Yellow Stripe-Like Transporters Research Articles

A mutation in the promoter of the yellow stripe-like transporter gene in cucumber results in a yellow cotyledon phenotype

Leaf color mutants in higher plants are considered to be ideal materials for studying the chlorophyll biosynthesis, photosynthesis mechanism and chloroplast development. Herein, we identified a spontaneous mutant, yc412, in cultivated cucumber that exhibited yellow cotyledons. The yellow-lethal mutant was diagnosed with an abnormal chloroplast ultrastructure, and reduced photosynthetic capacity and pigment content. Through bulked segregant analysis-based whole-genome sequencing and fine genetic mapping, we narrowed the yellow cotyledons (yc) locus to a 96.8 kb interval on chromosome 3. By resequencing and molecular cloning, we showed that Csyc is a potential candidate gene, which encodes a yellow stripe-like (YSL) transporter. The T to C mutation in the promoter region of Csyc caused the yellow cotyledon phenotype in yc412. Compared to YZU027A (WT), the expression of Csyc was significantly downregulated in the cotyledons of yc412. Silencing of Csyc in cucumber via virus-induced gene silencing resulted in chlorotic leaves, mainly by suppressing the chlorophyll content. Furthermore, a comparative transcriptome analysis revealed that chloroplast-related genes and chlorophyll biosynthesis genes were significantly downregulated in yc412 cotyledons. Our results provide new insights into the molecular function of the YSL transporter in plant chloroplast development and chlorophyll synthesis.

MsYSL6, A Metal Transporter Gene of Alfalfa, Increases Iron Accumulation and Benefits Cadmium Resistance.

Iron (Fe) is necessary for plant growth and development. The mechanism of uptake and translocation in Cadmium (Cd) is similar to iron, which shares iron transporters. Yellow stripe-like transporter (YSL) plays a pivotal role in transporting iron and other metal ions in plants. In this study, MsYSL6 and its promoter were cloned from leguminous forage alfalfa. The transient expression of MsYSL6-GFP indicated that MsYSL6 was localized to the plasma membrane and cytoplasm. The expression of MsYSL6 was induced in alfalfa by iron deficiency and Cd stress, which was further proved by GUS activity driven by the MsYSL6 promoter. To further identify the function of MsYSL6, it was heterologously overexpressed in tobacco. MsYSL6-overexpressed tobacco showed better growth and less oxidative damage than WT under Cd stress. MsYSL6 overexpression elevated Fe and Cd contents and induced a relatively high Fe translocation rate in tobacco under Cd stress. The results suggest that MsYSL6 might have a dual function in the absorption of Fe and Cd, playing a role in the competitive absorption between Fe and Cd. MsYSL6 might be a regulatory factor in plants to counter Cd stress. This study provides a novel gene for application in heavy metal enrichment or phytoremediation and new insights into plant tolerance to toxic metals.

Identification of temporally distributed candidate genes for high iron (Fe) and zinc (Zn) content in wheat (Triticum aestivum L.)

Micronutrient deficiency causes a major global public health issues. Increasing micronutrient content through biofortification can be achieved by exploring molecular regulatory networks for micronutrients metabolism in cereal crops. The present study provides regulatory gene networks for micronutrients homeostasis through transcriptome analysis at two seed developmental stages (14 and 28 days after anthesis) in three micronutrients variant wheat genotypes, ‘FAR 4’, ‘HD 3226’ and ‘K 8027’ at Illumina Hiseq-2500 platform. Differential gene expression (DGE) with gene ontology and KEGG pathway analysis, revealed the role of iron binding protein ferritin, nicotianamine synthase, and Yellow Stripe-Like transporters in maintaining higher grain micronutrient contents in ‘FAR 4’ genotype. The gene regulatory network analysis suggested the possible interactions between the protein and associated transcription factors and revealed NAC TFs, as the major regulatory factor in ‘FAR 4’ responsible for micronutrient homeostasis. Further, 11 F-box and 12 NBS-LRR disease resistance protein genes were abundant transcriptionally across the two seed development stages in all these wheat genotypes. The potential candidate genes may be further exploited to increase the micronutrient contents in cereal grains making them more nutrition for human consumption.

