide wheat panel and comparable to or higher than those of other widely used Rht genes, suggesting it had been subjected to positive selection. These findings not only identify a valuable Rht gene for wheat improvement but also develop a functionally diagnostic tool for marker-assisted breeding.

Molecular identification and validation of four stable QTL for adult-plant resistance to powdery mildew in Chinese wheat cultivar Bainong 64.

KEY MESSAGE: Four stable QTL for adult-plant resistance (APR) to powdery mildew were identified on chromosome arms 1DL, 2BS, 2DL, and 6BL in the widely grown Chinese wheat cultivar Bainong 64. These QTL had no effect on response to stripe rust or leaf rust. Wheat powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt), is a devastating fungal disease. Seedlings of Chinese wheat Bainong 64 are susceptible to Bgt, but adult plants have maintained resistance since it was released in 1996. A population of 171 recombinant inbred lines (RILs) developed from cross Jingshuang 16/Bainong 64 (JS16/BN64) was used to dissect genetic components of powdery mildew resistance. A genetic map comprising 5383 polymorphic markers was constructed using the 15 K SNP chip and kompetitive allele-specific PCR (KASP) markers. Composite interval mapping identified four stable QTL with favorable alleles all from BN64 on chromosome arms 1DL, 2BS, 2DL, and 6BL in at least four environments. They accounted for 8.3%, 13.8%, 14.4%, and 9.0% of the total phenotypic variation explained (PVE) in maximum, respectively. QPmjbr.caas-1DL, situated about 22 Mb from centromere, is probably a new QTL. QPmjbr.caas-2DL located near the end of arm 2DL and explained the largest PVE. Using genetic maps populated with KASP markers, QPmjbr.caas-2BS and QPmjbr.caas-6BL were fine mapped to a 1.8 cM genetic intervals spanning 13.6 Mb (76.0-89.6 Mb) and 1.7 cM and 4.9 Mb (659.9-664.8 Mb), respectively. The four QTL independent of stripe rust and leaf rust resistance were validated for powdery mildew resistance in another RIL population related to BN64 and a cultivar panel using representative KASP markers. Since BN64 has been a leading cultivar and an important breeding parent in China, the QTL and markers reported in this study will be useful for marker-assisted selection of APR.

Molecular characterization of stable QTL and putative candidate genes for grain zinc and iron concentrations in two related wheat populations.

KEY MESSAGE: Major QTL for grain zinc and iron concentrations were identified on the long arm of chromosomes 2D and 6D. Gene-based KASP markers were developed for putative candidate genes TaIPK1-2D and TaNAS10-6D. Micronutrient malnutrition is one of the most common public health problems in the world. Biofortification, the most attractive and sustainable solution to surmount malnutrition requires the development of micronutrient enriched new crop cultivars. In this study, two recombinant inbred line (RIL) populations, ZM175/XY60 and ZM175/LX987, were used to identify QTL for grain zinc concentration (GZnC), grain iron concentration (GFeC) and thousand grain weight (TGW). Eight QTL for GZnC, six QTL for GFeC and five QTL for TGW were detected. Three QTL on chromosomes 2DL and 4BS and chromosome 6A showed pleiotropic effects on all three traits. The 4BS and 6A QTL also increased plant height and might be Rht-B1a and Rht25a, respectively. The 2DL locus within a suppressed recombination region was identified in both RIL populations and the favorable allele simultaneously increasing GZnC, GFeC and TGW was contributed by XY60 and LX987. A QTL on chromosome 6DL associated only with GZnC was detected in ZM175/XY60 and was validated in JD8/AK58 RILs using kompetitive allele-specific PCR (KASP) marker K_AX-110119937. Both the 2DL and 6DL QTL were new loci for GZnC. Based on gene annotations, sequence variations and expression profiles, the phytic acid biosynthesis gene TaIPK1-2D and nicotianamine synthase gene TaNAS10-6D were predicted as candidate genes. Their gene-based KASP markers were developed and validated in a cultivar panel of 343 wheat accessions. This study investigated the genetic basis of GZnC and GFeC and provided valuable candidate genes and markers for breeding Zn- and Fe-enriched wheat.

Genetic dissection of grain morphology and yield components in a wheat line with defective grain filling.

