Two gene clusters and their positive regulator SlMYB13 that have undergone domestication-associated negative selection control phenolamide accumulation and drought tolerance in tomato

Among plant metabolites,phenolamides,which are conjugates of hydroxycinnamic acid derivatives and polyamines,play important roles in plant adaptation to abiotic and biotic stresses.However,the molecular mechanisms underlying phenolamide metabolism and regulation as well as the effects of domestication and breeding on phenolamide diversity in tomato remain largely unclear.In this study,we performed a metabolite-based genome-wide association study and identified two biosynthetic gene clusters(BGC7 and BGC11)containing 12 genes involved in phenolamide metabolism,including four biosynthesis genes(two 4CL genes,one C3H gene,and one CPA gene),seven decoration genes(five AT genes and two UGT genes),and one transport protein gene(DTX29).Using gene co-expression network analysis we further discovered that SlMYB13 positively regulates the expression of two gene clusters,thereby promoting phenolamide accumulation.Genetic and physiological analyses showed that BGC7,BGC11 and SlMYB13 enhance drought tolerance by enhancing scavenging of reactive oxygen species and increasing abscisic acid content in tomato.Natural variation analysis suggested that BGC7,BGC11 and SlMYB13 were negatively selected during tomato domestication and improvement,leading to reduced phenolamide content and drought tolerance of cultivated tomato.Collectively,our study discovers a key mechanism of phenolamide biosynthesis and regulation in tomato and reveals that crop domestication and improvement shapes metabolic diversity to affect plant environmental adaptation.

Conserved H3K27me3-associated chromatin looping mediates physical interactions of gene clusters in plants

Higher-order chromatin organization is essential for transcriptional regulation,genome stability maintenance,and other genome functions.Increasing evidence has revealed significant differences in 3D chromatin organization between plants and animals.However,the extent,pattern,and rules of chromatin organization in plants are still unclear.In this study,we systematically identified and characterized long-range chromatin loops in the Arabidopsis 3D genome.We identified hundreds of long-range cis chromatin loops and found their anchor regions are closely associated with H3K27me3 epigenetic modifications.Furthermore,we demonstrated that these chromatin loops are dependent on Polycomb group(PcG)proteins,suggesting that the Polycomb repressive complex2(PRC2)complex is essential for establishing and maintaining these novel loops.Although most of these PcG-medicated chromatin loops are stable,many of these loops are tissue-specific or dynamically regulated by different treatments.Interestingly,tandemly arrayed gene clusters and metabolic gene clusters are enriched in anchor regions.Long-range H3K27me3-marked chromatin interactions are associated with the coregulation of specific gene clusters.Finally,we also identified H3K27me3-associated chromatin loops associated with gene clusters in Oryza sativa and Glycine max,indicating that these long-range chromatin loops are conserved in plants.Our results provide novel insights into genome evolution and transcriptional coregulation in plants.

Heat-inducible SlWRKY3 confers thermotolerance by activating the SlGRXS1 gene cluster in tomato

High temperature stress is one of the major environmental factors that affect the growth and development of plants. Although WRKY transcription factors play a critical role in stress responses, there are few studies on the regulation of heat stress by WRKY transcription factors,especially in tomato. Here, we identified a group I WRKY transcription factor, SlWRKY3, involved in thermotolerance in tomato. First, SlWRKY3 was induced and upregulated under heat stress. Accordingly, overexpression of SlWRKY3 led to an increase, whereas knock-out of SlWRKY3 resulted in decreased tolerance to heat stress. Overexpression of SlWRKY3 accumulated less reactive oxygen species(ROS), whereas knock-out of SlWRKY3 accumulated more ROS under heat stress. This indicated that SlWRKY3 positively regulates heat stress in tomato. In addition,SlWRKY3 activated the expression of a range of abiotic stress-responsive genes involved in ROS scavenging, such as a SlGRXS1 gene cluster.Further analysis showed that SlWRKY3 can bind to the promoters of the SlGRXS1 gene cluster and activate their expression. Collectively, these results imply that SlWRKY3 is a positive regulator of thermotolerance through direct binding to the promoters of the SlGRXS1 gene cluster and activating their expression and ROS scavenging.

