NRAMP1 Research Articles

NRAMP6 and NRAMP1 cooperatively regulate root growth and manganese translocation under manganese deficiency in Arabidopsis.

The essential micronutrient manganese (Mn) in plants regulates multiple biological processes including photosynthesis and oxidative stress. Some Natural Resistance-Associated Macrophage Proteins (NRAMPs) have been reported to play critical roles in Mn uptake and reutilization in low Mn conditions. NRAMP6 was demonstrated to regulate cadmium tolerance and iron utilization in Arabidopsis. Nevertheless, it is unclear whether NRAMP6 plays a role in Mn nutrition. Here, we report that NRAMP6 cooperates with NRAMP1 in Mn utilization. Mutation of NRAMP6 in nramp1 but not in a wild-type background reduces root growth and Mn translocation from the roots to shoots under Mn deficient conditions. Grafting experiments revealed that NRAMP6 expression in both the roots and shoots is required for root growth and Mn translocation under Mn deficiency. We also showed that NRAMP1 could replace NRAMP6 to sustain root growth under Mn deficiency, but not vice versa. Mn deficiency does not affect the transcript level of NRAMP6, but is able to increase and decrease the protein accumulation of NRAMP6 in roots and shoots, respectively. Furthermore, NRAMP6 can be localized to both the plasma membrane and endomembranes including the endoplasmic reticulum, and Mn deficiency enhances the localization of NRAMP6 to the plasma membrane in Arabidopsis plants. NRAMP6 could rescue the defective growth of the yeast mutant Δsmf2, which is deficient in endomembrane Mn transport. Our results reveal the important role of NRAMP6 in Mn nutrition and in the long-distance signaling between the roots and shoots under Mn deficient conditions.

IDENTIFICATION OF GENES REGULATED IN RESPONSE TO Cu EXPOSURE IN Brassica nigra L.

Copper (Cu) is one of the essential trace metals required for plant growth. High amount of Cu in the media inhibits plant growth and is toxic to the plants. Brassica nigra L., a Cu accumulator, can tolerate a high amount of Cu and have specific mechanisms to relocate Cu within the cell compartments and keep the toxic amount of Cu away from the cytoplasm. This study aimed to evaluate the Cu-induced gene expression pattern of B. nigra Diyarbakir ecotype subjected to low Cu treatment. The Arabidopsis ATH1 genome array was used to determine the Cu-induced gene expression in the leaves of B. nigra grown at 25 µM Cu. Ninety-five genes were upregulated, and seventy-two genes were downregulated in the leaves of plants grown under 25 µM Cu. Cu responsive genes, such as glutathione S-transferase, glutathione reductase, heavy metal transporters, natural resistance-associated macrophage proteins, cytochrome p450, MYB-like transcription factor, copper/zinc, and Fe superoxide dismutases, and some protein kinases were highly expressed in the leaves of Cu-treated plants. The present work provides the global gene expression pattern in facultative metallophyte B. nigra, which could serve as a molecular tool for future phytoremediation studies.

Physiological and molecular characterization of strategy-I responses and expression of Fe-transporters in Fe-deficient soybean

