Oligopeptide Transporter Genes Research Articles

A Remarkable Expansion of Oligopeptide Transporter Genes in Rust Fungi (Pucciniales) Suggests a Specialization in Nutrient Acquisition for Obligate Biotrophy.

Nutrient acquisition by rust fungi during their biotrophic growth has been assigned to a few transporters expressed in haustorial infection structures. We performed a comparative genomic analysis of all transporter genes (hereafter termed transportome) classified according to the Transporter Classification Database, focusing specifically on rust fungi (order Pucciniales) versus other species in the Dikarya. We also surveyed expression of transporter genes in the poplar rust fungus for which transcriptomics data are available across the whole life cycle. Despite a significant increase in gene number, rust fungi presented a reduced transportome compared with most fungi in the Dikarya. However, a few transporter families in the subclass Porters showed significant expansions. Notably, three metal transport-related families involved in the import, export, and sequestration of metals were expanded in Pucciniales and expressed at various stages of the rust life cycle, suggesting a tight regulation of metal homeostasis. The most remarkable gene expansion in the Pucciniales was observed for the oligopeptide transporter (OPT) family, with 25 genes on average compared with seven to 14 genes in the other surveyed taxonomical ranks. A phylogenetic analysis showed several specific expansion events at the root of the order Pucciniales with subsequent expansions in rust taxonomical families. The OPT genes showed dynamic expression patterns along the rust life cycle and more particularly during infection of the poplar host tree, suggesting a possible specialization for the acquisition of nitrogen and sulfur through the transport of oligopeptides from the host during biotrophic growth.

Candida tropicalis oligopeptide transporters assist in the transmembrane transport of the antimicrobial peptide CGA-N9

Candida tropicalis is often reported as the second or third most common pathogen causing fungal infections. Antimicrobial peptides (AMPs) have attracted increasing attention for their broad-spectrum antimicrobial properties and low cytotoxicity. Our previous studies have shown that CGA-N9, a non-membrane-rupturing AMP, crosses the cell membrane to exert anticandidal activity. We speculate that there are some related transporters that assist in the transmembrane transport of CGA-N9. In this study, the relationship between CGA-N9 lethality kinetics and its real-time transmembrane amount in C. tropicalis cells was investigated. The results demonstrated that there was a positive correlation between its candicidal activity and transmembrane amount. A total of 12 oligopeptide transporter (OPT) coding sequences (CDSs) were cloned from C. tropicalis by using the conservative OPT gene sequences of Candida spp. to design primers and were named C. tropicalis OPTs (CtOPTs). The results of RT‒qPCR demonstrated that the expression levels of CtOPT1, CtOPT9 and CtOPT12 were correlated with the CGA-N9 transmembrane amount in a time-dependent manner. The results of molecular docking demonstrated that CtOPT1, CtOPT9 and CtOPT12 interact strongly with CGA-N9. Therefore, CtOPT1, CtOPT9 and CtOPT12 were predicted to assist in the transmembrane transport of the AMP CGA-N9.

The genome sequence of the Champagne-Epernay-Geisenheim wine yeast reveals its hybrid nature.

Lager yeasts are hybrids between Saccharomyces cerevisiae and S. eubayanus. Wine yeast biodiversity, however, has only recently been discovered to include besides pure S. cerevisiae strains also hybrids between different Saccharomyces yeasts as well as introgressions from non-Saccharomyces species. Here we analysed the genome of the Champagne-Epernay-Geisenheim (CEG) wine yeast. This yeast is an allotetraploid (4n-1) hybrid of S. cerevisiae harbouring a substantially reduced S. kudriavzevii genome contributing only 1/3 of a full genome equivalent. We identified a novel oligopeptide transporter gene, FOT4, in CEG located on chromosome XVI. FOT genes were originally derived from Torulaspora microellipsoides and FOT4 arose by non-allelic recombination between adjacent FOT1 and FOT2 genes. Fermentations of CEG in Riesling and Müller-Thurgau musts were compared with the S. cerevisiae Geisenheim wine yeast GHM, which does not carry FOT genes. At low temperature (10°C), CEG completed fermentations faster and produced increased levels of higher alcohols (e.g. isoamyl alcohol). At higher temperature (18°C), CEG produced higher amounts of the pineapple-like alkyl esters i-butyric and propionic acid ethyl esters compared to GHM. The hybrid nature of CEG thus provides advantages in grape must fermentations over S. cerevisiae wine yeasts, especially with regard to aroma production.

