Characterization of Two Metal Transporters HMA3 and NRAMP1 in Two Ecotypes of the Zinc/cadmium Hyperaccumulator Thlaspi Caerulescens Compared with Arabidopsis Thaliana PDF Download
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Author: Maria Clemencia Zambrano Mendoza
Publisher:
ISBN:
Category :
Languages : en
Pages : 498
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Book Description
Accumulation of a given metal in plants depends on a delicate and precise balance of various biological processes. Some plants have developed strategies that allow them to tolerate heavy metals in extreme conditions without suffering toxicity. This research focuses on the characterization of two metal transporters, a member of the P 1B -type (ATPase) transporter family (HMA3) and a member the Natural Resistance Associated Macrophage Protein (NRAMP) Nramp1 family. These transporters have proposed roles in ion homeostasis and mineral nutrition. The work here sought to determine if these transporters might have characteristics that suggest a role in heavy metal transport and tolerance in metal hyperaccumulating plants. These proteins are very well conserved among different taxa. Nonetheless, as little as a single amino acid change has the potential to modify their capacity to take up non essential metals such as Cd, or Pb, and/or increase affinity for other mineral nutrients. These transporters were cloned from a non-accumulator ( Arabidopsis thaliana L) and two ecotypes (Prayon and Ganges) of the hyperaccumulator Noccaea caerulescens (formerly= Thlaspi caerulescens). The full cDNA of an ortholog of either Nramp1 or HMA3 was expressed in yeast in order to provide a heterologous model to elucidate how polymorphisms between the orthologs might translate into functional differences between the protein sequences. A comparison of the HMA3 sequences to each other, or the Nramp1 sequences to each other, demonstrated that major motifs and domains in each protein were highly conserved but that there were numerous single amino acid polymorphisms. Few of these polymorphisms corresponded to positions in a protein that are known to be critical for transporter function. However, metal accumulation, tolerance and cell growth assays showed that the Nramp1 and HMA3 genes from Arabidopsis encoded proteins with the expected broad selectivity for divalent ion transport. In contrast, the genes from the Thlaspi ecotypes encoded proteins that showed more selectivity for ion transport. The Thlaspi ecotypes showed high selectivity for cadmium but the accumulation of other elements differed between the Thlaspi orthologs. These results suggest that the polymorphisms present in the Thlaspi sequences have produced differences in the transport characteristics of both the HMA3 and the Nramp1 transporters.
Author: Maria Clemencia Zambrano Mendoza
Publisher:
ISBN:
Category :
Languages : en
Pages : 498
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Book Description
Accumulation of a given metal in plants depends on a delicate and precise balance of various biological processes. Some plants have developed strategies that allow them to tolerate heavy metals in extreme conditions without suffering toxicity. This research focuses on the characterization of two metal transporters, a member of the P 1B -type (ATPase) transporter family (HMA3) and a member the Natural Resistance Associated Macrophage Protein (NRAMP) Nramp1 family. These transporters have proposed roles in ion homeostasis and mineral nutrition. The work here sought to determine if these transporters might have characteristics that suggest a role in heavy metal transport and tolerance in metal hyperaccumulating plants. These proteins are very well conserved among different taxa. Nonetheless, as little as a single amino acid change has the potential to modify their capacity to take up non essential metals such as Cd, or Pb, and/or increase affinity for other mineral nutrients. These transporters were cloned from a non-accumulator ( Arabidopsis thaliana L) and two ecotypes (Prayon and Ganges) of the hyperaccumulator Noccaea caerulescens (formerly= Thlaspi caerulescens). The full cDNA of an ortholog of either Nramp1 or HMA3 was expressed in yeast in order to provide a heterologous model to elucidate how polymorphisms between the orthologs might translate into functional differences between the protein sequences. A comparison of the HMA3 sequences to each other, or the Nramp1 sequences to each other, demonstrated that major motifs and domains in each protein were highly conserved but that there were numerous single amino acid polymorphisms. Few of these polymorphisms corresponded to positions in a protein that are known to be critical for transporter function. However, metal accumulation, tolerance and cell growth assays showed that the Nramp1 and HMA3 genes from Arabidopsis encoded proteins with the expected broad selectivity for divalent ion transport. In contrast, the genes from the Thlaspi ecotypes encoded proteins that showed more selectivity for ion transport. The Thlaspi ecotypes showed high selectivity for cadmium but the accumulation of other elements differed between the Thlaspi orthologs. These results suggest that the polymorphisms present in the Thlaspi sequences have produced differences in the transport characteristics of both the HMA3 and the Nramp1 transporters.