Iron (Fe) deficiency is one of the major nutritional disorders that affect soybean production. This study aimed to better understand the adaptive responses and gene characterization in Fe-deficient soybean. The experiment was conducted on 14-day-old soybean plants in a soil pot under Fe sufficiency and bicarbonate-induced Fe deficiency. In soybean, Fe deficiency caused severe root and shoot morphological retardation, which was accompanied by the reductions in chlorophyll score, Fv/Fm (quantum yield efficiency of PSII), Pi_ABS (photosynthetic performance index) as well as an increase in electrolyte leakage and cell death. This is in accordance with the lower Fe status in the root and shoot of Fe-deficient plants. Further, the expression of GmIRT1 (Fe-regulated transporter) was significantly induced, while the members of NRAMP (natural resistance-associated macrophage proteins) transporters were either downregulated (GmNRAMP2a, GmNRAMP2b, GmNRAMP5a, GmNRAMP7) or constitutively expressed (GmNRAMP5b, GmNRAMP6a, GmNRAMP6b) in roots of Fe-deficient soybean relative to controls. Furthermore, the activity of Fe-chelate reductase and the expression of GmFRO2 (ferric reductase oxidase) in roots of Fe-deficient soybean were both significantly reduced, indicating the impairment of the ability of soybean plants to maintain Fe availability in the rhizosphere. However, plants showed a decrease in rhizospheric pH and upregulation of GmATPase4 in roots, indicating that rhizosphere acidification is one of the adaptive approaches that soybean plants possess in response to Fe-deficiency. In bioinformatics analysis, soybean Fe-uptake genes showed partnership with Arabidopsis FRO2, FRU, NRAMP1, bHLH38, and bHLH39 genes. In addition, Fe-starved plants demonstrated elevated H2O2 and O2•– in roots which were consistent with the inefficient oxidative defense of antioxidant enzymes. These findings could aid in the development of Fe-efficient soybean by breeding or genome editing techniques.

Genotypic structure of four cattle breeds raised in Turkey by loci related to several diseases

This study aims to reveal the genotypic structure of four cattle breeds; Holstein (HS), Turkish Grey Steppe (TGS), Anatolian Black (AB) and East Anatolian Red (EAR), raised in Turkey in terms of CD14, MBL, ITGB6, SLC11A1 and TLR2 genes and to evaluate their usefulness in Marker Assisted Selection (MAS). It also assesses whether the loci associated with resistance to diseases are suitable for phylogenetic analysis. Desired alleles and/or genotypes were detected in native Turkish cattle breeds at different frequencies in terms of polymorphisms of CD14, MBL, ITGB6, SLC11A1 and TLR2 genes which were previously reported to be associated with mastitis, foot-and-mouth disease and tuberculosis. These variations offer opportunities to improve selection strategies against diseases in the future. These results preliminary indicate that associated studies between these variations and disease resistance in native Turkish cattle breeds should be conducted. On the other hand, phylogenetic tree constructed based on genetic distance clearly separated native Turkish cattle breeds from HS breed. The gene regions related to diseases can be used to distinguish native cattle breeds from exotic ones.

Polymorphisms of SLC11A1(NRAMP1) rs17235409 associated with and susceptibility to spinal tuberculosis in a southern Han Chinese population.

To investigate the association between SLC11A1 (NRAMP1) rs17235409 (D543N) polymorphisms and susceptibility to spinal tuberculosis (STB) in the Han population in southern China. This study included 227 STB patients and 516 controls. Polymorphisms of SLC11A1 rs17235409 were genotyped using a SNPscan™ kit, and the protein was detected by western blotting. The genotype and allele frequency distributions of SLC11A1 rs17235409 differed significantly between the STB group and the control group(χ2=17.650, P=0.000). The distribution of GA genotype(GA vs. GG: P = 0.000, OR [95% CI] = 2.203[1.520-3.192] was significantly different between STB group and control group, but there was no significant difference in the distribution of AA genotypes(AA vs. GG: P=0.889, OR [95%CI]=0.674[0.142-3.208]). The A allele was more common in the STB group than in the control group (A vs. G: P=0.001, OR [95%CI]=1.767[1.273-2.452]). Under the dominant model, the GA+AA genotype was more common in the STB group than in the control group (GA+AA vs. GG: P=0.000, OR [95%CI]=2.067[1.438-2.971]). However, under the recessive model, there was no difference in GA+GG genotype between the STB and control groups(GA+GG vs. AA: P=0.701, OR [95%CI] =1.772[0.373-8.409]). NRAMP1 protein expression in the STB group(n=9) was significantly higher than that in the control group(n=9) (t=5.292,P=0.001). Variant genotypes at the rs17235409 locus of the SLC11A1 gene are associated with STB in the southern Han Chinese population. NRAMP1 protein expression is increased in patients with spinal tuberculosis, and the presence of the A allele increases the risk of developing STB.

