Phenotyping of single plant cells on a microfluidic cytometry platform with fluorescent, mechanical, and electrical modules.

Compared to animal cells, phenotypic characterization of single plant cells on microfluidic platforms is still rare. In this work, we collated population statistics on the morphological, biochemical, physical and electrical properties of Arabidopsis protoplasts under different external and internal conditions, using progressively improved microfluidic platforms. First, we analyzed the different effects of three phytohormones (auxin, cytokinin and gibberellin) on the primary cell wall (PCW) regeneration process using a microfluidic flow cytometry platform equipped with a single-channel fluorescence sensor. Second, we correlated the intracellular reactive oxygen species (ROS) level induced by heavy metal stress with the concurrent PCW regeneration process by using a dual-channel fluorescence sensor. Third, by integrating contraction channels, we were able to effectively discriminate variations in cell size while monitoring the intensity of intracellular ROS signaling. Fourth, by combining an electrical impedance electrode with the contraction channel, we analyzed the differences in electrical and mechanical properties of wild-type and mutant plant cells before and after primary cell wall regeneration. Overall, our work demonstrates the feasibility and sensitivity of microfluidic flow cytometry in high-throughput phenotyping of plant cells and provides a reference for assessing metabolic and physiological indicators of individual plant cells in multiple dimensions.

Association between MTHFR C677T Gene Polymorphisms and the Efficacy of Vitamin Therapy in lowering Homocysteine Levels among Stroke Patients with Hyperhomocysteinemia.

BACKGROUND: The impact of the methylenetetrahydrofolate reductase (MTHFR) C677T mutation on the relationship between plasma homocysteine (Hcy) levels and stroke has been extensively studied and documented in previous study. However, it remains unclear whether the MTHFR C677T mutation can affect the response to Hcy lowering treatment in stroke patients with hyperhomocysteinemia (HHcy). Understanding the impact of genetic factors on treatment response can help optimize personalized treatment strategies for stroke patients with HHcy. We aimed to investigate the potential association between the MTHFR C677T gene polymorphisms and the effectiveness of Hcy lowering treatment using vitamin therapy in stroke patients with HHcy. METHODS: The MTHFR C677T genotype polymorphisms were identified using polymerase chain reaction-restriction fragment length polymorphism, and the distribution of three genotypes in the MTHFR C677T gene locus was compared. The treatment effects of Hcy lowering agents were compared among patients with different genotypes. RESULTS: Among the 320 stroke patients enrolled in the study, 258 (80.6%) were diagnosed with HHcy. Of these, 162 patients (Effective Group) responded well to the clinical Hcy lowering treatment, while 96 patients (Invalid Group) failed to achieve sufficient response even after taking combination supplements of folic acid, Vitamin B6, and methylcobalamin for one month. Significant differences were observed in terms of age (p < 0.001), hypertension (p = 0.034), dyslipidemia (p = 0.022), hyperuricemia (p = 0.013) and genotype distribution of MTHFR C677T gene polymorphism (p < 0.001) between the Invalid group and the Effective group. The multivariate regression analysis revealed that the T allele (odd rations [OR], 1.327; 95% confidence interval [CI], 1.114-1.580; p = 0.0015) was independently associated with an insufficient Hcy lowering treatment effect. Additionally, the TT genotype was independently associated with insufficient response in both the codominant model (OR, 1.645; 95% CI, 1.093-2.476; p = 0.017) and the recessive model (TT versus CC + CT; OR, 1.529; 95% CI, 1.145-2.042; p = 0.004). However, no relationship was observed between CT + TT genotypes and poor treatment effect in the dominate model. CONCLUSIONS: Our findings suggested that the TT genotype and T allele of MTHFR C677T polymorphism were independently associated with an insufficient Hcy lowering treatment effect in stroke patients with HHcy.

Variations of sediment organic phosphorus and organic carbon during the outbreak and decline of algal blooms in Lake Taihu, China.