GST family genes in jujube actively respond to phytoplasma infection

Jujube witches’broom(JWB)caused by phytoplasma has a severely negative effect on multiple metabolisms in jujube.The GST gene family in plants participates in the regulation of a variety of biotic and abiotic stresses.This study aims to identify and reveal the changes in the jujube GST gene family in response to phytoplasma infection.Here,70 ZjGSTs were identified in the jujube genome and divided into 8 classes.Among them,the Tau-class,including 44 genes,was the largest.Phylogenetic analysis indicated that Tau-class genes were highly conserved among species,such as Arabidopsis,cotton,chickpea,and rice.Through chromosome location analysis,37.1%of genes were clustered,and 8 of 9 gene clusters were composed of Tau class members.Through RT-PCR,qRT-PCR and enzyme activity detection,the results showed that the expression of half(20/40)of the tested ZjGSTs was inhibited by phytoplasma infection in field and tissue culture conditions,and GST activity was also significantly reduced.In the resistant and susceptible varieties under phytoplasma infection,ZjGSTU49-ZjGSTU54 in the cluster IV showed opposite expression patterns,which may be due to functional divergence during evolution.Some upregulated genes(ZjGSTU45,ZjGSTU49,ZjGSTU59,and ZjGSTU70)might be involved in the process of jujube against JWB.The yeast two-hybrid results showed that all 6 Tauclass proteins tested could form homodimers or heterodimers.Overall,the comprehensive analysis of the jujube GST gene family revealed that ZjGSTs responded actively to phytoplasma infection.Furthermore,some screened genes(ZjGSTU24,ZjGSTU49-52,ZjGSTU70,and ZjDHAR10)will contribute to further functional studies of jujube-phytoplasma interactions.

FunGeneClusterS: Predicting fungal gene clusters from genome and transcriptome data

Introduction:Secondary metabolites of fungi are receiving an increasing amount of interest due to their prolific bioactivities and the fact that fungal biosynthesis of secondary metabolites often occurs from co-regulated and co-located gene clusters.This makes the gene clusters attractive for synthetic biology and industrial biotechnology applications.We have previously published a method for accurate prediction of clusters from genome and transcriptome data,which could also suggest cross-chemistry,however,this method was limited both in the number of parameters which could be adjusted as well as in userfriendliness.Furthermore,sensitivity to the transcriptome data required manual curation of the predictions.In the present work,we have aimed at improving these features.Results:FunGeneClusterS is an improved implementation of our previous method with a graphical user interface for off-and on-line use.The new method adds options to adjust the size of the gene cluster(s)being sought as well as an option for the algorithm to be flexible with genes in the cluster which may not seem to be co-regulated with the remainder of the cluster.We have benchmarked the method using data from the well-studied Aspergillus nidulans and found that the method is an improvement over the previous one.In particular,it makes it possible to predict clusters with more than 10 genes more accurately,and allows identification of co-regulated gene clusters irrespective of the function of the genes.It also greatly reduces the need for manual curation of the prediction results.We furthermore applied the method to transcriptome data from A.niger.Using the identified best set of parameters,we were able to identify clusters for 31 out of 76 previously predicted secondary metabolite synthases/synthetases.Furthermore,we identified additional putative secondary metabolite gene clusters.In total,we predicted 432 co-transcribed gene clusters in A.niger(spanning 1.323 genes,12%of the genome).Some of these had functions related to primary metabolism,e.g.we have identified a cluster for biosynthesis of biotin,as well as several for degradation of aromatic compounds.The data identifies that suggests that larger parts of the fungal genome than previously anticipated operates as gene clusters.This includes both primary and secondary metabolism as well as other cellular maintenance functions.Conclusion:We have developed FunGeneClusterS in a graphical implementation and made the method capable of adjustments to different datasets and target clusters.The method is versatile in that it can predict co-regulated clusters not limited to secondary metabolism.Our analysis of data has shown not only the validity of the method,but also strongly suggests that large parts of fungal primary metabolism and cellular functions are both co-regulated and co-located.