The vacuolar transporter OsNRAMP2 mediates Fe remobilization during germination and affects Cd distribution to rice grain

Background and aimsIron (Fe) deficiency in plants is a common problem affecting agricultural production. Cadmium (Cd) is a toxic metal that can be taken up and transported within plants by transporters for divalent metals including Fe(II). The present study aims to investigate the functions of OsNRAMP2 (Natural Resistance-Associated Macrophage Protein 2) in the remobilization and distribution of Fe and Cd in rice.MethodsThe expression pattern of OsNRAMP2 was determined by quantitative real-time PCR and pOsNRAMP2::GUS assay. Knockout mutants of OsNRAMP2 were generated by using CRISPR/Cas9 gene editing. Localization of Fe in the vacuolar globoids of germinating seeds was imaged by high-resolution transmission electron microscopy coupled with energy-dispersive X-ray spectroscopy. Distributions of Fe and Cd between different plant tissues were investigated in hydroponic and soil pot experiments.ResultsOsNRAMP2 was mainly expressed in the embryo of germinating seeds, roots, leaf sheaths and leaf blades. OsNRAMP2 was localized at the tonoplast. Knockout of OsNRAMP2 delayed seed germination and produced chlorotic seedling leaves. Remobilization of Fe stored in the protein storage vacuoles in the scutellum of germinating seeds was restricted in osnramp2 mutants compared with wild type. Expression of genes related to Fe uptake was enhanced in the seedlings of osnramp2 mutants. Knockout of OsNRAMP2 significantly decreased the distribution of Cd, but not Fe, from leaves and straws to rice grains.ConclusionsOsNRAMP2 plays an important role in remobilizing vacuolar Fe during seed germination and affects translocation of Cd from vegetative tissues to rice grains.

Genetic Analysis of TB Susceptibility Variants in Ghana Reveals Candidate Protective Loci in SORBS2 and SCL11A1 Genes.

Despite advancements made toward diagnostics, tuberculosis caused by Mycobacterium africanum (Maf) and Mycobacterium tuberculosis sensu stricto (Mtbss) remains a major public health issue. Human host factors are key players in tuberculosis (TB) outcomes and treatment. Research is required to probe the interplay between host and bacterial genomes. Here, we explored the association between selected human/host genomic variants and TB disease in Ghana. Paired host genotype datum and infecting bacterial isolate information were analyzed for associations using a multinomial logistic regression. Mycobacterium tuberculosis complex (MTBC) isolates were obtained from 191 TB patients and genotyped into different phylogenetic lineages by standard methods. Two hundred and thirty-five (235) nondisease participants were used as healthy controls. A selection of 29 SNPs from TB disease-associated genes with high frequency among African populations was assayed using a TaqMan® SNP Genotyping Assay and iPLEX Gold Sequenom Mass Genotyping Array. Using 26 high-quality SNPs across 326 case-control samples in an association analysis, we found a protective variant, rs955263, in the SORBS2 gene against both Maf and Mtb infections (P BH = 0.05; OR = 0.33; 95% CI = 0.32–0.34). A relatively uncommon variant, rs17235409 in the SLC11A1 gene was observed with an even stronger protective effect against Mtb infection (MAF = 0.06; PBH = 0.04; OR = 0.05; 95% CI = 0.04–0.05). These findings suggest SLC11A1 and SORBS2 as a potential protective gene of substantial interest for TB, which is an important pathogen in West Africa, and highlight the need for in-depth host-pathogen studies in West Africa.

SLC11A1 genetic variation and low expression may cause immune response impairment in TB patients.