In this study, sediment organic phosphorus (OP) and organic carbon (OC) in Lake Taihu, China, as well as their relationships, were analyzed during the outbreak and decline of algal blooms (ABs) over a five-month field study. The results showed synchronous temporal changes in the sediment OP and OC contents with the development of ABs. In addition, there was a significant positive correlation between the sediment OP and OC (p < 0.01), suggesting simultaneous deposition and consumption during the ABs outbreak. The sediment OP and OC contents decreased significantly at the early and last stages of the ABs outbreak and increased at the peak of the ABs outbreak and during the ABs decline. These temporal variation patterns suggest that the sediment OC and OP contents did not consistently increase during the ABs outbreak, even though algae are an important source of organic matter in sediments. The depletion or enrichment of OC and OP in sediments may also depend on the scale of the ABs outbreak. The obtained results revealed significant differences in the sediment OC and OP contents between the months (p < 0.05). In addition, OP in the sediments was dominated by orthophosphate diester (phospholipids and DNA-P) and orthophosphate monoester during the ABs outbreak and decline, respectively. The active OC contents and proportions in the sediments in the ABs outbreak were significantly lower than those observed in the ABs decline period, demonstrating the significant impacts of the ABs outbreak and decline on the sediment OC and OP in Lake Taihu.

Arabidopsis seedlings respond differentially to nutrient efficacy of three rock meals by regulating root architecture and endogenous auxin homeostasis.

BACKGROUND: Plants show developmental plasticity with variations in environmental nutrients. Considering low-cost rock dust has been identified as a potential alternative to artificial fertilizers for more sustainable agriculture, the growth responses of Arabidopsis seedlings on three rock meals (basalt, granite, and marlstone) were examined for the different foraging behavior, biomass accumulation, and root architecture. RESULTS: Compared to ½ MS medium, basalt and granite meal increased primary root length by 13% and 38%, respectively, but marlstone caused a 66% decrease, and they all drastically reduced initiation and elongation of lateral roots but lengthened root hairs. Simultaneous supply of organic nutrients and trace elements increased fresh weight due to the increased length of primary roots and root hairs. When nitrogen (N), phosphorus (P), and potassium (K) were supplied individually, N proved most effective in improving fresh weight of seedlings growing on basalt and granite, whereas K, followed by P, was most effective for those growing on marlstone. Unexpectedly, the addition of N to marlstone negatively affected seedling growth, which was associated with repressed auxin biosynthesis in roots. CONCLUSIONS: Our data indicate that plants can recognize and adapt to complex mineral deficiency by adjusting hormonal homeostasis to achieve environmental sensitivity and developmental plasticity, which provide a basis for ecologically sound and sustainable strategies to maximize the use of natural resources and reduce the production of artificial fertilizers.

Mitochondrial Genome Sequence of Salvia officinalis (Lamiales: Lamiaceae) Suggests Diverse Genome Structures in Cogeneric Species and Finds the Stop Gain of Genes through RNA Editing Events.

Our previous study was the first to confirm that the predominant conformation of mitochondrial genome (mitogenome) sequence of Salvia species contains two circular chromosomes. To further understand the organization, variation, and evolution of Salvia mitogenomes, we characterized the mitogenome of Salvia officinalis. The mitogenome of S. officinalis was sequenced using Illumina short reads and Nanopore long reads and assembled using a hybrid assembly strategy. We found that the predominant conformation of the S. officinalis mitogenome also had two circular chromosomes that were 268,341 bp (MC1) and 39,827 bp (MC2) in length. The S. officinalis mitogenome encoded an angiosperm-typical set of 24 core genes, 9 variable genes, 3 rRNA genes, and 16 tRNA genes. We found many rearrangements of the Salvia mitogenome through inter- and intra-specific comparisons. A phylogenetic analysis of the coding sequences (CDs) of 26 common protein-coding genes (PCGs) of 11 Lamiales species and 2 outgroup taxa strongly indicated that the S. officinalis was a sister taxon to S. miltiorrhiza, consistent with the results obtained using concatenated CDs of common plastid genes. The mapping of RNA-seq data to the CDs of PCGs led to the identification of 451 C-to-U RNA editing sites from 31 PCGs of the S. officinalis mitogenome. Using PCR amplification and Sanger sequencing methods, we successfully validated 113 of the 126 RNA editing sites from 11 PCGs. The results of this study suggest that the predominant conformation of the S. officinalis mitogenome are two circular chromosomes, and the stop gain of rpl5 was found through RNA editing events of the Salvia mitogenome.