KEY MESSAGE: We identified stable QTL for grain morphology and yield component traits in a wheat defective grain filling line and validated genetic effects in a panel of cultivars using breeding-relevant markers. Grain filling capacity is essential for grain yield and appearance quality in cereal crops. Identification of genetic loci for grain filling is important for wheat improvement. However, there are few genetic studies on grain filling in wheat. Here, a defective grain filling (DGF) line wdgf1 characterized by shrunken grains was identified in a population derived from multi-round crosses involving nine parents and a recombinant inbreed line (RIL) population was generated from the cross between wdgf1 and a sister line with normal grains. We constructed a genetic map of the RIL population using the wheat 15K single nucleotide polymorphism chip and detected 25 stable quantitative trait loci (QTL) for grain morphology and yield components, including three for DGF, eleven for grain size, six for thousand grain weight, three for grain number per spike and two for spike number per m2. Among them, QDGF.caas-7A is co-located with QTGW.caas-7A and can explain 39.4-64.6% of the phenotypic variances, indicating that this QTL is a major locus controlling DGF. Sequencing and linkage mapping showed that TaSus2-2B and Rht-B1 were candidate genes for QTGW.caas-2B and the QTL cluster (QTGW.caas-4B, QGNS.caas-4B, and QSN.caas-4B), respectively. We developed kompetitive allele-specific PCR markers tightly linked to the stable QTL without corresponding to known yield-related genes, and validated their genetic effects in a diverse panel of wheat cultivars. These findings not only lay a solid foundation for genetic dissection underlying grain filling and yield formation, but also provide useful tools for marker-assisted breeding.

Fine mapping of reduced height locus RHT26 in common wheat.

KEY MESSAGE: We fine mapped RHT26 for plant height in wheat, confirmed its genetic effects in a panel of wheat cultivars and predicted candidate genes. Development of wheat cultivars with appropriate plant height (PH) is an important goal in breeding. Utilization of semi-dwarfing genes Rht-B1b and Rht-D1b triggered wheat Green Resolution in the 1960s. Since these genes also bring unfavorable features, such as reduced coleoptile length and grain weight, it is necessary to identify alternative reduced height genes without yield penalty. Here we constructed a high-density genetic map of a recombinant inbred line population derived from the cross of Zhongmai175 and Lunxuan987 and detected a stable genetic locus for PH, designated RHT26, on chromosome arm 3DL in all of six environments, accounting for 6.8-14.0% of the phenotypic variances. RHT26 was delimited to an approximate 1.4 Mb physical interval (517.1-518.5 Mb) using secondary mapping populations derived from 22 heterozygous recombinant plants and 24 kompetitive allele-specific PCR markers. Eleven high-confidence genes were annotated in the physical interval according to the Chinese Spring reference genome, and four of them were predicted as candidates for RHT26 based on genome and transcriptome sequencing analyses. We also confirmed that RHT26 had significant effects on PH, but not grain yield in a panel of wheat cultivars; its dwarfing allele has been frequently used in wheat breeding. These findings lay a sound foundation for map-based cloning of RHT26 and provide a breeding-applicable tool for marker-assisted selection.

High Resolution Genome Wide Association Studies Reveal Rich Genetic Architectures of Grain Zinc and Iron in Common Wheat (Triticum aestivum L.).

Biofortification is a sustainable strategy to alleviate micronutrient deficiency in humans. It is necessary to improve grain zinc (GZnC) and iron concentrations (GFeC) in wheat based on genetic knowledge. However, the precise dissection of the genetic architecture underlying GZnC and GFeC remains challenging. In this study, high-resolution genome-wide association studies were conducted for GZnC and GFeC by three different models using 166 wheat cultivars and 373,106 polymorphic markers from the wheat 660K and 90K single nucleotide polymorphism (SNP) arrays. Totally, 25 and 16 stable loci were detected for GZnC and GFeC, respectively. Among them, 17 loci for GZnC and 8 for GFeC are likely to be new quantitative trait locus/loci (QTL). Based on gene annotations and expression profiles, 28 promising candidate genes were identified for Zn/Fe uptake (8), transport (11), storage (3), and regulations (6). Of them, 11 genes were putative wheat orthologs of known Arabidopsis and rice genes related to Zn/Fe homeostasis. A brief model, such as genes related to Zn/Fe homeostasis from root uptake, xylem transport to the final seed storage was proposed in wheat. Kompetitive allele-specific PCR (KASP) markers were successfully developed for two major QTL of GZnC on chromosome arms 3AL and 7AL, respectively, which were independent of thousand kernel weight and plant height. The 3AL QTL was further validated in a bi-parental population under multi-environments. A wheat multidrug and toxic compound extrusion (MATE) transporter TraesCS3A01G499300, the ortholog of rice gene OsPEZ2, was identified as a potential candidate gene. This study has advanced our knowledge of the genetic basis underlying GZnC and GFeC in wheat and provides valuable markers and candidate genes for wheat biofortification.