Tuberculosis (TB) is caused by Mycobacterium tuberculosis. Host genetic factors are important for the detection of TB susceptibility. SLC11A1 is located in monocyte phagolysosomes that help to limit M. tuberculosis growth by transferring divalent cations across the membrane. Genetic variation in SLC11A1 may alter its expression and increase the susceptibility of individuals to TB. The current study aimed to provide insight into host genetic variations and gene expression in TB patients. A total of 164 TB patients and 85 healthy controls were enrolled in this study. SLC11A1 polymorphisms were detected by PCR-RFLP. Real-time qPCR was used for SLC11A1 gene expression, and ELISA was used for protein estimation. GTEx Portal was used for quantitative trait loci analysis, while the STRING (v.11) web platform was used for gene interactive network construction. Data were analyzed using SPSS, GraphPad Prism, Haploview, and SNPstats. SLC11A1 polymorphisms and combinatorial genotypes were strongly associated with TB susceptibility, which may explain the greater prevalence of tuberculosis in the local population. Polymorphisms in SLC11A1 have also been linked to gene expression variation. Furthermore, the expression of SLC11A1 was downregulated in TB patients, which may influence the function of other associated genes and may impair the immunological response to tuberculosis.

Mechanics of Nramp-family transition metal transporters

Transition metal ions (like Mn2+, Fe2+ and Co2+) are nutrients crucial to metabolism in all living cells. The natural resistance associated macrophage protein (Nramp) transporters, found across the tree of life, enable cells to take up a range of essential divalent transition metals. Human Nramps are required for the dietary uptake and endosomal recycling of non-heme iron, and in the innate immune response to intracellular pathogens. Nramps are pH dependent and co-transport protons, and thus have been labeled as proton-coupled symporters.

TmNAS3 from Triticum monococum directly regulated by TmbHLH47 increases Fe content of wheat grain.

Biofortification is an effective way to enhance wheat grain Fe content. However, Fe overload inhibits the growth and development of wheat. In this work, the impact of Triticum monococcum nicotianamine synthase 3 (TmNAS3) on Fe accumulation in wheat grain was analyzed. Transgenic wheat expressing TmNAS3 was obtained via Agrobacterium-mediated transformation. The concentrations of Fe in the grains of two transgenic wheat lines were 62.42μg/g and 68.75μg/g, while that in the non-transgenic line (NT) was only 29.51μg/g. Exogenous Fe application induced the expression of natural resistance-associated macrophage protein 3 (NRAMP3), NRAMP6, yellow stripe-like protein 3 (YSL3), YSL6, and vacuolar iron transporter 2 in transgenic wheat. The transcription factor that bound to the TmNAS3 promoter was identified, and the findings suggested that TmbHLH47 directly interacted and promoted the transcription of TmNAS3. Moreover, TmbHLH47 was observed to bind directly to the G-box in TmNAS3 promoter and regulated the transcriptional level of TmNAS3. Our findings contribute a TmbHLH47/TmNAS3 transcriptional pathway and thereby provide a potential strategy for improving the Fe concentration of wheat through genetic engineering.

The NtNRAMP1 transporter is involved in cadmium and iron transport in tobacco (Nicotiana tabacum)

Plant natural resistance-associated macrophage protein (NRAMP) plays an important role in maintaining intracellular metal homeostasis and coping with environmental heavy metal stress. Until now, studies on NRAMP in tobacco have been limited. In this study, NtNRAMP1 was cloned from tobacco cultivar TN90, and the highest expression level was observed in the roots, which was strongly induced by Fe deficiency. Heterologously expressed NtNRAMP1 significantly increased the Cd sensitivity of the yeast Δycf1 mutant. Three overexpressed NtNRAMP1 lines were generated to reveal the biofunction of NtNRAMP1. In the soil pot experiments under natural conditions, the contents of Fe and total chlorophyll were increased in the leaves of transgenic tobacco compared with the WT. To reveal the characteristics of NtNRAMP1 in metal transport, transgenic plants were cultured in hydroponic solution with 50 μM Cd and 200 μM Fe. Compared with the WT, the Cd concentrations in transgenic plants increased by 1.26–2.02-fold in the roots. Interestingly, the Cd content in the shoots of transgenic plants was slightly reduced compared with that of the WT. Overexpression of NtNRAMP1 did not promote Fe uptake from the external environment into the roots but enhanced the transfer of Fe from the roots to shoots. Additionally, Fe overload in the leaves of transgenic tobacco resulted in increased levels of MDA and H2O2 while Fe toxicity may be relieved by POD. In conclusion, overexpression of NtNRAMP1 in tobacco could promote Cd uptake and Fe transport from the roots to shoots while disturbing Fe homeostasis in the leaves of transgenic tobacco.