Improved efficiency of Sedum lineare (Crassulaceae) in remediation of arsenic-contaminated soil by phosphate-dissolving strain P-1 in association with phosphate rock.

The selection of appropriate plants and growth strategies is a key factor in improving the efficiency and universal applicability of phytoremediation. Sedum lineare grows rapidly and tolerates multiple adversities. The effects of inoculation of Acinetobacter sp. phosphate solubilizing bacteria P-1 and application of phosphate rock (PR) as additives on the remediation efficiency of As-contaminated soil by S. lineare were investigated. Compared with the control, both the single treatment and the combination of inoculation with strain P-1 and application of PR improved the biomass by 30.7-395.5%, chlorophyll content by 48.1-134.8%, total protein content by 12.5-92.4% and total As accumulation by 45.1-177.5%, and reduced the As-induced oxidative damage. Inoculation with strain P-1 increased the activities of superoxide dismutases and catalases of S. lineare under As stress, decreased the accumulation of reactive oxygen species in plant tissues and promoted the accumulation of As in roots. In contrast, simultaneous application of PR decreased As concentration in S. lineare tissues, attenuated As-induced lipid peroxidation and improved As transport to shoots. In addition, the combined application showed the best performance in improving resistance and biomass, which significantly increased root length by 149.1%, shoot length by 33%, fresh weight by 395.5% and total arsenic accumulation by 159.2%, but decreased the malondialdehyde content by 89.1%. Our results indicate that the combined application of strain P-1 and PR with S. lineare is a promising bioremediation strategy to accelerate phytoremediation of As-contaminated soils.

Overexpression of SHORT-ROOT2 transcription factor enhances the outgrowth of mature axillary buds in poplar trees.

SHORT-ROOT (SHR) transcription factors play important roles in asymmetric cell division and radial patterning of Arabidopsis roots. In hybrid poplar (P. tremula × P. alba clone INRA 717-1B4), PtaSHR2 was preferentially expressed in axillary buds (AXBs) and transcriptionally up-regulated during AXB maturation and activation. Overexpression of SHR2 (PtSHR2OE) induced an enhanced outgrowth of AXBs below the bud maturation point, with a simultaneous transition of an active shoot apex into an arrested terminal bud. The larger and more mature AXBs of PtSHR2OE trees revealed altered expression of genes involved in axillary meristem initiation and bud activation, as well as a higher ratio of cytokinin to auxin. To elucidate the underlying mechanism of PtSHR2OE-induced high branching, subsequent molecular and biochemical studies showed that compared with wild-type trees, decapitation induced a quicker bud outburst in PtSHR2OE trees, which could be fully inhibited by exogenous application of auxin or cytokinin biosynthesis inhibitor, but not by N-1-naphthylphthalamic acid. Our results indicated that overexpression of PtSHR2B disturbed the internal hormonal balance in AXBs by interfering with the basipetal transport of auxin, rather than causing auxin biosynthesis deficiency or auxin insensitivity, thereby releasing mature AXBs from apical dominance and promoting their outgrowth.

De Novo Hybrid Assembly of the Salvia miltiorrhiza Mitochondrial Genome Provides the First Evidence of the Multi-Chromosomal Mitochondrial DNA Structure of Salvia Species.