Author: Maria Clemencia Zambrano Mendoza
Publisher:
ISBN:
Category :
Languages : en
Pages : 267
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Book Description
Accumulation of a given metal in plants depends on a delicate and precise balance of various biological processes. Some plants have developed strategies that allow them to tolerate heavy metals in extreme conditions without suffering toxicity. This research focuses on the characterization of two metal transporters, a member of the P 1B -type (ATPase) transporter family (HMA3) and a member the Natural Resistance Associated Macrophage Protein (NRAMP) Nramp1 family. These transporters have proposed roles in ion homeostasis and mineral nutrition. The work here sought to determine if these transporters might have characteristics that suggest a role in heavy metal transport and tolerance in metal hyperaccumulating plants. These proteins are very well conserved among different taxa. Nonetheless, as little as a single amino acid change has the potential to modify their capacity to take up non essential metals such as Cd, or Pb, and/or increase affinity for other mineral nutrients. These transporters were cloned from a non-accumulator ( Arabidopsis thaliana L) and two ecotypes (Prayon and Ganges) of the hyperaccumulator Noccaea caerulescens (formerly= Thlaspi caerulescens). The full cDNA of an ortholog of either Nramp1 or HMA3 was expressed in yeast in order to provide a heterologous model to elucidate how polymorphisms between the orthologs might translate into functional differences between the protein sequences. A comparison of the HMA3 sequences to each other, or the Nramp1 sequences to each other, demonstrated that major motifs and domains in each protein were highly conserved but that there were numerous single amino acid polymorphisms. Few of these polymorphisms corresponded to positions in a protein that are known to be critical for transporter function. However, metal accumulation, tolerance and cell growth assays showed that the Nramp1 and HMA3 genes from Arabidopsis encoded proteins with the expected broad selectivity for divalent ion transport. In contrast, the genes from the Thlaspi ecotypes encoded proteins that showed more selectivity for ion transport. The Thlaspi ecotypes showed high selectivity for cadmium but the accumulation of other elements differed between the Thlaspi orthologs. These results suggest that the polymorphisms present in the Thlaspi sequences have produced differences in the transport characteristics of both the HMA3 and the Nramp1 transporters.
Author: Matthew J Milner
Publisher:
ISBN:
Category :
Languages : en
Pages : 0
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Thlaspi caerulescens (J&C Presl) is a Zn/Cd hyperaccumulator which tolerates Zn and Cd toxic soil environments and accumulates both metals in the shoot to extremely high levels without any toxicity symptoms (30,000 ppm Zn and 12,000 ppm Cd on a leaf dry weight basis). This ability to hyperaccumulate heavy metals has intrigued plant biologists for many years for the possible value T. caerulescens may have in the phytoremediation of Zn- and Cd-contaminated soils. However, it's slow growth and diminished shoot biomass limits its usefulness for phytoremediation. Exploiting the genetic potential of this species by transferring metal hyperaccumulating traits to plants with a higher biomass may be an effective approach to generate new and novel metal accumulator plants. To accomplish this goal, a better understanding of the basic molecular and physiological mechanisms responsible for Zn/Cd hyperaccumulation is needed. Also, because we had previously shown that Zn hyperaccumulation in T. caerulescens is related to the elevated or altered Zn-dependent expression of a number of metal transport and metal-related genes, this plant may serve as a useful tool for studying plant mechanisms of Zn sensing and homeostasis. In an attempt to better understand how the transport of Zn and Cd from the soil to the shoot is altered in metal hyperaccumulators, T. caerulescens was compared in this study to two closely related nonaccumulator plants, Thlaspi arvense and Arabidopsis thaliana. Additionally ecotypic variation within T. caerulescens was exploited. These two approaches were used to identify key proteins involved in Zn and Cd uptake, regulation and sequestration. One component of the research was the functional