COPT2, a plasma membrane located copper transporter, is involved in the uptake of Au in Arabidopsis.
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ABSTRACT:
SUBMITTER: Tiwari M
PROVIDER: S-EPMC5595958 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Members of the aquaporin (AQP) family have been suggested to transport aluminum (Al) in plants; however, the Al form transported by AQPs and the roles of AQPs in Al tolerance remain elusive. Here we report that NIP1;2, a plasma membrane-localized member of the Arabidopsis nodulin 26-like intrinsic protein (NIP) subfamily of the AQP family, facilitates Al-malate transport from the root cell wall into the root symplasm, with subsequent Al xylem loading and root-to-shoot translocation, which are critical steps in an internal Al tolerance mechanism in Arabidopsis We found that NIP1;2 transcripts are expressed mainly in the root tips, and that this expression is enhanced by Al but not by other metal stresses. Mutations in NIP1;2 lead to hyperaccumulation of toxic Al3+ in the root cell wall, inhibition of root-to-shoot Al translocation, and a significant reduction in Al tolerance. NIP1;2 facilitates the transport of Al-malate, but not Al3+ ions, in both yeast and Arabidopsis We demonstrate that the formation of the Al-malate complex in the root tip apoplast is a prerequisite for NIP1;2-mediated Al removal from the root cell wall, and that this requires a functional root malate exudation system mediated by the Al-activated malate transporter, ALMT1. Taken together, these findings reveal a critical linkage between the previously identified Al exclusion mechanism based on root malate release and an internal Al tolerance mechanism identified here through the coordinated function of NIP1;2 and ALMT1, which is required for Al removal from the root cell wall, root-to-shoot Al translocation, and overall Al tolerance in Arabidopsis.
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Project description:The dopamine transporter (DAT) is essential for the reuptake of the released neurotransmitter dopamine (DA) in the brain. Psychostimulants, methamphetamine and cocaine, have been reported to induce the formation of DAT multimeric complexes in vivo and in vitro. The interpretation of DAT multimer function has been primarily in the context of compounds that induce structural and functional modifications of the DAT, complicating the understanding of the significance of DAT multimers. To examine multimerization in the absence of DAT ligands as well as in their presence, we developed a novel, optogenetic fusion chimera of cryptochrome 2 and DAT with an mCherry fluorescent reporter (Cry2-DAT). Using blue light to induce Cry2-DAT multimeric protein complex formation, we were able to simultaneously test the functional contributions of DAT multimerization in the absence or presence of substrates or inhibitors with high spatiotemporal precision. We found that blue light-stimulated Cry2-DAT multimers significantly increased IDT307 uptake and MFZ 9-18 binding in the absence of ligands as well as after methamphetamine and nomifensine treatment. Blue light-induced Cry2-DAT multimerization increased colocalization with recycling endosomal marker Rab11 and had decreased presence in Rab5-positive early endosomes and Rab7-positive late endosomes. Our data suggest that the increased uptake and binding results from induced and rapid trafficking of DAT multimers to the plasma membrane. Our data suggest that DAT multimers may function to help maintain DA homeostasis.
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Project description:Transporters belonging to the natural resistance-associated macrophage protein (Nramp) family play important roles in metal uptake and homeostasis. Although Nramp members have been functionally characterized in plants, the role of Nramp in the important tropical forage legume Stylosanthes guianensis (stylo) is largely unknown. This study aimed to determine the responses of Nramp genes to metal stresses and investigate its metal transport activity in stylo. Five SgNramp genes were identified from stylo. Expression analysis showed that SgNramp genes exhibited tissue preferential expressions and diverse responses to metal stresses, especially for manganese (Mn), suggesting the involvement of SgNramps in the response of stylo to metal stresses. Of the five SgNramps, SgNramp1 displayed the highest expression in stylo roots. A close correlation between SgNramp1 expression and root Mn concentration was observed among nine stylo cultivars under Mn limited condition. The higher expression of SgNramp1 was correlated with a high Mn uptake in stylo. Subsequent subcellular localization analysis showed that SgNramp1 was localized to the plasma membrane. Furthermore, heterologous expression of SgNramp1 complemented the phenotype of the Mn uptake-defective yeast (Saccharomyces cerevisiae) mutant Δsmf1. Mn concentration in the yeast cells expressing SgNramp1 was higher than that of the empty vector control, suggesting the transport activity of SgNramp1 for Mn in yeast. Taken together, this study reveals that SgNramp1 is a plasma membrane-localized transporter responsible for Mn uptake in stylo.