Ammonium-nitrogen addition at the seedling stage does not reduce grain cadmium concentration in two common wheat (Triticum aestivum L.) cultivars

High cadmium (Cd) concentration in common wheat (Triticum aestivum L.) grains poses potential health risks. Several management strategies have been used to reduce grain Cd concentration. However, limited information is available on the use of ammonium-nitrogen (NH4+-N) as a strategy to manage Cd concentration in wheat grains. In this study, NH4+-N addition at the seedling stage unchanged the grain Cd concentration in the high-Cd accumulator, Zhoumai 18 (ZM18), but dramatically increased that in the low-Cd accumulator, Yunmai 51 (YM51). Further analysis revealed that the effects of NH4+-N addition on whole-plant Cd absorption, root-to-shoot Cd translocation, and shoot-to-grain Cd remobilization were different between the two wheat cultivars. In ZM18, NH4+-N addition did not change whole-plant Cd absorption, but inhibited root-to-shoot Cd translocation and Cd remobilization from lower internodes, lower leaves, node 1, and internode 1 to grains via the down-regulation of yellow stripe-like transporters (YSL), zinc transporters (ZIP5, ZIP7, and ZIP10), and heavy-metal transporting ATPases (HMA2). This inhibition decreased the grain Cd content by 29.62%, which was consistent with the decrease of the grain dry weight by 23.26%, leading to unchanged grain Cd concentration in ZM18. However, in YM51, NH4+-N addition promoted continuous Cd absorption during grain filling, root-to-shoot Cd translocation and whole-plant Cd absorption. The absorbed Cd was directly transported to internode 1 via the xylem and then re-transported to grains via the phloem by up-regulated YSL, ZIP5, and copper transporters (COPT4). This promotion increased the grain Cd content by 245.35%, which was higher than the increased grain dry weight by 132.89%, leading to increased grain Cd concentration in YM51. Our findings concluded that the addition of NH4+-N fertilizer at the seedling stage is not suitable for reducing grain Cd concentration in common wheat cultivars.

The role of the yellow stripe-like transporter BdYSL3 in copper homeostasis in Brachypodium.

The role of the yellow stripe-like transporter BdYSL3 in copper homeostasis in Brachypodium.

TaYS1A, a Yellow Stripe-Like Transporter Gene, Is Required for Wheat Resistance to Puccinia striiformis f. sp. Tritici.

Yellow stripe-like (YSL) transporters are required for the transportation of metal-phytosiderophores and are structurally related to metal-nicotianamine complexes. Some studies also reported the involvement of YSL transporters in pathogen-induced defense. However, the molecular mechanisms of YSL genes involved in biotic stress responses are still not clear, especially in cereal crops. This study aimed to functionally characterize TaYS1A during the interaction of wheat and Puccinia striiformis f. sp. tritici (Pst), the causal agent of stripe rust disease. TaYS1A was localized in the cell membrane of wheat protoplasts and Nicotiana benthamiana cells. TaYS1A was significantly up-regulated in wheat leaves after being infected with the avirulent Pst isolate CYR23 and after treatment with salicylic acid (SA). Silencing of TaYS1A by the virus-induced gene silencing method enhanced the susceptibility of wheat to Pst accompanied by reducing the accumulation of SA and H2O2 and down-regulating the transcriptions of TaPR1 and TaPR2. In addition, TaYS1A was found to interact with TaNH2, a homolog of OsNH2, by yeast-two-hybrid assay, and silencing of TaYS1A diminished the expression of TaNH2. Our findings suggested the existence of positive regulation of TaYS1A in providing resistance against Pst by modulating SA-induced signaling and offered new insight into the biological role of YSL in wheat against pathogens.

Integrative analysis of hexaploid wheat roots identifies signature components during iron starvation.