Polymorphism of TLR1, TLR4, and SLC11A1 genes in populations of different cattle breeds of Ukrainian selection

Aim. To study the specificities of population genetic structure of Ukrainian Black-and-White dairy breed, Ukrainian Red-and-White dairy breed, and Ukrainian Grey cattle breed by polymorphism of TLR1, TLR4, and SLC11A1 genes. Methods. The study was conducted using the method of polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP). Results. The results of the study demonstrated that in all the experimental populations, the locus TLR4 by mutations 8732G>A, 8834G>C, and 2021C>T was monomorphic, while loci TLR1 (1596G>A) and SLC11A1 (7400C>G and 7808A>T) were polymorphic. For the TLR4 gene by 8732G>A mutation, only individu- als with genotype BB were found; by 8834G>C – with genotype GG; by 2021C>T – with genotype CC. By BclI- polymorphism in the first exon of TLR1 (1596G>A) the A and G allelic variants were found in each experimental population. The deviation from the Hardy-Weinberg equilibrium state was revealed for the populations of Ukrainian Black-and-White and Ukrainian Grey cattle breeds. A considerable excess of heterozygous individuals was fixed in both cattle populations (31 and 39 % respectively). Considering PstI-polymorphism in exon 11 of SLC11A1 gene, allelic variants C and G were found for SNP5 (7400C>G), as well as A and T for SNP6 (7808A>T) in all experimen- tal populations. As for SNP5 (7400C>G), a considerable excess (from 15 to 30 %) of heterozygous individuals was determined for all experimental groups. Unlike other breeds, there was no excess (Fis = 0,04) of heterozygotes for SNP6 (7808A>T) in the population of Ukrainian Grey cattle. Conclusions. The parameters of genetic variability in the different cattle populations of Ukrainian selection by TLR1, TLR4, and SLC11A1 loci were studied. The results of the study showed the impossibility of using TLR4 locus by 8732G>A, 8834G>C and 2021C>T mutations in the programs of marker-associated selection of the investigated cattle breeds due to its monomorphic nature. The analysis of the allele and genotype distribution by TLR1 and SLC11A1 loci (presence of individuals with different genotypes in all experimental cattle breeds) indicates the possibility of using different allelic variants of these genes in the breeding programs for the studied cattle populations.

Genetic system underlying responses of Cryptococcus neoformans to cadmium.

Cryptococcus neoformans, basidiomycetous pathogenic yeast, is basically an environmental fungus and, therefore, challenged by ever changing environments. In this study, we focused on how C. neoformans responds to stress caused by cadmium that is one of high-risk pollutants. By tracking phenotypes of the resistance or sensitivity to cadmium, we undertook forward and reverse genetic studies to identify genes involved in cadmium metabolism in C. neoformans. We found that the main route of Cd2+ influx is through Mn2+ ion transporter, Smf1, which is an ortholog of Nramp (natural resistance-associated macrophage protein 1) of mouse. We found that serotype A strains are generally more resistant to cadmium than serotype D strains and that cadmium resistance of H99, a representative of serotype A strains, was found to be due to a partial defect in SMF1. We found that calcium channel has a subsidiary role for cadmium uptake. We also showed that Pca1 (P-type-ATPase) functions as an extrusion pump for cadmium. We examined the effects of some metals on cadmium toxicity and suggested (i) that Ca2+ and Zn2+ could exert their protective function against Cd2+ via restoring cadmium-inhibited cellular processes and (ii) that Mg2+ and Mn2+ could have antagonistic roles in an unknown Smf1-independent Cd2+ uptake system. We proposed a model for Cd2+-response of C. neoformans, which will serve as a platform for understanding how this organism copes with the toxic metal.