Genome-Wide Identification and Transcript Analysis Reveal Potential Roles of Oligopeptide Transporter Genes in Iron Deficiency Induced Cadmium Accumulation in Peanut.

The oligopeptide transporter (OPT) family is a group of proton-coupled symporters that play diverse roles, including metal homeostasis. However, little is known about this family of peanuts. To reveal the potential roles of AhOPT genes in Fe/Cd interactions, peanut AhOPT genes were genome-widely identified, and the relationships between gene expression and Cd accumulation were detected in two contrasting peanut cultivars (Fenghua 1 and Silihong) under Fe-sufficient or Fe-deficient conditions. A total of 40 AhOPT genes were identified in peanuts, which were divided into two subfamilies (PT and YS). Most AhOPT genes underwent gene duplication events predominated by whole-genome duplication. Clustered members generally have similar protein structures. However, gene structural divergences occurred in most of the duplicated genes. Transcription analysis revealed that AhOPT3.2/3.4 and AhYSL3.1/3.2 might be responsible for Fe deficiency tolerance, while AhOPT3.1/3.4, AhOPT7.1/7.2, and AhYSL1.1 be involved in Fe/Cd interactions. These genes might be regulated by transcription factors, including ATHB-12, ATHB-6, DIVARICATA, MYB30, NAC02, DOF3.4, IDD7, and LUX. Reduced expressions of AhYSL3.1/3.2 and higher expressions of AhOPT3.4 might contribute to higher Fe-deficiency tolerance in Silihong. Higher expression of AhOPT7.3 and AhOPT6.1 might be responsible for low Cd accumulation in Fenghua 1. Our results confirmed that AhOPT3/6/7 and AhYSL1/3 might be involved in the transport of Fe and/or Cd in peanuts and provided new clues to understanding potential mechanisms of Fe/Cd interactions.

Function and transcriptome analysis of an oligopeptide transporter CgOPT2 in the rubber anthracnose fungus Colletotrichum gloeosporioides

Colletotrichum gloeosporioides is a main pathogen causing rubber anthracnose, which brings huge economic loss to natural rubber industry. Oligopeptide transporter (OPT) is a kind of peptide transport protein, which usually exhibits various biological functions in fungi. In this study, an oligopeptide transporter gene CgOPT2 is identified from the rubber anthracnose fungus C. gloeosporioides as a homolog of OPT2 in Saccharomyces cerevisiae. CgOPT2 encodes a 990-amino acid protein, which has an OPT domain harboring 15 transmembrane domains. The gene knock-out mutant of CgOPT2 had reduced vegetative growth on PDA and CM media, and was more sensitive to H2O2, SDS and Congo Red than the wild-type strain. Moreover, the mutant had obviously lagged conidia germination and attenuated virulence on rubber leaves. Transcriptome analysis showed CgOPT2 could affect the expression of many genes involved in cellular transport, amino acid metabolism and signal transduction, etc. In conclusion, CgOPT2 is an important oligopeptide transporter of C. gloeosporioides, which participates in the process of vegetative growth, cell wall integrity, oxidative stress, conidia germination and pathogenicity.

Genome-Wide Identification and Comparative Analysis for OPT Family Genes in Panax ginseng and Eleven Flowering Plants.

Herb genomics and comparative genomics provide a global platform to explore the genetics and biology of herbs at the genome level. Panax ginseng C.A. Meyer is an important medicinal plant for a variety of bioactive chemical compounds of which the biosynthesis may involve transport of a wide range of substrates mediated by oligopeptide transporters (OPT). However, information about the OPT family in the plant kingdom is still limited. Only 17 and 18 OPT genes have been characterized for Oryza sativa and Arabidopsis thaliana, respectively. Additionally, few comprehensive studies incorporating the phylogeny, gene structure, paralogs evolution, expression profiling, and co-expression network between transcription factors and OPT genes have been reported for ginseng and other species. In the present study, we performed those analyses comprehensively with both online tools and standalone tools. As a result, we identified a total of 268 non-redundant OPT genes from 12 flowering plants of which 37 were from ginseng. These OPT genes were clustered into two distinct clades in which clade-specific motif compositions were considerably conservative. The distribution of OPT paralogs was indicative of segmental duplication and subsequent structural variation. Expression patterns based on two sources of RNA-Sequence datasets suggested that some OPT genes were expressed in both an organ-specific and tissue-specific manner and might be involved in the functional development of plants. Further co-expression analysis of OPT genes and transcription factors indicated 141 positive and 11 negative links, which shows potent regulators for OPT genes. Overall, the data obtained from our study contribute to a better understanding of the complexity of the OPT gene family in ginseng and other flowering plants. This genetic resource will help improve the interpretation on mechanisms of metabolism transportation and signal transduction during plant development for Panax ginseng.