The low expression of NUP62CL indicates good prognosis and high level of immune infiltration in lung adenocarcinoma.

A primary factor in tumor morbidity and mortality, lung adenocarcinoma (LUAD) is known to be a major subtype of lung cancer, having the lowest survival rate among all other cancers. Using The Cancer Genome Atlas (TCGA) database the relationship between the immune infiltrate and the NUP62CL was explored and the value of the NUP62CL expression in the prognosis and diagnosis LUAD was examined. Using the logistic regression and the Wilcoxon signed-rank test the relationship between the NUP62CL and the clinico-pathological features was analyzed. There was a significant correlation between the clinical stage (p = 0.005), the N (p = 0.004), and the decreased expression of NUP62CL. The prognosis of LUAD with high NUP62CL expression was revealed to be worse than that with low NUP62CL expression (p < 0.001) by the Kaplan-Meier survival analysis. The potentiality of NUP62CL to be a significant factor of prognosis for LUAD was indicated by the analyses of the multivariate and the univariate Cox regression models. In LUAD, the crucial role of recombination and maintenance of telomere as a significant pathway for NUP62CL was suggested by the Gene Set Enrichment Analysis (GSEA). To analyze the correlation between the genes and the tumor infiltrating immune cells the CIBERSORT was used. Moreover the positive correlation with the NUP62CL expression in LUAD of the infiltration level of the tumor infiltrating B lymphocytes and memory CD4+ T cells was exhibited by CIBERSORT. Therefore, NUP62CL may be a new valuable prognostic indicator for LUAD.

Dissection of Molecular Processes and Genetic Architecture Underlying Iron and Zinc Homeostasis for Biofortification: From Model Plants to Common Wheat.

The micronutrients iron (Fe) and zinc (Zn) are not only essential for plant survival and proliferation but are crucial for human health. Increasing Fe and Zn levels in edible parts of plants, known as biofortification, is seen a sustainable approach to alleviate micronutrient deficiency in humans. Wheat, as one of the leading staple foods worldwide, is recognized as a prioritized choice for Fe and Zn biofortification. However, to date, limited molecular and physiological mechanisms have been elucidated for Fe and Zn homeostasis in wheat. The expanding molecular understanding of Fe and Zn homeostasis in model plants is providing invaluable resources to biofortify wheat. Recent advancements in NGS (next generation sequencing) technologies coupled with improved wheat genome assembly and high-throughput genotyping platforms have initiated a revolution in resources and approaches for wheat genetic investigations and breeding. Here, we summarize molecular processes and genes involved in Fe and Zn homeostasis in the model plants Arabidopsis and rice, identify their orthologs in the wheat genome, and relate them to known wheat Fe/Zn QTL (quantitative trait locus/loci) based on physical positions. The current study provides the first inventory of the genes regulating grain Fe and Zn homeostasis in wheat, which will benefit gene discovery and breeding, and thereby accelerate the release of Fe- and Zn-enriched wheats.

Development and Validation of a Clinical Prognostic Model Based on Immune-Related Genes Expressed in Clear Cell Renal Cell Carcinoma.

Background: Clear cell renal cell carcinoma (ccRCC) is the most frequent and terminal subtype of RCC. Reliable markers associated with the immune response are not available to predict the prognosis of patients with ccRCC. We exploited the extensive number of ccRCC samples from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) repository to perform a comprehensive analysis of immune-related genes (IRGs). Methods: Based on TCGA data, we incorporated IRGs and their expression profiles of 72 normal and 539 ccRCC samples. Univariate Cox analysis was used to evaluate the relationship between overall survival (OS) and IRGs expression. The Lasso Cox regression model identified prognostic genes used to establish a clinical immune prognostic model. The TF-IRG network was used to study the potential molecular mechanisms of action and properties of ccRCC-specific IRGs. Multivariate Cox analysis established a clinical prognostic model of IRGs. Results: We found a significant correlation among 15 differentially expressed IRGs with the OS of patients with ccRCC. Gene function enrichment analysis showed that these IRGs are significantly associated with response to receptor ligand activity. Lasso Cox regression analysis identified 10 genes with the greatest prognostic value. A clinical prognostic model based on six IRGs, which performed well for predicting prognosis, revealed significant associations of patients' survival with age, sex, stage, tumor, node, and metastasis. Moreover, these findings reflect the infiltration of tumors by various immune cells. Conclusion: We identified six clinically significant IRGs and incorporated them into a clinical prognostic model with great significance for monitoring and predicting prognosis of ccRCC.