Efficient inversions and duplications of mammalian regulatory DNA elements and gene clusters by CRISPR/Cas9

The human genome contains millions of DNA regulatory elements and a large number of gene clusters,most of which have not been tested experimentally.The clustered regularly interspaced short palindromic repeats(CRISPR)/CRISPR-associated nuclease 9(Cas9)programed with a synthetic single-guide RNA(sgRNA)emerges as a method for genome editing in virtually any organisms.Here we report that targeted DNA fragment inversions and duplications could easily be achieved in human and mouse genomes by CRISPR with two sgRNAs.Specifically,we found that,in cultured human cells and mice,efficient precise inversions of DNA fragments ranging in size froma few tens of bp to hundreds of kb could be generated.In addition,DNA fragment duplications and deletions could also be generated by CRISPR through trans-allelic recombination between the Cas9-induced double-strand breaks(DSBs)on two homologous chromosomes(chromatids).Moreover,junctions of combinatorial inversions and duplications of the protocadherin(Pcdh)gene clusters induced by Cas9 with four sgRNAs could be detected.In mice,we obtained founders with alleles of precise inversions,duplications,and deletions of DNA fragments of variable sizes by CRISPR.Interestingly,we found that very efficient inversions were mediated by microhomology-mediated end joining(MMEJ)through short inverted repeats.We showed for the first time that DNA fragment inversions could be transmitted through germlines in mice.Finally,we applied this CRISPR method to a regulatory element of the Pcdha cluster and found a new role in the regulation of members of the Pcdhg cluster.This simple and efficient method should be useful in manipulating mammalian genomes to study millions of regulatory DNA elements as well as vast numbers of gene clusters.

Assembly and features of secondary metabolite biosynthetic gene clusters in Streptomyces ansochromogenes

A draft genome sequence of Streptomyces ansochromogenes 7100 was generated using 454 sequencing technology. In combination with local BLAST searches and gap filling techniques, a comprehensive antiSMASH-based method was adopted to assemble the secondary metabolite biosynthetic gene clusters in the draft genome of S. ansochromogenes. A total of at least 35 putative gene clusters were identified and assembled. Transcriptional analysis showed that 20 of the 35 gene clusters were expressed in either or all of the three different media tested, whereas the other 15 gene clusters were silent in all three different media. This study provides a comprehensive method to identify and assemble secondary metabolite biosynthetic gene clusters in draft genomes of Streptomyces, and will significantly promote functional studies of these secondary metabolite biosynthetic gene clusters.

Optimization of Charging/Battery-Swap Station Location of Electric Vehicles with an Improved Genetic Algorithm-Based Model

The joint location planning of charging/battery-swap facilities for electric vehicles is a complex problem.Considering the differences between these two modes of power replenishment,we constructed a joint location-planning model to minimize construction and operation costs,user costs,and user satisfaction-related penalty costs.We designed an improved genetic algorithm that changes the crossover rate using the fitness value,memorizes,and transfers excellent genes.In addition,the present model addresses the problem of“premature convergence”in conventional genetic algorithms.A simulated example revealed that our proposed model could provide a basis for optimized location planning of charging/battery-swapping facilities at different levels under different charging modes with an improved computing efficiency.The example also proved that meeting more demand for power supply of electric vehicles does not necessarily mean increasing the sites of charging/battery-swap stations.Instead,optimizing the level and location planning of charging/battery-swap stations can maximize the investment profit.The proposed model can provide a reference for the government and enterprises to better plan the location of charging/battery-swap facilities.Hence,it is of both theoretical and practical value.

Recent Progress in Elucidating the Structure, Function and Evolution of Disease Resistance Genes in Plants

Plants employ multifaceted mechanisms to fight with numerous pathogens in nature. Resistance （R） genes are the most effective weapons against pathogen invasion since they can specifically recognize the corresponding pathogen effectors or associated protein（s） to activate plant immune responses at the site of infection. Up to date, over 70 R genes have been isolated from various plant species. Most R proteins contain conserved motifs such as nucleotide-binding site （NBS）, leucine-rich repeat （LRR）, Toll-interleukin-1 receptor domain （TIR, homologous to cytoplasmic domains of the Drosophila Toll protein and the manamalian intefleukin-1 receptor）, coiled-coil （CC） or leucine zipper （LZ） structure and protein kinase domain （PK）. Recent results indicate that these domains play significant roles in R protein interactions with effector proteins from pathogens and in activating signal transduction pathways involved in innate immunity. This review highlights an overview of the recent progress in elucidating the structure, function and evolution of the isolated R genes in different plant-pathogen interaction systems.