Genome-wide analysis indicates diverse physiological roles of the turnip (Brassica rapa var. rapa) oligopeptide transporters gene family

Oligopeptide transporters (OPTs) encode integral membrane-localized proteins and have a broad range of substrate transport capabilities. Here, 28 BrrOPT genes were identified in the turnip. Phylogenetic analyses revealed two well-supported clades in the OPT family, containing 15 BrrOPTs and 13 BrrYSLs. The exon/intron structure of OPT clade are conserved but the yellow stripe-like (YSL) clade was different. The exon/intron of the YSL clade possesses structural differences, whereas the YSL class motifs structure are conserved. The OPT genes are distributed unevenly among the chromosomes of the turnip genome. Phylogenetic and chromosomal distribution analyses revealed that the expansion of the OPT gene family is mainly attributable to segmental duplication. For the expression profiles at different developmental stages, a comprehensive analysis provided insights into the possible functional divergence among members of the paralog OPT gene family. Different expression levels under a variety of ion deficiencies also indicated that the OPT family underwent functional divergence during long-term evolution. Furthermore, BrrOPT8.1, BrrYSL1.2, BrrYSL1.3, BrrYSL6 and BrrYSL9 responded to Fe(II) treatments and BrrYSL7 responded to calcium treatments, BrrYSL6 responded to multiple treatments in root, suggesting that turnip OPTs may be involved in mediating cross-talk among different ion deficiencies. Our data provide important information for further functional dissection of BrrOPTs, especially in transporting metal ions and nutrient deficiency stress adaptation.

Transport Deficiency Is the Molecular Basis of Candida albicans Resistance to Antifungal Oligopeptides.

Oligopeptides incorporating N3-(4-methoxyfumaroyl)-L-2,3-diaminopropanoic acid (FMDP), an inhibitor of glucosamine-6-phosphate synthase, exhibited growth inhibitory activity against Candida albicans, with minimal inhibitory concentration values in the 0.05–50 μg mL-1 range. Uptake by the peptide permeases was found to be the main factor limiting an anticandidal activity of these compounds. Di- and tripeptide containing FMDP (F2 and F3) were transported by Ptr2p/Ptr22p peptide transporters (PTR) and FMDP-containing hexa-, hepta-, and undecapeptide (F6, F7, and F11) were taken up by the oligopeptide transporters (OPT) oligopeptide permeases, preferably by Opt2p/Opt3p. A phenotypic, apparent resistance of C. albicans to FMDP-oligopeptides transported by OPT permeases was triggered by the environmental factors, whereas resistance to those taken up by the PTR system had a genetic basis. Anticandidal activity of longer FMDP-oligopeptides was strongly diminished in minimal media containing easily assimilated ammonium sulfate or L-glutamine as the nitrogen source, both known to downregulate expression of the OPT genes. All FMDP-oligopeptides tested were more active at lower pH and this effect was slightly more remarkable for peptides F6, F7, and F11, compared to F2 and F3. Formation of isolated colonies was observed inside the growth inhibitory zones induced by F2 and F3 but not inside those induced by F6, F7, and F11. The vast majority (98%) of those colonies did not originate from truly resistant cells. The true resistance of 2% of isolates was due to the impaired transport of di- and to a lower extent, tripeptides. The resistant cells did not exhibit a lower expression of PTR2, PTR22, or OPT1–3 genes, but mutations in the PTR2 gene resulting in T422H, A320S, D119V, and A320S substitutions in the amino acid sequence of Ptr2p were found.

Transcriptional responses in Lactococcus lactis subsp. cremoris to the changes in oxygen and redox potential during milk acidification.

