CDF Family Research Articles

CDF family of zinc transporters ZRC-1, MSC-2, and ZRG-17 are involved in survival at high zinc conditions, vegetative development, and cellulase utilization in Neurospora crassa

CDF family of zinc transporters ZRC-1, MSC-2, and ZRG-17 are involved in survival at high zinc conditions, vegetative development, and cellulase utilization in Neurospora crassa

Zinc homeostasis in Schizosaccharomyces pombe.

Most metal ions such as iron, calcium, zinc, or copper are essential for all eukaryotes. Organisms must maintain homeostasis of these metal ions because excess or deficiency of metal ions could cause damage to organisms. The steady state of many metal ions such as iron and copper has been well studied in detail. However, how to regulate zinc homeostasis in Schizosaccharomyces pombe is still confusing. In this review, we provide an overview of the molecular mechanisms that how S. pombe is able to maintain the balance of zinc levels in the changes of environment. In response to high levels of zinc, the transcription factor Loz1 represses the expression of several genes involved in the acquisition of zinc. Meanwhile, the CDF family proteins transport excess zinc to the secretory pathway. When zinc levels are limited, Loz1 was inactivated and could not inhibit the expression of zinc acquisition genes, and zinc stored in the secretory pathway is released for use by the cells. Besides, other factors that regulate zinc homeostasis are also discussed.

Insights Into Histoplasma capsulatum Behavior on Zinc Deprivation.

Histoplasma capsulatum is a thermodimorphic fungus that causes histoplasmosis, a mycosis of global incidence. The disease is prevalent in temperate and tropical regions such as North America, South America, Europe, and Asia. It is known that during infection macrophages restrict Zn availability to H. capsulatum as a microbicidal mechanism. In this way the present work aimed to study the response of H. capsulatum to zinc deprivation. In silico analyses showed that H. capsulatum has eight genes related to zinc homeostasis ranging from transcription factors to CDF and ZIP family transporters. The transcriptional levels of ZAP1, ZRT1, and ZRT2 were induced under zinc-limiting conditions. The decrease in Zn availability increases fungicidal macrophage activity. Proteomics analysis during zinc deprivation at 24 and 48 h showed 265 proteins differentially expressed at 24 h and 68 at 48 h. Proteins related to energy production pathways, oxidative stress, and cell wall remodeling were regulated. The data also suggested that low metal availability increases the chitin and glycan content in fungal cell wall that results in smoother cell surface. Metal restriction also induces oxidative stress triggered, at least in part, by reduction in pyridoxin synthesis.

The photoperiodic response of hypocotyl elongation involves regulation of CDF1 and CDF5 activity.

Hypocotyl elongation relies on directional cell expansion, a process under light and circadian clock control. Under short photoperiods (SD), hypocotyl elongation in Arabidopsis thaliana follows a rhythmic pattern, a process in which circadian morning-to-midnight waves of the transcriptional repressors PSEUDO-RESPONSE REGULATORS (PRRs) jointly gate PHYTOCHROME-INTERACTING FACTOR (PIF) activity to dawn. Previously, we described CYCLING DOF FACTOR 5 (CDF5) as a target of this antagonistic PRR/PIF dynamic interplay. Under SD, PIFs induce CDF5 accumulation specifically at dawn, when it promotes the expression of positive cell elongation regulators such as YUCCA8 to induce growth. In contrast to SD, hypocotyl elongation under long days (LD) is largely reduced. Here, we examine whether CDF5 is an actor in this photoperiod specific regulation. We report that transcription of CDF5 is robustly induced in SD compared to LD, in accordance with PIFs accumulating to higher levels in SD, and in contrast to other members of the CDF family, whose expression is mainly clock regulated and have similar waveforms in SD and LD. Notably, when CDF5 was constitutively expressed under LD, CDF5 protein accumulated to levels comparable to SD but was inactive in promoting cell elongation. Similar results were observed for CDF1. Our findings indicate that both CDFs can promote cell elongation specifically in shorter photoperiods, however, their activity in LD is inhibited at the post-translational level. These data not only expand our understanding of the biological role of CDF transcription factors, but also identify a previously unrecognized regulatory layer in the photoperiodic response of hypocotyl elongation.

Identification of CDF family members in apple and their expression in response to sucrose during floral induction

ABSTRACT Plant-specific CDF genes encode a class of transcription factors which play crucial roles in plant flowering, especially in photoperiod-regulated flowering. But little is known about the potential functions of MdCDFs during flower induction in apple. In this study, 11 putative MdCDFs were identified and named systematically. Bioinformatic analysis revealed a functional and structural similarity among the MdCDFs. Analysis of tandem and synteny revealed several paired CDF homologs between apple and Arabidopsis. RT-qPCR revealed that MdCDFs exhibited distinct expression patterns in different apple tissues and most of them displayed high expression levels in flowers and leaves. Further expression profiles of MdCDFs were investigated in different-flowering varieties of apple and in response to sugar treatment. The results indicated that MdCDFs were responsive to floral induction. Meanwhile, the expressions of all MdCDFs were inhibited in buds during different floral induction stages after sugar treatment. The expressions of flowering-related and photoperiod pathway-related genes showed differences and may indicate that most of the MdCDFs may function as a light response flowering inhibitor and could be repressed by the sugar-mediated floral induction. Collectively, the comprehensive analysis of MdCDFs provides valuable information on their expression during floral induction that will serve as a foundation for further studies.