Salvia miltiorrhiza has been an economically important medicinal plant. Previously, an S. miltiorrhiza mitochondrial genome (mitogenome) assembled from Illumina short reads, appearing to be a single circular molecule, has been published. Based on the recent reports on the plant mitogenome structure, we suspected that this conformation does not accurately represent the complexity of the S. miltiorrhiza mitogenome. In the current study, we assembled the mitogenome of S. miltiorrhiza using the PacBio and Illumina sequencing technologies. The primary structure of the mitogenome contained two mitochondrial chromosomes (MC1 and MC2), which corresponded to two major conformations, namely, Mac1 and Mac2, respectively. Using two approaches, including (1) long reads mapping and (2) polymerase chain reaction amplification followed by Sanger sequencing, we observed nine repeats that can mediate recombination. We predicted 55 genes, including 33 mitochondrial protein-coding genes (PCGs), 3 rRNA genes, and 19 tRNA genes. Repeat analysis identified 112 microsatellite repeats and 3 long-tandem repeats. Phylogenetic analysis using the 26 shared PCGs resulted in a tree that was congruent with the phylogeny of Lamiales species in the APG IV system. The analysis of mitochondrial plastid DNA (MTPT) identified 16 MTPTs in the mitogenome. Moreover, the analysis of nucleotide substitution rates in Lamiales showed that the genes atp4, ccmB, ccmFc, and mttB might have been positively selected. The results lay the foundation for future studies on the evolution of the Salvia mitogenome and the molecular breeding of S. miltiorrhiza.

Phylogenetic analysis and development of molecular markers for five medicinal Alpinia species based on complete plastome sequences.

BACKGROUND: Alpinia species are widely used as medicinal herbs. To understand the taxonomic classification and plastome evolution of the medicinal Alpinia species and correctly identify medicinal products derived from Alpinia species, we systematically analyzed the plastome sequences from five Alpinia species. Four of the Alpinia species: Alpinia galanga (L.) Willd., Alpinia hainanensis K.Schum., Alpinia officinarum Hance, and Alpinia oxyphylla Miq., are listed in the Chinese pharmacopeia. The other one, Alpinia nigra (Gaertn.) Burtt, is well known for its medicinal values. RESULTS: The four Alpinia species: A. galanga, A. nigra, A. officinarum, and A. oxyphylla, were sequenced using the Next-generation sequencing technology. The plastomes were assembled using Novoplasty and annotated using CPGAVAS2. The sizes of the four plastomes range from 160,590 bp for A. galanga to 164,294 bp for A. nigra, and display a conserved quadripartite structure. Each of the plastomes encodes a total of 111 unique genes, including 79 protein-coding, 28 tRNA, and four rRNA genes. In addition, 293-296 SSRs were detected in the four plastomes, of which the majority are mononucleotides Adenine/Thymine and are found in the noncoding regions. The long repeat analysis shows all types of repeats are contained in the plastomes, of which palindromic repeats occur most frequently. The comparative genomic analyses revealed that the pair of the inverted repeats were less divergent than the single-copy region. Analysis of sequence divergence on protein-coding genes showed that two genes (accD and ycf1) had undergone positive selection. Phylogenetic analysis based on coding sequence of 77 shared plastome genes resolves the molecular phylogeny of 20 species from Zingiberaceae. In particular, molecular phylogeny of four sequenced Alpinia species (A. galanga, A. nigra, A. officinarum, and A. oxyphylla) based on the plastome and nuclear sequences showed congruency. Furthermore, a comparison of the four newly sequenced Alpinia plastomes and one previously reported Alpinia plastomes (accession number: NC_048461) reveals 59 highly divergent intergenic spacer regions. We developed and validated two molecular markers Alpp and Alpr, based on two regions: petN-psbM and psaJ-rpl33, respectively. The discrimination success rate was 100 % in validation experiments. CONCLUSIONS: The results from this study will be invaluable for ensuring the effective and safe uses of Alpinia medicinal products and for the exploration of novel Alpinia species to improve human health.