Milk acidification and metabolic activity of the starter cultures are affected by oxygen; however, molecular factors related to the redox changes are poorly defined. The objective of the study was to investigate transcriptional responses in Lactococcus lactis subsp. cremoris CHCCO2 grown in milk to the shifts of oxygen and redox potential (Eh7 ). Transcriptomic studies were performed with the use of Illumina HiSeq 2000 mRNA sequencing and validated by the real-time quantitative PCR. In total 105 differentially expressed genes were assigned functional gene names. Most of the differentially expressed genes were detected during aerobic reduction phase. Upregulated genes were implicated in lactose utilization, glycogen biosynthesis, amino sugar metabolism, oxidation-reduction, pyrimidine biosynthesis and DNA integration processes. Genes of purine nucleotide biosynthesis and genes encoding amino acid, multidrug resistance and ion ABC transporters were mostly downregulated, while oligopeptide transporter genes were reduced during oxygen depletion and induced at minimum Eh7 . Understanding of gene responses in starter cultures to the changes of oxidation-reduction state is important for the better control and reproducibility of dairy fermentations. We applied mRNA sequencing by Illumina HiSeq 2000 to investigate gene expression profile in a dairy strain of Lactococcus lactis subsp. cremoris during milk acidification. Novelty of this study lies in linking transcriptional responses to oxygen depletion and the changes of redox potential with the fermentation kinetics and clarification of molecular factors specifically expressed in milk which might be essential for bacterial performance and the final quality of cheeses.

Effects of Collagen Ingestion and their Biological Significance

Collagen is the most abundant extracellular matrix protein in animal tissues, and heat-denatured collagen (gelatin) and its hydrolysate (collagen peptide) are frequently used as food supplements. Clinical trials revealed that supplemental ingestion of 5-10 g collagen peptide daily improved the properties of facial skin, suppressed ultraviolet-induced skin erythema, and improved the T-cell-related immune status of Japanese people who ingest an average of 1.9 g collagen from their daily diet. Ingested collagen is digested and absorbed partly as prolylhydroxyproline (Pro-Hyp) and hydroxyprolylglycine (Hyp-Gly), resulting in their rather high concentrations in blood. The beneficial effects of collagen peptide ingestion appear to be mediated at least partly by these collagenderived dipeptides. Collagen-derived Pro-Hyp exhibits physiological activity in the in vitro differentiation processes of skin fibroblasts, chondrocytic cells, and pre-adipocytes; in addition, the expression of oligopeptide transporter genes has been confirmed in pre-adipocytes. Thus, it is tempting to speculate that the target cells of Pro-Hyp are precursor cells. Pro-Hyp is generated endogenously when collagen turnover is enhanced, such as in an ear inflamed by contact dermatitis, and ingested Pro-Hyp appears in the same dermatitic ear when ingested orally. Thus, endogenous and food-derived Pro-Hyp co-localizes in the same tissue and may co-interact. Further studies on Pro- Hyp may reveal novel interactions between homeostasis of the tissue and this animal-derived nutrient.

Regulatory Networks Controlling Nitrogen Sensing and Uptake in Candida albicans

Nitrogen is one of the key nutrients for microbial growth. During infection, pathogenic fungi like C. albicans need to acquire nitrogen from a broad range of different and changing sources inside the host. Detecting the available nitrogen sources and adjusting the expression of genes for their uptake and degradation is therefore crucial for survival and growth as well as for establishing an infection. Here, we analyzed the transcriptional response of C. albicans to nitrogen starvation and feeding with the infection-relevant nitrogen sources arginine and bovine serum albumin (BSA), representing amino acids and proteins, respectively. The response to nitrogen starvation was marked by an immediate repression of protein synthesis and an up-regulation of general amino acid permeases, as well as an up-regulation of autophagal processes in its later stages. Feeding with arginine led to a fast reduction in expression of general permeases for amino acids and to resumption of protein synthesis. The response to BSA feeding was generally slower, and was additionally characterized by an up-regulation of oligopeptide transporter genes. From time-series data, we inferred network interaction models for genes relevant in nitrogen detection and uptake. Each individual network was found to be largely specific for the experimental condition (starvation or feeding with arginine or BSA). In addition, we detected several novel connections between regulator and effector genes, with putative roles in nitrogen uptake. We conclude that C. albicans adopts a particular nitrogen response network, defined by sets of specific gene-gene connections for each environmental condition. All together, they form a grid of possible gene regulatory networks, increasing the transcriptional flexibility of C. albicans.