Effect of breastfeeding on children's health and its relationship to NRAMP1 expression: A cross-sectional study

Toddlers with exclusive breastfeeding can increase immunity in preventing infectious diseases such as Upper Respiratory Tract Infections (ARI). The body's resistance to disease is controlled by Natural Resistance-Associated Macrophage Protein 1 (NRAMP1). NRAMP1 contributes to the pathophysiology of several intercellular infections, including ARI. The purpose of this study was to determine the effect of breastfeeding on children's health and its relationship to NRAMP1 expression. A cross-sectional study was conducted on 124 toddlers in October 2020–June 2021. Toddlers were selected purposively from three Community Health Centers in Central Jakarta. The first group (n = 62) was exclusive breastfeeding and the second group (n = 62) was not exclusive breastfeeding. The characteristics of mothers and toddlers were collected using a structured questionnaire supported by medical record data. Meanwhile, NRAMP1 expression and NRAMP1 protein levels were obtained from the examination of blood samples. Examination of NRAMP1 gene mRNA expression by real-time PCR method and serum NRAMP1 protein levels by ELISA method. Data were analyzed using t-test, ANOVA, and multiple linear regression. The results obtained that the average mRNA expression of NRAMP1 gene and protein levels of NRAMP1 in infants who were not exclusive breastfeeding were 6.88 fold change (FC) and 315.02 pg/ml compared to those who received exclusive breastfeeding of 11.36 FC and 1087.74 pg./ml. Parity, immunization history, exclusive breastfeeding, and frequency of ARI were significantly associated with NRAMP1 gene mRNA expression and NRAMP1 protein levels (P < 0.05). Maternal and under-five age, gender, and nutritional status were not significantly related (P > 0.05). Exclusive breastfeeding was the dominant factor influencing NRAMP1 gene mRNA expression (OR: 4268) and NRAMP1 protein content (OR: 737,362). Antibodies obtained from exclusive breastfeeding in synergy with the NRAMP1 gene form the body's immunity in infants suffering from ARI.

Effect of selected single nucleotide polymorphisms in SLC11A1, ANKRA2, IFNG and PGLYRP1 genes on host susceptibility to Mycobacterium avium subspecies paratuberculosis infection in Indian cattle.

Paratuberculosis (PTB) is a chronic infectious enteritis of ruminants, caused by Mycobacterium avium subspecies paratuberculosis (MAP) that brings huge economic loss to the dairy farmers. The study was conducted to explore the association of selected SNPs in IFNG, SLC11A1, ANKRA2 and PGLYRP1 genes with resistance to PTB disease in Indian cattle population. A case-control resource population was established based on the results of diagnostic tests used for detection of MAP infection status viz. ELISA, Johnin PPD test, faecal microscopy and IS900 blood PCR. The PCR-RFLP method was used for genotyping of SNPs. SNPs rs109453173 in SLC11A1, rs110853455 in IFNG and rs41933863 in ANKRA2 genes were significantly (P<0.05) associated with resistance to MAP infection. For SNP rs109453173, GG genotype and G allele was found to be associated with resistance against MAP infection than CC and CG genotypes and C allele, respectively. For SNP rs110853455, AG genotype was found to be associated with susceptibility to MAP infection than AA and GG genotype. For SNP rs41933863, the AG genotype provided three and six times more resistance against MAP infection than GG and AA genotype. The results of this study are suggestive of SNPs rs109453173, rs110853455 and rs41933863 as potential markers for screening MAP resistant cattle and a breeding programme favouring GG genotype and G allele for rs109453173, AG genotype for rs41933863 and against AG genotype for rs110853455 might confer resistance against MAP infection in Indian cattle. However, investigation of these SNPs in an independent and larger population will warrant the strength of association for resistance against MAP infection in cattle.