Rational Engineering of Secondary Metabolic Pathways in a Heterologous Host to Enable the Biosynthesis of Hibarimicin Derivatives with Enhanced Anti-Melanomic Activity

A 61-kb biosynthetic gene cluster(BGC),which is accountable for the biosynthesis of hibarimicin(HBM)B from Microbispora rosea subsp.hibaria TP-A0121,was heterologously expressed in Streptomyces coelicolor M1154,which generated a trace of the target products but accumulated a large amount of shunt products.Based on rational analysis of the relevant secondary metabolism,directed engineering of the biosynthetic pathways resulted in the high production of HBM B,as well as new HBM derivates with improved antitumor activity.These results not only establish a biosynthetic system to effectively synthesize HBMs-a class of the largest and most complex Type-Ⅱpolyketides,with a unique pseudo-dimeric structure-but also set the stage for further engineering and deep investigation of this complex biosynthetic pathway toward potent anticancer drugs.

Bruton’s tyrosine kinase inhibitors in primary central nervous system lymphoma:New hopes on the horizon

In this editorial,we comment on the article by Wang et al.This manuscript explores the potential synergistic effects of combining zanubrutinib,a novel oral inhibitor of Bruton’s tyrosine kinase,with high-dose methotrexate(HD-MTX)as a therapeutic intervention for primary central nervous system lymphoma(PCNSL).The study involves a retrospective analysis of 19 PCNSL patients,highlighting clinicopathological characteristics,treatment outcomes,and genomic biomarkers.The results indicate the combination’s good tolerance and strong antitumor activity,with an 84.2%overall response rate.The authors emphasize the potential of zanubrutinib to modulate key genomic features of PCNSL,particularly mutations in myeloid differentiation primary response 88 and cluster of differentiation 79B.Furthermore,the study investigates the role of circulating tumor DNA in cerebrospinal fluid for disease surveillance and treatment response monitoring.In essence,the study provides valuable insights into the potential of combining zanubrutinib with HD-MTX as a frontline therapeutic regimen for PCNSL.The findings underscore the importance of exploring alternative treatment modalities and monitoring genomic and liquid biopsy markers to optimize patient outcomes.While the findings suggest promise,the study’s limitations should be considered,and further research is needed to establish the clinical relevance of this therapeutic approach for PCNSL.

A novel and complete gene cluster involved in the degradation of aniline by Delftia sp. AN3

A recombinant strain, Escherichia coli JM109-AN1, was obtained by constructing of a genomic library of the total DNA of Delftia sp. AN3 in E. coli JM109 and screening for catechol 2,3-dioxygenase activity. This recombinant strain could grow on aniline as sole carbon, nitrogen and energy source. Enzymatic assays revealed that the exogenous genes including aniline dioxygenase （AD） and catechol 2,3-dioxygenase （C230） genes could well express in the recombinant strain with the activities of AD and C230 up to 0.31 U/mg wet cell and 1.92 U/mg crude proteins, respectively. The AD or C23O of strain AN3 could only catalyze aniline or catechol but not any other substituted substrates. This recombinant strain contained a recombinant plasmid, pKC505-AN1, in which a 29.7-kb DNA fragment from Delftia sp. AN3 was inserted. Sequencing and open reading frame （orfs） analysis of this 29.7 kb fragment revealed that it contained at least 27 orfs, among them a gene cluster （consisting of at least 16 genes, named danQTA1A2BRDCEFG1HIJKG2） was responsible for the complete metabolism of aniline to TCA-cycle intermediates. This gene cluster could be divided into two main parts, the upper sequences consisted of 7 genes （danQTA1A2BRD） were predicted to encode a multi-component aniline dioxygenase and a LysR-type regulator, and the central genes （danCEFG1HIJKG2） were expected to encode meta-cleavage pathway enzymes for catechol degradation to TCA-cycle intermediates. Unlike clusters tad from Delftia tsuruhatensis AD9 and tdn from Pseudomonas putida UCC22, in this gene cluster, all the genes were in the same transcriptional direction. There was only one set of C230 gene （danC） and ferredoxin-like protein gene （danD）. The presence of only one set of these two genes and specificity of AD and C230 might be the reason for strain AN3 could only degrade aniline. The products of danQTA1A2BRDC showed 99%-100% identity to those from Delftia acidovorans 7N, and 50%-85% identity to those of tad cluster from D. tsuruhatensis AD9 in amino acid residues. Besides this dan cluster, the 29.7 kb fragment also contained genes encoding the trans-membrane transporter and transposases which might be needed for transposition of the gene cluster. Pulsed-field gel electrophoresis （PFGE） and plasmid curing experiments suggested that the dan cluster might be encoded on the chromosome of strain AN3. The GenBank accession number for the dan cluster of Delftia sp. AN3 is DQ661649.