Comparison of Metabolic Capacities and Inference of Gene Content Evolution in Mosquito-Associated Spiroplasma diminutum and S. taiwanense

Mosquitoes are hosts of several Spiroplasma species that belong to different serogroups. To investigate the genetic mechanisms that may be involved in the utilization of similar hosts in these phylogenetically distinct bacteria, we determined the complete genome sequences of Spiroplasma diminutum and S. taiwanense for comparative analysis. The genome alignment indicates that their chromosomal organization is highly conserved, which is in sharp contrast to the elevated genome instabilities observed in other Spiroplasma lineages. Examination of the substrate utilization strategies revealed that S. diminutum can use a wide range of carbohydrates, suggesting that it is well suited to living in the gut (and possibly the circulatory system) of its mosquito hosts. In comparison, S. taiwanense has lost several carbohydrate utilization genes and acquired additional sets of oligopeptide transporter genes through tandem duplications, suggesting that proteins from digested blood meal or lysed host cells may be an important nutrient source. Moreover, one glycerol-3-phosphate oxidase gene (glpO) was found in S. taiwanense but not S. diminutum. This gene is linked to the production of reactive oxygen species and has been shown to be a major virulence factor in Mycoplasma mycoides. This finding may explain the pathogenicity of S. taiwanense observed in previous artificial infection experiments, while no apparent effect was found for S. diminutum. To infer the gene content evolution at deeper divergence levels, we incorporated other Mollicutes genomes for comparative analyses. The results suggest that the losses of biosynthetic pathways are a recurrent theme in these host-associated bacteria.

Submergence confers immunity mediated by the WRKY22 transcription factor in Arabidopsis.

Transcriptional control plays an important role in regulating submergence responses in plants. Although numerous genes are highly induced during hypoxia, their individual roles in hypoxic responses are still poorly understood. Here, we found that expression of genes that encode members of the WRKY transcription factor family was rapidly and strongly induced upon submergence in Arabidopsis thaliana, and this induction correlated with induction of a large portion of innate immunity marker genes. Furthermore, prior submergence treatment conferred higher resistance to the bacterial pathogen Pseudomonas syringae in Arabidopsis. Among the WRKY genes tested, WRKY22 had the highest level of induction during the early stages of submergence. Compared with the wild type, WRKY22 T-DNA insertion mutants wrky22-1 and wrky22-2 had lower disease resistance and lower induction of innate immunity markers, such as FLG22-INDUCED RECEPTOR-LIKE KINASE1 (FRK1) and WRKY53, after submergence. Furthermore, transcriptomic analyses of wrky22-2 and chromatin immunoprecipitation identified several potential targets of WRKY22, which included genes encoding a TIR domain-containing protein, a plant peptide hormone, and many OLIGO PEPTIDE TRANSPORTER genes, all of which may lead to induction of innate immunity. In conclusion, we propose that submergence triggers innate immunity in Arabidopsis via WRKY22, a response that may protect against a higher probability of pathogen infection either during or after flooding.

An oligopeptide transporter gene family in Phanerochaete chrysosporium

Oligopeptide transporters are believed to translocate their substrates across the membrane from the extracellular environment into cell. In the present study, sixteen oligopeptide transporter (OPT) orthologs (PcOPT1 to PcOPT16) were identified in Phanerochaete chrysosporium strain BKMF1767. They were predicted to encode integral membrane proteins with 13 to 16 transmembrane domains. The cDNA and genomic sequences of these genes were confirmed by clone sequencing and Transcriptome Sequencing. Based on sequence similarities, the PcOPT genes can be grouped into three different subfamilies. Comprehensive analysis revealed correlations between gene groupings and gene structures. The PcOPT genes showed differential expression patterns at different metabolic stages or in various stress conditions, suggesting that they may play different functional roles. PcOPT14 was able to mediate the uptake of tetrapeptide KLGL when heterologously expressed in Saccharomyces cerevisiae. This study provides some basic genomic information for OPT genes in P. chrysosporium and will be useful for elucidating their roles in growth, stress response and survival of this organism. Moreover, the results can help us to better understand the complexity of metabolism of P. chrysosporium.