Molecular Hydrogen Increases Quantitative and Qualitative Traits of Rice Grain in Field Trials.

How to use environmentally friendly technology to enhance rice field and grain quality is a challenge for the scientific community. Here, we showed that the application of molecular hydrogen in the form of hydrogen nanobubble water could increase the length, width, and thickness of brown/rough rice and white rice, as well as 1000-grain weight, compared to the irrigation with ditch water. The above results were well matched with the transcriptional profiles of representative genes related to high yield, including up-regulation of heterotrimeric G protein β-subunit gene (RGB1) for cellular proliferation, Grain size 5 (GS5) for grain width, Small grain 1 (SMG1) for grain length and width, Grain weight 8 (GW8) for grain width and weight, and down-regulation of negatively correlated gene Grain size 3 (GS3) for grain length. Meanwhile, although total starch content in white rice is not altered by HNW, the content of amylose was decreased by 31.6%, which was parallel to the changes in the transcripts of the amylose metabolism genes. In particular, cadmium accumulation in white rice was significantly reduced, reaching 52% of the control group. This phenomenon was correlated well with the differential expression of transporter genes responsible for Cd entering plants, including down-regulated Natural resistance-associated macrophage protein (Nramp5), Heavy metal transporting ATPase (HMA2 and HMA3), and Iron-regulated transporters (IRT1), and for decreasing Cd accumulation in grain, including down-regulated Low cadmium (LCD). This study clearly showed that the application of molecular hydrogen might be used as an effective approach to increase field and grain quality of rice.

Iron-deficiency response and differential expression of iron homeostasis related genes in spring wheat (

Iron (Fe) is essential for plant growth and human health. Fe deficiency reduces yield and quality traits of wheat (Triticum aestivum L.). Grains of modern bread wheat varieties contain low levels of Fe, and Fe uptake and translocation in wheat grown in Fe conditions have not been studied in detail. This study investigates Fe homeostasis and biofortification in genetically stable spring wheat Almaken and Zhenis M5 mutant lines, developed with 200 Gy for higher grain Fe content. Mutant lines and parents were analysed for the expression of genes involved in Fe homeostasis under normal and deficient Fe. Wheat homologues of genes that participated in phytosiderophore (PS) synthesis and transport were significantly upregulated in the Fe-limited roots of Almaken M/1 and both Zhenis M/2 mutant lines, emphasising the role of deoxymugineic acid (DMA) in iron acquisition. The combined overexpression of SAMS, NAS1, TaNAAT, DMAS and TOM was also revealed in the roots of Almaken M/1 and both Zhenis M5 mutant lines, suggesting their involvement in PS synthesis, Fe chelation and transport. Under Fe deficiency, levels of TaYS1A encoding the wheat homologues of the metal-NA transporter YSL, also showed 2.6-, 5.1- and 5.9-fold increases in the roots of Almaken M/1 and both Zhenis M5 mutant lines, respectively. Vacuolar iron transporters (VIT2), natural resistance associated-macrophage protein (NRAMP) genes and the transcription factor basic-loop-helix (bHLH) were significantly upregulated under Fe starvation in shoots. Fe-deficiency-related genotype-dependent and tissue-specific gene expression differences provide new insights into genes involved in iron homeostasis and biofortification genes in wheat.