The ability to manipulate ROS metabolism in pepper may affect aphid virulence.

Myzus persicae has severe economic impact on pepper (Capsicum) cultivation. Previously, we identified two populations of M. persicae, NL and SW, that were avirulent and virulent, respectively on C. baccatum accession PB2013071. The transcriptomics approach used in the current study, which is the first study to explore the pepper-aphid interaction at the whole genome gene expression level, revealed genes whose expression is differentially regulated in pepper accession PB2013071 upon infestation with these M. persicae populations. The NL population induced ROS production genes, while the SW population induced ROS scavenging genes and repressed ROS production genes. We also found that the SW population can induce the removal of ROS which accumulated in response to preinfestion with the NL population, and that preinfestation with the SW population significantly improved the performance of the NL population. This paper supports the hypothesis that M. persicae can overcome the resistance in accession PB2013071 probably because of its ability to manipulate plant defense response especially the ROS metabolism and such ability may benefit avirulent conspecific aphids.

FIBCD1 overexpression predicts poor prognosis in patients with hepatocellular carcinoma.

Fibrinogen C domain-containing 1 (FIBCD1) is an acetyl-recognition receptor that affects the occurrence and development of certain tumors. However, the prognostic significance of FIBCD1 in hepatocellular carcinoma (HCC) remains unclear. This study aimed to explore FIBCD1 expression in HCC and to determine the prognostic value of FIBCD1 in patients with HCC. A total of 1,058 liver tissue samples with detailed and complete clinical information were collected, including 495 HCC samples. Tissue microarray immunohistochemistry analysis was used to evaluate FIBCD1 protein expression in the collected tissues. The Kaplan-Meier plotter online tool was used to investigate the association between FIBCD1 expression and prognosis of patients with HCC. Oncomine and the Gene Expression Profiling Interactive Analysis database were used for bioinformatics analysis of FIBCD1. Results showed that FIBCD1 expression was higher in HCC and was associated with tumor diameter (P=0.002), tumor number (P=0.001), tumor node metastasis stage (P<0.001), primary tumor (T; P<0.001), lymph node metastases (N; P=0.002), distant metastases (M; P=0.023), differentiation degree (P=0.003), vascular invasion (P<0.001) and liver cirrhosis (P=0.011). Patients with HCC and high FIBCD1 expression had worse overall survival than those with low FIBCD1 expression. High FIBCD1 expression (P<0.001), TNM stage (P=0.003), T (P<0.001), N (P=0.014), and vascular invasion (P<0.001) were independent prognostic factors in HCC. Hence, FIBCD1 may be a novel biomarker for prognosis evaluation of HCC.

Aphid populations showing differential levels of virulence on Capsicum accessions.

The green peach aphid, Myzus persicae, is one of the most threatening pests in pepper cultivation and growers would benefit from resistant varieties. Previously, we identified two Capsicum accessions as susceptible and three as resistant to M. persicae using an aphid population originating from the Netherlands (NL). Later on we identified an aphid population originating from a different geographical region (Switserland, SW) that was virulent on all tested Capsicum accessions. The objective of the current work is to describe in detail different aspects of the interaction between two aphid populations and two selected Capsicum accessions (one that was susceptible [PB2013046] and one that was resistant [PB2013071] to population NL), including biochemical processes involved. Electrical penetration graph (EPG) recordings showed similar feeding activities for both aphid populations on PB2013046. On accession PB2013071 the aphid population SW was able to devote significantly more time to phloem ingestion than population NL. We also studied plant defense response and found that plants of accession PB2013046 could not induce an accumulation of reactive oxygen species and callose formation after infestation with either aphid population. However, plants of PB2013071 induced a stronger defense response after infestation by population NL than after infestation by population SW. Based on these results, population SW of M. persicae seems to have overcome the resistance of PB2013071 that prevented feeding of aphids from NL population. The potential mechanism by which SW population overcomes the resistance is discussed.

Aphid resistance in Capsicum maps to a locus containing LRR-RLK gene analogues.