Iron (Fe) is an essential micronutrient for all organisms. In crop plants, Fe deficiency can decrease crop yield significantly; however, our current understanding of how major crops respond to Fe deficiency remains limited. Herein, the effect of Fe deprivation at both the transcriptomic and metabolic level in hexaploid wheat was investigated. Genome-wide gene expression reprogramming was observed in wheat roots subjected to Fe starvation, with a total of 5854 genes differentially expressed. Homoeologue and subgenome-specific analysis unveiled the induction-biased contribution from the A and B genomes. In general, the predominance of genes coding for nicotianamine synthase, yellow stripe-like transporters, metal transporters, ABC transporters, and zinc-induced facilitator-like protein was noted. Expression of genes related to the Strategy II mode of Fe uptake was also predominant. Our transcriptomic data were in agreement with the GC-MS analysis that showed the enhanced accumulation of various metabolites such as fumarate, malonate, succinate, and xylofuranose, which could be contributing to Fe mobilization. Interestingly, Fe starvation leads to a significant temporal increase of glutathione S-transferase at both the transcriptional level and enzymatic activity level, which indicates the involvement of glutathione in response to Fe stress in wheat roots. Taken together, our result provides new insight into the wheat response to Fe starvation at the molecular level and lays the foundation to design new strategies for the improvement of Fe nutrition in crops.

Copper accumulation, subcellular partitioning and physiological and molecular responses in relation to different copper tolerance in apple rootstocks.

To unravel the physiological and molecular regulation mechanisms underlying the variation in copper (Cu)accumulation, translocation and tolerance among five apple rootstocks, seedlings were exposed to either basal or excess Cu. Excess Cu suppressed plant biomass and root architecture, which was less pronounced in Malus prunifolia Borkh., indicating its relatively higher Cu tolerance. Among the five apple rootstocks, M. prunifolia exhibited the highest Cu concentration and bio-concentration factor in roots but the lowest translocation factor, indicating its greater ability to immobilize Cu and restrict translocation to the aerial parts. Higher Cu concentration in cell wall fraction but lower Cu proportion in membrane-containing and organelle-rich fractions were found in M. prunifolia. Compared with the other four apple rootstocks under excess Cu conditions, M. prunifolia had a lower increment of hydrogen peroxide in roots and leaves and malondialdehyde in roots, but higher concentrations of carbohydrates and enhanced antioxidants. Transcript levels of genes involved in Cu uptake, transport and detoxification revealed species-specific differences that are probably related to alterations in Cu tolerance. M. prunifolia had relatively higher gene transcript levels including copper transporters 2 (COPT2), COPT6 and zinc/iron-regulated transporter-related protein 2 (ZIP2), which probably took part in Cu uptake, and C-type ATP-binding cassette transporter 2 (ABCC2), copper chaperone for Cu/Zn superoxide dismutase (CCS), Cu/Zn superoxide dismutase 1 (CSD1) and metallothionein 2 (MT2) probably implicated in Cu detoxification, and relatively lower mRNA levels of yellow stripe-like transporter 3 (YSL3) and heavy metal ATPase 5 (HMA5) involved in transport of Cu to aerial parts. These results suggest that M. prunifolia is more tolerant to excess Cu than the other four apple rootstocks under the current experimental conditions, which is probably attributed to more Cu retention in roots, subcellular partitioning, well-coordinated antioxidant defense mechanisms and transcriptional expression of genes involved in Cu uptake, translocation and detoxification.

Genome-wide analysis of oligopeptide transporters and detailed characterization of yellow stripe transporter genes in hexaploid wheat.