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Project description:The serotonin (5-HT) uptake system is supposed to play a crucial part in vascular functions by "fine-tuning" the local concentration of 5-HT in the vicinity of 5-HT(2) receptors in vascular smooth muscle cells. In this study, the mechanism of 5-HT uptake in human brain vascular smooth muscle cells (HBVSMCs) was investigated. [(3)H]5-HT uptake in HBVSMCs was Na(+)-independent. Kinetic analyses of [(3)H]5-HT uptake in HBVSMCs revealed a K(m) of 50.36 ± 10.2 mM and a V(max) of 1033.61 ± 98.86 pmol/mg protein/min. The specific serotonin re-uptake transporter (SERT) inhibitor citalopram, the specific norepinephrine transporter (NET) inhibitor desipramine, and the dopamine transporter (DAT) inhibitor GBR12935 inhibited 5-HT uptake in HBVSMCs with IC(50) values of 97.03 ± 40.10, 10.49 ± 5.98, and 2.80 ± 1.04 μM, respectively. These IC(50) values were 100-fold higher than data reported by other authors, suggesting that those inhibitors were not blocking their corresponding transporters. Reverse transcription-polymerase chain reaction results demonstrated the presence of mRNA for organic cation transporter (OCT)-3 and plasma membrane monoamine transporter (PMAT), but the absence of OCT-1, OCT-2, SERT, NET, and DAT. siRNA knockdown of OCT-3 and PMAT specifically attenuated 5-HT uptake in HBVSMCs. It is concluded that 5-HT uptake in HBVSMCs was mediated predominantly by a low-affinity and Na(+)-independent mechanism. The most probable candidates are OCT-3 and PMAT, but not the SERT.
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Project description:Copper is an essential micronutrient. Following entry via the human copper transporter 1 (hCTR1), copper is delivered to several copper chaperones, which subsequently transfer the metal to specific targets via protein:protein interactions. It is has been assumed, but not demonstrated, that chaperones acquire copper directly from hCTR1. However, some reports have pointed to an intermediary role for glutathione (GSH), an abundant copper-binding tri-peptide. To address the issue of how transported copper is acquired by the copper chaperones in vivo, we measured the initial rate of (64)Cu uptake in cells in which the cellular levels of copper chaperones or GSH were substantially depleted or elevated. Knockdown or overexpression of copper chaperones ATOX1, CCS, or both had no effect on the initial rate of (64)Cu entry into HEK293 cells having endogenous or overexpressed hCTR1. In contrast, depleting cellular GSH using L-buthionine-sulfoximine (BSO) caused a 50% decrease in the initial rate of (64)Cu entry in HEK293 cells and other cell types. This decrease was reversed by washout of BSO or GSH replenishment with a permeable ester. BSO treatment under our experimental conditions had no significant effects on the viability, ATP levels, or metal content of the cells. Attenuated (64)Cu uptake in BSO was not due to oxidation of the cysteine in the putative metal-binding motif (HCH) at the intracellular hCTR1 COOH terminus, because a mutant lacking this motif was fully active, and (64)Cu uptake was still reduced by BSO treatment. Our data suggest that GSH plays an important role in copper handling at the entry step.
| S-EPMC3625801 | biostudies-literature
Project description:The TRPM7 (transient receptor potential melastatin 7) ion channel has been implicated in the uptake of Mg2+ into vertebrate cells, as elimination of TRPM7 expression through gene targeting in DT40 B-lymphocytes renders them unable to grow in the absence of supplemental Mg2+. However, a residual capacity of TRPM7-deficient cells to accumulate Mg2+ and proliferate when provided with supplemental Mg2+ suggests the existence of Mg2+ uptake mechanism(s) other than TRPM7. Evaluation of the expression of several members of the SLC41 (solute carrier family 41) family, which exhibit homology with the MgtE class of prokaryotic putative bivalent-cation transporters, demonstrated that one, SLC41A2 (solute carrier family 41 member 2), is expressed in both wild-type and TRPM7-deficient DT40 cells. Characterization of heterologously expressed SLC41A2 protein indicated that it is a plasma-membrane protein with an N-terminus-outside/C-terminus-inside 11-TM (transmembrane)-span topology, consistent with its functioning as a trans-plasma-membrane transporter. In contrast with a previous report of ion-channel activity associated with SLC41A2 expression in oocytes, investigation of whole cell currents in SLC41A2-expressing DT40 cells revealed no novel currents of any type associated with SLC41A2 expression. However, expression of SLC41A2 in TRPM7-deficient cells under the control of a doxycycline-inducible promoter was able to conditionally enhance their net uptake of 26Mg2+ and conditionally and dose-dependently provide them with the capacity to grow in the absence of supplemental Mg2+, observations strongly supporting a model whereby SLC41A2 directly mediates trans-plasma-membrane Mg2+ transport. Overall, our results suggest that SLC41A2 functions as a plasma-membrane Mg2+ transporter in vertebrate cells.
| S-EPMC1820800 | biostudies-literature