Evolutionary expansion and functional diversification of oligopeptide transporter gene family in rice

BackgroundOligopeptide transporters (OPTs) play important roles in the mobilization of organic nitrogenous compounds and usually associate with tissues that show signs of rapid protein hydrolysis, such as germinating seeds and senescing leaves. This study is to investigate rice OPT genes.ResultsA total of sixteen OsOPT genes (Os for Oryza sative L.) were identified in the rice genome, which were then classified into six sections that belong to two subfamilies (the PT and YSL subfamily). The major mechanisms for evolutionary expansion of the sixteen genes during the rice genome evolution include segmental and tandem duplication. Calculation of the duplication event dates indicated that the sixteen genes originated from nine original OsOPT genes, and the duplication events could be classified into three evolutionary stages. The first evolutionary stage occurred approximately 50 million years ago (Mya) and involved the evolution of four new genes. The second evolutionary stage was approximately 20 Mya and was marked by the appearance of two new genes, and the third evolutionary stage was approximately 9 Mya when two new genes evolved. Mining of the expression database and RT-PCR analysis indicated that the expression of most duplicated OsOPT genes showed high tissue specificities. Diverse expression patterns for the sixteen genes were evaluated using both semi-quantitative RT-PCR and the MPSS data. Expression levels of some OsOPT genes were regulated by abiotic and biotic stresses suggesting the potential involvement of these gene products in rice stress adaptation. Five OsOPT gene mutants showed abnormal development and growth, the primary analysis of five OsOPT gene mutants suggested that they may be necessary for rice development.ConclusionsThese results suggested that rice-specific OsOPT genes might be potentially useful in improving rice.Electronic supplementary materialThe online version of this article (doi:10.1186/1939-8433-5-12) contains supplementary material, which is available to authorized users.

Repressive processing of antihypertensive peptides, Val-Pro-Pro and Ile-Pro-Pro, in Lactobacillus helveticus fermented milk by added peptides

Lactobacillus helveticus can release the antihypertensive peptides, Val-Pro-Pro (VPP) and Ile-Pro-Pro (IPP), from casein in fermented milk by a specific proteolytic system. To better understand the regulation of gene expression of the proteolytic enzymes thought to link to the processing of both antihypertensive peptides in L. helveticus, microarray analysis for whole gene expression in the presence and absence of added peptides in the fermented milk was studied. The productivity of both VPP and IPP in L. helveticus CM4 fermented milk was repressed by adding 2% quantity of Peptone as peptide mixture to the milk. Among the selected 13 amino acids, Gly, Ile, Leu, Phe, Met, Ser and Val were effective in the repression of the productivity of VPP and IPP in the fermented milk. The activity of the cell wall-associated proteinase, which may play a key role in the processing of the two antihypertensive peptides, was significantly repressed by the addition of the 2% quantity of Peptone into the fermented milk. By DNA microarray analysis it was found that prtH2 corresponding to the cell wall-associated proteinase gene, most of the endopeptidase genes such as pepE, pepO1, pepO2 and pepO3, most of the oligopeptide transporter genes, such as dppA2, dppB, dppC, dppD and dppF, most likely involved in the processing of VPP and IPP were down-regulated. These results suggest that amino acids released from milk peptides in the fermented milk might down-regulate the gene expressions of some of the proteolytic enzymes and may cause repression of the release of VPP and IPP in L. helveticus fermented milk.

Analyses of the oligopeptide transporter gene family in poplar and grape

BackgroundOligopeptide transporters (OPTs) are a group of membrane-localized proteins that have a broad range of substrate transport capabilities and that are thought to contribute to many biological processes. The OPT proteins belong to a small gene family in plants, which includes about 25 members in Arabidopsis and rice. However, no comprehensive study incorporating phylogeny, chromosomal location, gene structure, expression profiling, functional divergence and selective pressure analysis has been reported thus far for Populus and Vitis.ResultsIn the present study, a comprehensive analysis of the OPT gene family in Populus (P. trichocarpa) and Vitis (V. vinifera) was performed. A total of 20 and 18 full-length OPT genes have been identified in Populus and Vitis, respectively. Phylogenetic analyses indicate that these OPT genes consist of two classes that can be further subdivided into 11 groups. Gene structures are considerably conserved among the groups. The distribution of OPT genes was found to be non-random across chromosomes. A high proportion of the genes are preferentially clustered, indicating that tandem duplications may have contributed significantly to the expansion of the OPT gene family. Expression patterns based on our analyses of microarray data suggest that many OPT genes may be important in stress response and functional development of plants. Further analyses of functional divergence and adaptive evolution show that, while purifying selection may have been the main force driving the evolution of the OPTs, some of critical sites responsible for the functional divergence may have been under positive selection.ConclusionsOverall, the data obtained from our investigation contribute to a better understanding of the complexity of the Populus and Vitis OPT gene family and of the function and evolution of the OPT gene family in higher plants.