A mutagenic study identifying critical residues for the structure and function of rice manganese transporter OsMTP8.1.

Rice (Oryza sativa) MTP8.1 (OsMTP8.1) is a tonoplast-localized manganese transporter of the cation diffusion facilitator family. Here we present a structure-function analysis of OsMTP8.1 based on the site-directed and random mutagenesis and complementation assays in manganese hypersensitive yeast, in combination with three-dimensional (3D) structure modeling based on the crystal structure of the Escherichia coli CDF family member, EcYiiP. Two metal-binding sites are conserved in OsMTP8.1 with EcYiiP, one is between transmembrane helices TM2 and TM5, the other is the cytoplasmic C-terminus. In addition to these two metal-binding sites, there may exist other Mn-binding sites such as that at the very end of the CTD. Two residues (R167 and L296) may play an important role for the hinge-like movement of CTDs. Several mutations such as E357A and V374D may affect dimer formation, and S132A may induce a conformational change, resulting in a loss of transport function or modification in metal selectivity. The N-terminus of OsMTP8.1 was not functional for Mn transport activity, and the real function of NTD remains to be investigated in the future. The findings of the present study illustrate the structure-function relationship of OsMTP8.1 in Mn transport activity, which may also be applied to other plant Mn-CDF proteins.

Characterization of a vacuolar zinc transporter OZT1 in rice (Oryza sativa L.)

The CDF family is a ubiquitous family that has been identified in prokaryotes, eukaryotes, and archaea. Members of this family are important heavy metal transporters that transport metal ions out of the cytoplasm. In this research, a full length cDNA named Oryza sativa Zn Transporter 1 (OZT1) that closely related to rat ZnT-2 (Zn Transporter 2) gene was isolated from rice. The OZT1 encoding a CDF family protein shares 28.2%~84.3% of identities and 49.3%~90.9% of similarities with other zinc transporters such as RnZnT-2, HsZnT-8, RnZnT-8 and AtMTP1. OZT1 was constitutively expressed in various rice tissues. The OZT1 expression was significantly induced both in the seedlings of japonica rice Nipponbare and indica rice IR26 in response to Zn(2+) and Cd(2+) treatments. Besides, OZT1 expression was also increased when exposed to other excess metals, such as Cu(2+), Fe(2+) and Mg(2+). Subcellular localization analysis indicated that OZT1 localized to vacuole. Heterologous expression of OZT1 in yeast increased tolerance to Zn(2+) and Cd(2+) stress but not the Mg(2+) stress. Together, OZT1 is a CDF family vacuolar zinc transporter conferring tolerance to Zn(2+) and Cd(2+) stress, which is important to transporting and homeostasis of Zn, Cd or other heavy metals in plants.

Maricaulis maris cation diffusion facilitator: Achieving homogeneity through a mixed-micelle approach

One of the most common problems encountered during isolation and purification of homogenous membrane proteins is their aggregation which in turn makes the obtained material unsuitable for structural and functional studies. As various detergents can have a different impact on the protein stability and solubility, introducing the protein to different detergent micelles can result in the removal of such aggregation. Here we describe a method for retrieving homogenous samples of a putative member of the cation diffusion facilitator family from the marine bacterium Maricaulis maris (MmCDF3). A feature that makes this 23kDa protein particularly interesting to study is that it lacks the cytoplasmic domain that in other members of the CDF family protrudes into the cytoplasm and that was proposed to play a crucial role in the metal transport. The MmCDF3 was produced with the C-terminal hexahistidine tag in Escherichia coli and subsequently purified using affinity chromatography followed by gel-filtration yielding 7.5mg of the pure transporter. However, solubilization and purification of the protein in a single detergent or a complete detergent exchange to another single detergent invariably resulted in the formation of protein aggregates. Instead, if the protein was introduced into a micelle of multiple detergents, the aggregation level notably decreased. Purification of the protein in a mixture of n-dodecyl-β-D-maltoside and Fos-choline-12 at a ratio of 4 to 1 allowed for recovery of 3.7mg of homogenous, non-aggregated MmCDF3 from 1L of bacterial culture that could easily be separated from aggregated material.

Biodegradation of malachite green by strain Pseudomonas sp. K9 and cloning of the tmr2 gene associated with an ISPpu12.