Oligopeptide transporters (OPT) are integral cell membrane proteins that play a critical role in the transport of small peptides, secondary amino acids, glutathione conjugates, and mineral uptake. In the present study, 67 putative wheat yellow stripe-like transporter (YSL) proteins belonging to the subfamily of OPT transporters were identified. Phylogeny analysis resulted in the distribution of wheat YSLs into four discrete clades. The highest number of YSLs was present on the A genome and the chromosome 2 of hexaploid wheat. The identified wheat YSL genes showed differential expression in different tissues and during grain development suggesting the importance of this subfamily. Gene expression pattern of TaYSLs during iron starvation experiments suggested an early high transcript accumulation of TaYS1A, TaYS1B, TaYSL3, TaYSL5, and TaYSL6 in roots. In contrast, delayed expression was observed in shoots for TaYS1A, TaYS1B, TaYSL5, TaYSL12, and TaYSL19 as compared to control. Further, their expression under biotic and abiotic response emphasized their alternative functions during the plant growth and development. In conclusion, this work is the first comprehensive study of wheat YSL transporters and would be an important resource for prioritizing genes towards wheat biofortification.

Overexpression of a Miscanthus sacchariflorus yellow stripe-like transporter MsYSL1 enhances resistance of Arabidopsis to cadmium by mediating metal ion reallocation

The yellow stripe-like (YSL) family of transporters mediates the uptake, translocation, and distribution of various mineral elements in vivo by transferring metal ions chelated with phytosiderophore or nicotianamine (NA). However, little is known about the roles of the YSL genes against cadmium in planta. In this study, we first cloned and characterized a vital member of the YSL gene family, MsYSL1, from the bioenergy plant Miscanthus sacchariflorus. MsYSL1 localized in the plasma membrane and was widely expressed throughout the whole seedling with the highest expression level in the stem. In addition, its expression in the root was stimulated by excess manganese (Mn), cadmium (Cd), and lead, and a shortage of iron (Fe), zinc (Zn), and copper. Functional complementation in yeast indicated that MsYSL1 showed transport activity for Fe(II)–NA and Zn–NA, but not for Cd–NA. Although they exhibited no significant differences versus the wild type under normal cultivation conditions, MsYSL1-overexpressing Arabidopsis lines displayed a higher resistance to Cd accompanied by longer root lengths, lower Cd, Zn, and Mn levels in roots, and higher Cd, Fe, and Mn translocation ratios under Cd stress. Moreover, genes related to NA synthesis, metal translocation, long-distance transport, and Cd exclusion were highly induced in transgenic lines under Cd stress. Thus, MsYSL1 may be an essential transporter for diverse metal–NAs to participate in the Cd detoxification by mediating the reallocation of other metal ions.

Isolation and characterization of a novel cadmium-regulated Yellow Stripe-Like transporter (SnYSL3) in Solanum nigrum.

SnYSL3 encodes a plasma-localized transporter delivering various metal-nicotianamine complexes. The expression of SnYSL3 is up-regulated by excess Cd, suggesting an important role for SnYSL3 in response to Cd stress. The Yellow Stripe-Like (YSL) transporters have been proposed to participate in metal uptake and long-range transport in model plants. In this study, we isolated and characterized a novel member of the YSL gene family, SnYSL3, from the cadmium hyperaccumulator Solanum nigrum. SnYSL3 was constitutively expressed and encodes a plasma membrane-localized protein. In situ RNA hybridization localized the SnYSL3 transcripts predominantly in vascular tissues and epidermal cells of the roots and stems, while in leaves, the mRNA levels were high in the vasculature. The SnYSL3 expression level was up-regulated by excess Cd, excess Fe and Cu deficiency. Heterologous expression of SnYSL3 in yeast revealed that SnYSL3 transports nicotianamine complexes containing Fe(II), Cu, Zn and Cd. SnYSL3 overexpression in Arabidopsis thaliana decreased Fe and Mn concentrations in the roots and increased the root-to-shoot translocation ratios of Fe and Mn. Under Cd exposure, the transgenic plants showed increased translocation ratios of Fe and Cd, but no difference was observed in Mn translocation from roots to shoots between the transgenic and wild-type lines. Although the accurate function of SnYSL3 remains to be confirmed, these results suggest that SnYSL3 is a transporter delivering a broad range of metal-nicotianamine complexes and is potentially important for the response to heavy metal stress, especially due to Cd and Fe.