Form Gene Clustering Method about Pan-Ethnic-Group Products Based on Emotional Semantic

The use of pan-ethnic-group products form knowledge primarily depends on a designer＇s subjective experience without user participation. The majority of studies primarily focus on the detection of the perceptual demands of consumers from the target product category. A pan-ethnic-group products form gene clustering method based on emotional semantic is constructed. Consumers＇ perceptual images of the pan-ethnic-group products are obtained by means of product form gene extraction and coding and computer aided product form clustering technology. A case of form gene clustering about the typical pan-ethnic-group products is investigated which indicates that the method is feasible. This paper opens up a new direction for the future development of product form design which improves the agility of product design process in the era of Industry 4.0.

A pair of two－component regulatory genes ecrA1／A2 in S. coelicolor

Two-component genes are kinds of genetic elements involved in regulation of antibiotic production in Streptomyces coelicolor. DNA microarray analysis revealed that ecrA1/A2, which mapped at distant sites from red locus and encode respectively the kinase and regulator, expressed coordinately with genes of Red specific biosynthetic pathway, ecrA1 and ecrA2 gene-disruptive mutants were constructed using homogenotisation by reciprocal double crossover. Fermentation data showed that the undecylprodigiosin (Red) level of production was lower than that of wild-type strain. However, the change of the actinorhodin (Act) production level was not significant compared with wild type. Thus, these experiment results confirmed that the two-component system ecrA 1/A2 was positive regulatory element for red gene cluster.

Clustering of Major Genes Conferring Blast Resistance in a Durable Resistance Rice Cultivar Gumei 2

By using 304 recombinant inbred lines derived from indica rice cross Zhong 156/Gumei 2, a linkage map consisting of 177 marker loci and covering 12 rice chromosomes was constructed and employed for mapping genes conferring blast resistance in rice. Genomic location of gene Pi25(t) conferring neck blast resistance to the Chinese isolate 92-183 (race ZC15) was verified to be located between markers A7 and RG456 on chromosome 6, with genetic distances of 1.7 cM and 1.5 cM to A7 and RG456, respectively. Leaf blast resistance of Gumei 2 to the Philippine isolate Ca89 (lineage 4) was found to be controlled by a single gene. The gene tentatively designated as Pi26(\) was located between makers B10 and R674 on chromosome 6, with genetic distances of 5.7 cM and 25.8 cM to B10 and R674 respectively. Resistant alleles at both gene loci were derived from Gumei 2, indicating an existence of resistance gene cluster in Gumei 2.

Evolutionary dynamics of the Wnt gene family:implications for lophotrochozoans

Genes encoding Wnt ligands, which have important roles in cell communication and organ development, are restricted to multicellular animals. We systematically studied W nt genes from eumetazoan genomes, with emphasis on the poorly studied superphylum Lophotrochozoa(four annelids, seven mollusks, eight platyhelminths, one bdelloid rotifer, and one brachiopod species). Between 3 and 39 W nt loci were identified in each genome, and the protostome-specific loss of Wnt3 genes was confirmed. We identified gastropod-specific loss of Wnt8, refining the previously proposed mollusk-specific loss. Some duplicated Wnt genes belonging to a same subfamily or closely related subfamilies showed tandem distribution in the lophotrochozoan genomes, indicating tandem duplication events during Wnt family evolution. Members of the conserved Wnt10-Wnt6-Wnt1-Wnt9 cluster showed highly correlated expression patterns over time in two assayed lophotrochozoans, the oyster C rassostrea gigas and the brachiopod L ingula anatina, reflecting the possible similar function of the clustered W nt genes.

Floating Escherichia coli by Expressing Cyanobacterial Gas Vesicle Genes

Gas vesicles are hollow, air-filled polyprotein structures that provide the buoyancy to cells. They are found in a variety of prokaryotes. In this study, we isolated a partial gas vesicle protein gene cluster containing gvpA and gvpC20ψ from Planktothrix rubescens, and inserted it into an expression vector and expressed it in E. coli. The gas vesicle was developed in bacterial cells, which made bacterial cells to float on medium surface. We also amplified gvpA and gvpC20ψ separately and synthesized an artificial operon by fusing these two genes with the standardized gene expression controlling elements of E. coli. The artificial operon was expressed in E. coli, forming gas vesicles and floating bacteria cells. Our findings verified that the whole set of genes and the overall structure of gas vesicle gene cluster are not necessary for developing gas vesicles in bacteria cells. Two genes, gvpA and gvpC20ψ, of the gas vesicle gene cluster are sufficient for synthesizing an artificial operon that can develop gas vesicles in bacteria cells. Our findings provided a wide range of applications including easing the harvest of cultured microalgae and bacteria, as well as enriching and remediating aquatic pollutants by constructing gas vesicles in their ceils.