A bacterial strain K9 capable of degrading malachite green was isolated from the sludge of the wastewater treatment system of a chemical plant. It was identified preliminarily as Pseudomonas sp. Strain K9 was also able to degrade other triphenylmethane dyes, such as Crystal Violet and Basic Fuchsin. The gene tmr2, encoding the triphenylmethane reductase, was cloned from strain K9, and functionally expressed in E. coli. A 5946-bp DNA fragment including the tmr2 gene was cloned from the genomic DNA of strain K9 by chromosome walking. Its sequence analysis showed that tmr2 was associated with a typical mobile element ISPpu12 consisting of tnpA (encoding a transposase), lspA (encoding a lipoprotein signal peptidase) and orf1 (encoding a putative MerR family regulator), orf2 (encoding a CDF family heavy metal/H(+) antiporter). This association was also found in another malachite green-degrading strain Pseudomonas sp. MDB-1, which indicated that the tmr2 gene might be a horizontally transferable gene.

Isolation of a methyl parathion-degrading strain Stenotrophomonas sp. SMSP-1 and cloning of the ophc2 gene

A rod-shaped, gram-negative bacterium Stenotrophomonas sp. SMSP-1 was isolated from the sludge of a wastewater treating system of a pesticide manufacturer. Strain SMSP-1 could hydrolyze methyl parathion to p-nitrophenol (PNP) and dimethyl phosphorothioate but could not degrade PNP further. Strain SMSP-1 was able to hydrolyze other organophosphate pesticides, including fenitrothion, ethyl parathion, fenthion, and phoxim, but not chlorpyrifos. A 4395-bp DNA fragment, including an organophosphorus hydrolase encoding gene ophc2, was cloned from the chromosome of strain SMSP-1 using the shotgun technique. Its sequence analysis showed that ophc2 was associated with a typical mobile element ISPpu12 consisting of tnpA (encoding a transposase), lspA (encoding a lipoprotein signal peptidase), and orf1 (encoding a CDF family heavy metal/H(+) antiporter). The ophc2 gene was effectively expressed in E. coli. This is the second report of cloning the ophc2 gene and the first report of this gene from the genus of Stenotrophomonas.

Identification and characterization of MtMTP1, a Zn transporter of CDF family, in the Medicago truncatula

Zn is an essential micronutrient in plants, and the mechanisms of Zn homeostasis are under intensive study. In this report, we have identified MtMTP1, a Zn transporter of the CDF family in the legume model plant Medicago truncatula. The ORF of the MtMTP1 cDNA encodes a protein consisting of 407 amino acid residues with a predicted molecular mass of 45 kDa. Like other metal tolerance proteins (MTPs) in plants, heterologous expression of MtMTP1 can complement the Zn-susceptible zrc1 cot1 yeast double mutant. The expression pattern was studied by quantitative fluorescent PCR. The expression of MtMTP1 was detected in all vegetative organs with the highest level of expression observed in leaves. With Zn supplementation its expression in roots was reduced while its expression in stems was increased in the first 2 days. No obvious changes were detected in leaves. Inoculation with Rhizobium meliloti down-regulated its expression in roots.

Histidine residues in the region between transmembrane domains III and IV of hZip1 are required for zinc transport across the plasma membrane in PC-3 cells

The proteins from the ZIP and the CDF families of zinc transporters contain a histidine-rich sequence in a loop domain located between transmembrane domains III and IV for the ZIP family and transmembrane domains IV and V for the CDF family. Topological predictions suggest that these loops are located in the cytoplasm. The loops contain a histidine-rich sequence with a variable number of histidine residues depending on the transporter. The histidine-rich sequence was postulated to serve as an extra-membrane metal binding site in these proteins. hZip1 is a human zinc transporter ubiquitously expressed. The histidine-rich motif located in the large loop of this transporter is composed of the following sequence, H 158WHD 161. To determine if this motif is involved in the zinc transport activity of the protein, we performed site directed-mutagenesis to replace the loop histidines with alanines. Results suggest that both histidines are necessary for the zinc transport function and are not involved in the plasma membrane localization of the transporter as has been reported for the Zrt1 transporter in yeast. In addition, two histidine residues in transmembrane domains IV and V are also important in the zinc transport function. The results support an intermolecular exchange mechanism of zinc transport.

Seismic data compression based on integer wavelet transform

Due to the particularity of the seismic data, they must be treated by lossless compression algorithm in some cases. In the paper, based on the integer wavelet transform, the lossless compression algorithm is studied. Comparing with the traditional algorithm, it can better improve the compression rate. CDF (2, n) biorthogonal wavelet family can lead to better compression ratio than other CDF family, SWE and CRF, which is owe to its capability in canceling data redundancies and focusing data characteristics. CDF (2, n) family is suitable as the wavelet function of the lossless compression seismic data.

Zeroing in on zinc uptake in yeast and plants.

Zinc is an essential micronutrient. Genes responsible for zinc uptake have now been identified from yeast and plants. These genes belong to an extended family of cation transporters called the ZIP gene family. Zinc efflux genes that belong to another transporter family, the CDF family, have also been identified in yeast and Arabidopsis. It is clear that studies in yeast can greatly aid our understanding of zinc metabolism in plants.