characterization of the micronutrient transport TcZNT1, which had previously been suggested to be the root Zn/Cd uptake transporter in T. caerulescens. TcZNT1 was characterized in comparison to its closest sequence-based homolog in Arabidopsis, AtZIP4. From this research we obtained findings that TcZNT1 most likely is involved in root Zn uptake from the soil, but also found a possible new role for this transporter in long distance Zn transport based on high levels of TcZNT1 gene expression in the root and leaf vasculature. While it was found that AtZIP4 shares a number of similarities to TcZNT1 with regards to metal transport and tissue-specific expression, we identified a major difference in that AtZIP4 is localized to the chloroplast while TcZNT1 is a plasma membrane transporter. Hence AtZIP4 must play a role in chloroplast micronutrient homeostasis. A second component of the research focused on potential transcriptional regulators of T. caerulescens Zn transporter genes. Using yeast complementation assays, we identified intriguing candidates for this role based on the ability of T. caerulescens transcription factors to activate the expression of a yeast high affinity Zn uptake transporter. These proteins are members of the E2F family of transcription factors that are thought to be involved solely in cell cycle regulation. Here it was shown that one of these TcE2Fs binds with high affinity to a putative E2F element in the TcZNT1 promoter. TcZNT1 is closely related in sequence to the yeast Zn transporter gene that was activated by the TcE2F. However, to date, we have not shown direct activation of TcZNT1 expression by TcE2F2. The final component of this research project exploited natural ecotypic variation in Cd hyperaccumulation between two T. caerulescens ecotypes, and used a comparative transcriptomics approach between these ecotypes to identify a candidate transporter for the enhanced Cd accumulation in the one ecotype. This transporter, TcHMA3, is a member of the P-type ATPase family of metal transporters and was found to be a vacuolar Cd transporter expressed in both the roots and shoots of T. caerulescens. Overexpression of this vacuolar transporter in transgenic Arabidopsis led to increased Cd tolerance and increased root Cd accumulation.
Author: Melinda Anne Klein
Publisher:
ISBN:
Category :
Languages : en
Pages : 308
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Author:
Publisher:
ISBN:
Category :
Languages : en
Pages : 5
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Book Description
Metal ions are critical nutrients, yet overaccumulation of these same metals can also be toxic. To maintain appropriate intracellular levels, cells require specific metal uptake systems that are subject to precise homeostatic regulation. The long-range goal of our research is to define the molecular mechanism(s) and regulation of metal ion uptake in eukaryotic cells. Integrating genetic, molecular biological and biochemical approaches, we have examined these processes in the yeast Saccharomyces cerevisiae and the plant Arabidopsis thaliana. Both are proven model systems for studying fundamental cellular processes. Our work has focused on the ZIP family of metal transporters which we identified; this family has representatives in bacteria, fungi, plants and animals. IRT1, one of the founding members of the ZIP family, is an essential cation transporter that is expressed in the epidermal cells of iron deficient plant roots and is responsible for uptake of iron from the soil. We now know that t here are 15 ZIP genes in the Arabidopsis genome which can be divided into four different classes, based on their intron/exon arrangements and the similarities among their encoded gene products. The ZIP family members display different substrate specificities for metals and different tissue distributions in Arabidopsis. Moreover, the family members respond differentially to metal deficiencies. For example, IRT1, ZIP6 and ZIP9 mRNA are expressed mainly in the roots of iron deficient plants whereas ZIP4 responds to both iron and zinc deficiency. Work in both yeast and Arabidopsis has addressed substrate specificity as well as how these transporters are regulated in response to metal availability. Our project was broken down into four specific aims. Significant progress was made on all four aims. I have listed the publications which have resulted under the relevant specific aim.