Identification of anrF gene, a homology of admM of andrimid biosynthetic gene cluster related to the antagonistic activity of Enterobacter cloacae B8

AIM： To identify the gene （s） related to the antagonistic activity of Enterobacter cloacae B8 and to elucidate its antagonistic mechanism. METHODS： Transposon-mediated mutagenesis and tagging method and cassette PCR-based chromosomal walking method were adopted to isolate the mutant strain （s） of B8 that lost the antagonistic activity and to clone DNA fragments around Tn5 insertion site. Sequence compiling and open reading frame （ORF） finding were done with DNAStar program and homologous sequence and conserved domain searches were performed with BlastN or BlastP programs at www.ncbi.nlm.nih.gov. To verify the gene involved in the antagonistic activity, complementation of a full-length clone of the anrFgene to the mutant B8F strain was used. RESULTS： A 3 321 bp contig around the Tn5 insertion site was obtained and an ORF of 2 634 bp in length designated as anrFgene encoding for a 877 aa polyketide synthase-like protein was identified. It had a homology of 83% at the nucleotide level and 79% ID/87% SIM at the protein level, to the admM gene of Pantoea agglomerans andrimid biosynthetic gene cluster （AY192157）. The Tn5 was inserted at 2 420 bp of the gene corresponding to the COG3319 （the thioesterase domain of type I polyketide synthase） coding region on BSF. The antagonistic activity against Xanthomonas oryzae pv. oryzae was resumed with complementation of the full-length anrFgene to the mutant B8F. CONCLUSION： The anrFgene obtained is related to the antagonistic activity of BS, and the antagonistic substances produced by B8 are andrimid and/or its analogs.

Cytogenetic Mapping of Disease Resistance Genes and Analysis of Their Distribution Features on Chromosomes in Maize

Cytogenetic maps of four clusters of disease resistance genes were generated by ISH of the two RFLP markers tightly linked to and flanking each of maize resistance genes and the cloned resistance genes from other plant species onto maize chromosomes, combining with data published before. These genes include Helminthosporium turcium Pass resistance genes Ht1, Htn1 and Ht2, Helminthosporium maydis Nisik resistance genes Rhm1 and Rhm2, maize dwarf mosaic virus resistance gene Mdm1, wheat streak mosaic virus resistance gene Wsm1, Helminthosporium carbonum ULLstrup resistance gene Hml and the cloned Xanthomonas oryzae pv. Oryzae resistance gene Xa21 of rice, Cladosporium fulvum resistance genes Cf-9 and Cf-2.1 of tomato,and Pseudomonas syringae resistance gene RPS2 of Arabidopsis. Most of the tested disease resistance genes located on the four chromosomes, i.e., chromosomes1, 3, 6 and 8, and they closely distributed at the interstitial regions of these chromosomal long arms with percentage distances ranging 31.44(±3.72)-72.40(±3.25) except for genes Rhm1, Rhm2, Mdm1 and Wsm1 which mapped on the satellites of the short arms of chromosome6. It showed that the tested RFLP markers and genes were duplicated or triplicated in maize genome. Homology and conservation of disease resistance genes among species, and relationship between distribution features and functions of the genes were discussed. The results provide important scientific basis for deeply understanding structure and function of disease resistance genes and breeding in maize.

Involvement of clustered oyster Wnt genes in gut formation

Genes encoding Wnt ligands play important roles in organ development. The Wnt10-Wnt6-Wnt1-Wnt9 cluster widely presents in many metazoan genomes, indicating the importance of gene arrangement. Hypothesis has been proposed that they may be coordinately regulated. However, few expression correlations were identified in model animals. We analyzed the tissue expression pattern of clustered oyster Wnt10, Wnt6, Wnt1, and Wnt9 a genes in this study. The results indicated the highest expression level in adult gut system of these clustered W nt genes, except for Wnt6, which had highest expression in mantle. Further whole-mount immunofluorescence assay indicated that Wnt6 protein was restricted to gut region in oyster larvae. These results suggest the possible important role of the W nt10-Wnt6-Wnt1-Wnt9 cluster in oyster gut formation.