Arabidopsis OPT6 is an Oligopeptide Transporter with Exceptionally Broad Substrate Specificity

Oligopeptide transporters (OPTs) are found in fungi, bacteria and plants. The nine Arabidopsis thaliana OPT genes are expressed mainly in the vasculature and are thought to transport tetra- and pentapeptides, and peptide-like substrates such as glutathione. Expression of AtOPT6 in Xenopus laevis oocytes demonstrated that AtOPT6 transports many tetra- and pentapeptides. In addition, AtOPT6 transported reduced glutathione (GSH), a tripeptide, but not oxidized glutathione (GSSG). Recent data showed that Candida albicans OPTs can transport peptides up to eight amino acids in length. AtOPT6 transported mammalian signaling peptides up to 10 amino acids in length and, in addition, known plant development- and nematode pathogenesis-associated peptides up to 13 amino acids long. AtOPT6 displayed high affinity for penta- and dodecapeptides, but low affinity for GSH. In comparison the Saccharomyces cerevisiae ScOPT1 was incapable of transporting any of the longer peptides tested. These data demonstrate the necessity of experimentally determining substrate specificity of individual OPTs, and lay a foundation for structure/function studies. Characterization of the AtOPT6 substrate range provides a basis for investigating the possible physiological function of AtOPT6 in peptide signaling and thiol transport in response to stress.

Characterization of the PT Clade of Oligopeptide Transporters in Rice

Oligopeptide transporters (OPTs) are a group of membrane‐localized proteins that have a broad range of substrate transport capabilities and that are thought to contribute to many biological processes. Nine OPTs belonging to the peptide transport (PT) clade were identified in the rice (Oryza sativa L.) genome, based on their predicted amino acid sequences containing the two highly conserved motifs (NPG and KIPPR) that define the PT‐OPT subgroup. Protein identity analysis comparing the rice OPTs of the PT and yellow stripe (YS) clades revealed low levels of similarity between members of these two OPT subgroups. Sequence analysis of the predicted proteins of the PT clade, named OsOPT1 to OsOPT9, revealed all to be integral membrane proteins. Functional analysis of five of the OsOPTs, using a yeast complementation assay, demonstrated that OsOPT1, OsOPT3, OsOPT4, OsOPT5, and OsOPT7 could transport ferrous and/or ferric iron chelated to nicotianamine, a nonpeptidyl metal‐chelate. Semi quantitative reverse‐transcriptase polymerase chain reaction verified expression of the OsOPTs in roots, stems, leaves, hulls, pedicels, and embryos, but transcripts for each OsOPT were not found in every tissue. The identification of multiple PT‐clade OPT genes in rice with different expression patterns indicates that they may have different biological roles in the plant.

Expression Profiles of Arabidopsis thaliana in Mineral Deficiencies Reveal Novel Transporters Involved in Metal Homeostasis

Plants directly assimilate minerals from the environment and thus are key for acquisition of metals by all subsequent consumers. Limited bio-availability of copper, zinc and iron in soil decreases both the agronomic productivity and the nutrient quality of crops. Understanding the molecular mechanisms underlying metal homeostasis in plants is a prerequisite to optimizing plant yield and metal nutrient content. To absorb and maintain a balance of potentially toxic metal ions, plants utilize poorly understood mechanisms involving a large number of membrane transporters and metal binding proteins with overlapping substrate specificities and complex regulation. To better understand the function and the integrated regulation, we analyzed in Arabidopsis the expression patterns in roots and in leaves of 53 genes coding for known or potential metal transporters, in response to copper, zinc, and iron deficiencies in Arabidopsis. Comparative analysis of gene expression profiles revealed specific transcriptional regulation by metals of the genes contrasting with the known wide substrate specificities of the encoded transporters. Our analysis suggested novel transport roles for several gene products and we used functional complementation of yeast mutants to correlate specific regulation by metals with transport activity. We demonstrate that two ZIP genes, ZIP2 and ZIP4, are involved in copper transport. We also present evidence that AtOPT3, a member of the oligopeptide transporter gene family with significant similarities to the maize iron-phytosiderophore transporter YS1, is regulated by metals and heterologous expression AtOPT3 can rescue yeast mutants deficient in metal transport.