KEY MESSAGE: A QTL for aphid resistance on pepper chromosome 2 was identified and validated. This QTL affects aphid survival and reproduction, and was fine mapped to a locus containing LRR-RLK analogues. Myzus persicae is one of the most threatening insect pests that adversely affects pepper (Capsicum) cultivation. Resistance to aphids was previously identified in Capsicum baccatum. This study aimed at elucidating the genetics of aphid resistance in C. baccatum. A QTL analysis was carried out for M. persicae resistance in an F2 population derived from an intraspecific cross between a highly resistant plant and a susceptible plant. Survival and reproduction were used as resistance parameters. Interval mapping detected two QTLs affecting aphid survival (Rmpas-1) and reproduction (Rmprp-1), respectively, both localized in the same area and sharing the same top marker on chromosome 2. Use of this marker as co-factor in multiple-QTL mapping analysis revealed a second, minor QTL (Rmprp-2) only affecting aphid reproduction, on chromosome 4. Fine mapping confirmed the effects of Rmpas-1 and Rmprp-1 and narrowed the major QTL Rmprp-1 down to a genomic region of 96 kb which is predicted to encode four analogues of resistance genes of the receptor-like kinase family containing a leucine-rich repeat domain (LRR-RLKs). This work provides not only initial information for breeding aphid-resistant pepper varieties, but also forms the basis for future molecular analysis of gene(s) involved in aphid resistance.

Reduced phloem uptake of Myzus persicae on an aphid resistant pepper accession.

BACKGROUND: The green peach aphid (GPA), Myzus persicae, is economically one of the most threatening pests in pepper cultivation, which not only causes direct damage but also transmits many viruses. Breeding aphid resistant pepper varieties is a promising and environmentally friendly method to control aphid populations in the field and in the greenhouse. Until now, no strong sources of resistance against the GPA have been identified. Therefore the main aims of this study were to identify pepper materials with a good level of resistance to GPA and to elucidate possible resistance mechanisms. RESULTS: We screened 74 pepper accessions from different geographical areas for resistance to M. persicae. After four rounds of evaluation we identified one Capsicum baccatum accession (PB2013071) as highly resistant to M. persicae, while the accessions PB2013062 and PB2012022 showed intermediate resistance. The resistance of PB2013071 resulted in a severely reduced uptake of phloem compared to the susceptible accession, as determined by Electrical Penetration Graph (EPG) studies. Feeding of M. persicae induced the expression of callose synthase genes and resulted in callose deposition in the sieve elements in resistant, but not in susceptible plants. CONCLUSIONS: Three aphid resistant pepper accessions were identified, which will be important for breeding aphid resistant pepper varieties in the future. The most resistant accession PB2013071 showed phloem-based resistance against aphid infestation.

Identification and characterization of pheromone receptors and interplay between receptors and pheromone binding proteins in the diamondback moth, Plutella xyllostella.

Moths depend on olfactory cues such as sex pheromones to find and recognize mating partners. Pheromone receptors (PRs) and Pheromone binding proteins (PBPs) are thought to be associated with olfactory signal transduction of pheromonal compounds in peripheral olfactory reception. Here six candidate pheromone receptor genes in the diamondback moth, Plutella xyllostella were identified and cloned. All of the six candidate PR genes display male-biased expression, which is a typical characteristic of pheromone receptors. In the Xenopus-based functional study and in situ hybridization, PxylOR4 is defined as another pheromone receptor in addition to the previously characterized PxylOR1. In the study of interaction between PRs and PBPs, PxylPBPs could increase the sensitivity of the complex expressing oocyte cells to the ligand pheromone component while decreasing the sensitivity to pheromone analogs. We deduce that activating pheromone receptors in olfactory receptor neurons requires some role of PBPs to pheromone/PBP complex. If the chemical signal is not the pheromone component, but instead, a pheromone analog with a similar structure, the complex would have a decreased ability to activate downstream pheromone receptors.

Expression patterns and binding properties of three pheromone binding proteins in the diamondback moth, Plutella xyllotella.

Pheromone binding proteins (PBPs) play a key role in transporting hydrophobic sex pheromone components emitted by con-specific female across aqueous sensillar lymph to the surface of olfactory receptor neurons. A number of PBPs have been cloned, however, details of their function are still largely unknown. Here three pheromone binding protein genes in the diamondback moth, Plutella xyllotella were cloned. The three PxylPBP genes are not only expressed in chemosensory tissues but also expressed in female reproductive organs and male legs. To better understand the functions of PxylPBPs in the initial steps of pheromone recognition, three PxylPBPs were expressed in Escherichia coli and the ligand-binding specificities of purified recombinant PBPs were investigated. Fluorescence binding assays indicate that three PxylPBPs not only robustly bound all four sex pheromone components but also significantly bound pheromone analogs with at least one double bond, while weakly bound tested plant volatiles. Although pheromone analogs bound PBPs, they could not elicit the moth's electrophysiological response. These experiments provide evidence that PxylPBPs have limited selectivity of pheromone components and analogs and some downstream components such as odor receptors might be involved in selectivity and specificity of pheromone perception in P. xyllotella.