Author:
Publisher: Academic Press
ISBN: 0123946107
Category : Science
Languages : en
Pages : 477
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Book Description
This volume of Current Topics in Membranes focuses on metal transmembrane transporters and pumps, a recently discovered family of membrane proteins with many important roles in the physiology of living organisms. The book summarizes the most recent advances in the field of metal ion transport and provides a broad overview of the major classes of transporters involved in homeostasis of heavy metals. Various families of the transporters and metal specificities are discussed with the focus on the structural and mechanistic aspects of their function and regulation. The reader will access information obtained through a variety of approaches ranging from X-ray crystallography to cell biology and bioinformatics, which have been applied to transporters identified in diverse biological systems, such as pathogenic bacteria, plants, humans and others. Field is cutting-edge and a lot of the information is new to research community Wide breadth of topic coverage Contributors of high renown and expertise
Author: Aryadeep Roychoudhury
Publisher: Academic Press
ISBN: 0128179562
Category : Science
Languages : en
Pages : 272
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Book Description
Metal and Nutrient Transporters in Abiotic Stress focuses on the different forms of environmental stress related to heavy metal, metalloid and nutrient deficiency that have the potential to inflict major damages to crop plants, leading to a massive decrease in crop yield and productivity. The book presents the current state of knowledge of the biochemical and molecular regulation of several classes of membrane transporters related to the uptake of metals/metalloids and nutrient elements during different stresses and their probable mechanisms of operation in plant stress tolerance. Metal and Nutrient Transporters in Abiotic Stress provides a comprehensive discussion that will help in mitigating multiple forms of stresses utilizing transporter proteins. Edited by leading experts and written by a global team of knowledgeable contributors, this book will further stimulate research in the field of transporter proteins and will foster further interests for researchers, academicians and scientists worldwide. It is complimented by its companion book titled Transporters and Plant Osmotic Stress. Focuses exclusively on metal and nutrient transporters involved in multiple environmental stresses in plants Explains exploiting transporters in crop improvement programs through transgenic technology against different stresses such as heavy metal, metalloid and nutrient deficiency Serves as an important source of information in the field of abiotic stress
Author:
Publisher:
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Category :
Languages : en
Pages : 5
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Soils at many DOE sites are contaminated with metals and radionuclides. Such soils obviously pose a risk to human and animal health. Unlike organic wastes, which can be metabolized, metals are immutable and cannot be degraded into harmless constituents. Phytoremediation, the use of plants to remove toxic materials from soil and water, may prove to be an environmentally friendly and cost effective solution for cleaning up metal contaminated sites. The success of phytoremediation will rely on the availability of plants that absorb, translocate, and tolerate the contaminating metals. However, before we can engineer such plants, we need more basic information on how plants acquire metals. An important long term goal of our research program is to understand how metals such as zinc, cadmium and iron are transported across membranes. Our research is focused on a new family of metal transporters, which we have identified through combined studies in the yeast Saccharomyces cerevisiae and in the model plant Arabidopsis thaliana. We have identified a family of 24 presumptive metal transport genes in a variety of organisms including yeast, trypanosomes, plants, nematodes, and humans. This family, which we have designated the ''ZIP'' genes, provides a rich source of material with which to undertake studies on metal transport in eukar.
Author:
Publisher:
ISBN:
Category :
Languages : en
Pages : 3
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Book Description
Soils at many DOE sites are contaminated with metals and radionuclides. Such soils obviously pose a risk to human and animal health. Unlike organic wastes which can be metabolized, metals are immutable and cannot be degraded into harmless constituents. Phytoremediation, the use of plants to remove toxic materials from soil and water, may prove to be an environmentally friendly and cost effective solution for cleaning up metal-contaminated sites. The success of phytoremediation will rely on the availability of plants that absorb, translocate, and tolerate the contaminating metals. However, before the authors can engineer such plants, they need more basic information on how plants acquire metals. An important long term goal of the research program is to understand how metals such as zinc, cadmium and copper are transported across membranes. The research is focused on a new family of metal transporters which they have identified through combined studies in the yeast Saccharomyces cerevisiae and in the model plant Arabidopsis thaliana. They have identified a family of 19 presumptive metal transport genes in a variety of organisms including yeast, trypanosomes, plants, nematodes, and humans. This family, which the authors have designated the ZIP genes, provides a rich source of material with which to undertake studies on metal transport in eukaryotes. The project has three main objectives: Objective 1: Determine the sub-cellular location of the ZIP proteins in Arabidopsis. Objective 2: Carry out a structure/function analysis of the proteins encoded by the ZIP gene family to identify regions of the protein responsible for substrate specificity and affinity. Objective 3: Engineer plants to overexpress and underexpress members of the ZIP gene family and analyze these transgenic plants for alterations in metal accumulation. They now know that manipulation of transporter levels will also require an understanding of post-transcriptional control of ZIP gene expression. They are currently in year one of a three-year project.
Author: Nicole Suzann Pence
Publisher:
ISBN: 9780493502854
Category :
Languages : en
Pages : 220
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Book Description
In summary, our results suggest an alteration in the molecular regulation of a number of genes involved in heavy metal transport and metabolism. A Zn-dependent trans-acting factor and Zn responsive cis elements may result in upregulation of this suite of genes, which in turn causes the dramatic metal hyperaccumulation phenotype in Thlaspi caerulescens.