Epigenetic regulation of SLC11a1 gene in horses infected with cyathostomins

Cyathostomins are the most prevalent nematode parasites of horses worldwide and the SLC11a1 gene has been found to modulate the host immune response against parasite infections. The objective of this work was to determine the DNA methylation profile of the SLC11a1 gene in naturally infected horses (n = 6) and correlate it with the cyathostomin fecal egg count (FEC). In silico analyzes showed two CpG islands, located in the body of the gene and DNA sequencing showed a statistically significant difference between the methylation profile of islands 1 and 2 (P < 0.0001). The data revealed that island 1 was hypermethylated (>95%), while island 2 was hypomethylated (<10%). Although there were no statistical differences in animals with low or high FEC in relation to methylation of island 1 (P = 0.177) or island 2 (P = 0.231), the analysis of island 2 showed a pattern in which samples with higher FEC also presented higher methylation for that island. This data indicates a possible correlation between the cyathostome FEC and the increased methylation of island 2. Animals with a high methylation rate presented a decrease in FEC after 4 months. The preliminary results indicate for the first time the possibility of epigenetic regulation of the SLC11a1 gene in horses. Moreover, it is suggested that methylation of island 2 of the SLC11a1 cascade might indicate the prognostic of parasite infection. Epigenetic processes such as these might be used as biomarkers for identifying individual animals for target treatments and specific breeding purposes.

Role of Nramp transporter genes of Spirodela polyrhiza in cadmium accumulation

As a pollutant, Cd causes severe impact to the environment and damages living organisms. It can be uptaken from the environment by the natural resistance-associated macrophage protein (Nramp) in plants. However, the ion absorption function of Nramp transporter genes in Spirodela polyrhiza has not been reported. In this study, SpNramp1, SpNramp2, and SpNramp3 from S. polyrhiza were cloned and their functions were analyzed in S. polyrhiza and yeast. Growth parameters and physicochemical indices of wild-type and transgenic lines were measured under Cd stress. Results revealed that SpNramp1, SpNramp2, and SpNramp3 were identified as plasma membrane-localized transporters, and their roles in transporting Cd were verified in yeast. In S. polyrhiza, SpNramp1 overexpression significantly increased the content of Cd, Fe, Mn, and fresh weight. SpNramp2 overexpression increased Mn and Cd. SpNramp3 overexpression increased Fe and Mn concentrations. These results indicate that SpNramp1, SpNramp2, and SpNramp3 had a different preference for ion absorption. Two S. polyrhiza transgenic lines (OE1 and OE3) were obtained. One of them (OE1) showed a stronger accumulation ability, and the other one (OE3) exhibited tolerance capacity to Cd. This study provides new insight into the functions of SpNramp1, SpNramp2, and SpNramp3 and obtains important enrichment lines (OE1) for manipulating Cd accumulation, phytoremediation, and ecological safety.

Identification of NRAMP4 from Arabis paniculata enhance cadmium tolerance in transgenic Arabidopsis

Arabis paniculata has been reported as a hyperaccumulator and functions in cadmium (Cd) tolerance and accumulation. However, the genes involved in Cd stress resistance in A. paniculata are still unknown. In this work, genes of the natural resistanceassociated macrophage proteins (NRAMPs) were characterized in A. paniculata, and their evolutionary relationship and expression patterns were analysed. Expression profiles indicated that ApNRAMPs showed large differences in response to Cd stress. It was highly induced by Cd in root and shoot tissues. To investigate the function of ApNRAMP4 under Cd stress, ApNRAMP4 was cloned and expressed in yeast and Arabidopsis. The results indicated that yeast and Arabidopsis expressing ApNRAMP4 showed normal growth under Cd stress. In addition, transgenic yeast and Arabidopsis showed the ability to concentrate Cd. Under 20 μM CdCl2, Cd concentrations in wild type (WT) and transgenic yeast were 3.11 and 5.92 mg/kg, respectively. Cd concentrations in root tissues of WTand transgenic Arabidopsis were 0.18 and 0.54 mg/kg, respectively. In shoot tissues of WT and transgenic Arabidopsis, Cd concentrations were 0.13 and 0.49 mg/kg, respectively. This report provides genomic information on hyperaccumulator A. paniculata. In addition, the present work identified key NRAMP genes that may serve as resources for heavy metal phytoremediation.