In-silico characterisation and comparative mapping of yellow stripe like transporters in five grass species

Iron (Fe) and Zinc (Zn) are the key elements required for many of the biological process in plants and animals. Transporter proteins are essential for uptake, transport and accumulation for Fe and Zn in plants. The present investigation was undertaken to study and compare the structural and functional diversity and evolutionary significance of the yellow stripe-like (YSL) transporters through in-silico tools in five species (barley, Brachypodium, foxtail millet, maize and rice) of Poaceae. One hundred and two YSL transporters collected from public databases were used in the analysis. All YSL transporters possessed PF03169 domain which belongs to the oligo peptide transporters (OPT) super family. Molecular weight of YSL proteins ranged from 11.10 to 84.70 kDa while pI values ranged from 4.99 to 11.64. Scondary structure analysis identified that, alpha helix and random coils were the most common structures of the YSL proteins. Phylogenetic analysis revealed that the YSL transporters are highly conserved in these five grass species. Comparative mapping of genes of YSL transporters showed maximum synteny between Brachypodium and barley (30%) followed by Brachypodium and rice (25%). Neutrality test has in fact revealed the positive or Darwinian selection on YSL transporters. The results of the present investigation provided a significant understanding of the structural and biological role of YSL transporters as well as the evolutionary pattern in Poaceae family.

Phylogenetic and Expression Analysis of Pear Yellow Stripe-Like Transporters and Functional Verification of PbrYSL4 in Pear Pollen

Yellow stripe-like (YSL) family transporters, belonging to the oligopeptide transporter family, are significant iron transport proteins. In this study, we provided a genome-wide identification and analysis of the YSL gene family in Pyrus bretschneideri. We found eight YSL gene members in pear, clustered into four main groups in the phylogenetic tree. Segmental duplication has played a key role in the expansion of the pear YSL family. The pollen activity analysis indicated that the low concentration of iron ion was beneficial to both pear pollen germination and pollen tube growth. Among the eight YSL genes, PbrYSL4 had particularly high expression in all pear tissues; it was significantly responsive to change in the external iron ion supply in the pollen cultivation in vitro. Moreover, expression of PbrYSL4 in yeast mutant Δccc1 (Ca 2+ -sensitive cross-complementer 1 mutant) made Δccc1 restore growth in high iron medium. These data together suggest that PbrYSL4 was involved in the movement of iron in the pear pollen tube growth.

Successful Reproduction Requires the Function of Arabidopsis YELLOW STRIPE-LIKE1 and YELLOW STRIPE-LIKE3 Metal-Nicotianamine Transporters in Both Vegetative and Reproductive Structures

Several members of the Yellow Stripe-Like (YSL) family of proteins are transporters of metals that are bound to the metal chelator nicotianamine or the related set of mugineic acid family chelators known as phytosiderophores. Here, we examine the physiological functions of three closely related Arabidopsis (Arabidopsis thaliana) YSL family members, AtYSL1, AtYSL2, and AtYSL3, to elucidate their role(s) in the allocation of metals into various organs of Arabidopsis. We show that AtYSL3 and AtYSL1 are localized to the plasma membrane and function as iron transporters in yeast functional complementation assays. By using inflorescence grafting, we show that AtYSL1 and AtYSL3 have dual roles in reproduction: their activity in the leaves is required for normal fertility and normal seed development, while activity in the inflorescences themselves is required for proper loading of metals into the seeds. We further demonstrate that the AtYSL1 and AtYSL2 proteins, when expressed from the AtYSL3 promoter, can only partially rescue the phenotypes of a ysl1ysl3 double mutant, suggesting that although these three YSL transporters are closely related and have similar patterns of expression, they have distinct activities in planta. In particular, neither AtYSL1 nor AtYSL2 is able to functionally complement the reproductive defects exhibited by ysl1ysl3 double mutant plants.