Label-Free and Simultaneous Mechanical and Electrical Characterization of Single Plant Cells Using Microfluidic Impedance Flow Cytometry.

Despite that single-cell-type-level analyses have been extensively conducted on animal models to gain new insights into complex biological processes; the unique biological and physiological properties of plant cells have not been widely studied at single-cell resolution. In this work, an electrical impedance flow cytometry was fabricated based on microfluidics with constriction microchannel to simultaneously characterize the mechanical and electrical properties of single plant cells. Protoplasts from two model plant species, the herbaceous Arabidopsis thaliana and the woody Populus trichocarpa, could be readily discriminated by their respective mechanical traits, but not by electrical impedance. On the contrary, overexpression of a red fluorescent protein on plasma membrane resulted in changes in cell electrical impedance instead of cell deformability. During primary cell wall (PCW) regeneration, this extracellular layer outside of protoplasts introduced dramatic variations in both mechanical and electrical properties of single plant cells. Furthermore, the effects of auxin, an essential phytohormone regulating PCW reformation, were validated on this platform. Taken together, our results revealed a novel application of microfluidic impedance flow cytometry in the field of plant science to simultaneously characterize dual biophysical properties at single-cell resolution, which could be further developed as a powerful and reliable tool for plant cell phenotyping and cell fate specification.

TDIF peptides regulate root growth by affecting auxin homeostasis and PINs expression in Arabidopsis thaliana.

MAIN CONCLUSION: TDIF and TDIF-like peptides in excess simultaneously facilitate primary root elongation and lateral root formation through regulating auxin distribution and transport. Tracheary element differentiation inhibitory factor (TDIF) plays key roles in mediating cell-cell communication and stem cell maintenance during vascular development. Recently, TDIF has also been linked to lateral root (LR) organogenesis through Brassinosteroid Insensitive 2 (BIN2) action. In this work, by comparing the in vitro and in vivo activities of AtCLE41-encoded TDIF and one poplar-derived TDIF-like peptide in Arabidopsis thaliana, we demonstrated that both TDIFs promoted primary root (PR) growth and stimulated LR formation. Without affecting auxin biosynthesis and catabolism, TDIFs suppressed the auxin maxima at PR apex but intensified the auxin accumulation at LR initiation sites along the longitudinal axis of PR. TDIF did not alter root sensitivity to exogenous auxin and mutants with varied endogenous auxin levels responded to TDIF peptides in a wild-type manner but to a lesser extent. Intriguingly, TDIF specifically upregulated the transcript abundance of PINs and multiple pin mutants displayed insensitivity to TDIF, demonstrating that PIN-mediated polar auxin transport (PAT) is indispensably required for the TDIF-induced root phenotypes. Taken together, our results revealed that TDIF might target PAT via mobilizing auxin efflux carriers to dynamically regulate the auxin signaling output and hence facilitate PR growth and LR formation.

Overexpression of a Eutrema salsugineum phosphate transporter gene EsPHT1;4 enhances tolerance to low phosphorus stress in soybean.

OBJECTIVE: To enhance Pi absorption and utilization efficiency of soybean, a member of PHT1 gene family was isolated and characterized from E. salsugineum, which was a homologous gene of AtPHT1;4 and consequently designated as EsPHT1;4. RESULTS: Quantitative real-time PCR (qRT-PCR) analysis showed that the transcript level of EsPHT1;4 significantly increased both in roots and leaves of E. salsugineum under Pi deficient conditions. Furthermore, EsPHT1;4 was transferred to soybean cultivar "YD22" using an Agrobacterium-mediated cotyledonary-node transformation method. Overexpression of EsPHT1;4 in soybean not only promoted the increase of plant biomass and yield of transgenic plants upon low P stress, but also increased the accumulation and transportation of Pi from roots to leaves in the transgenic soybean lines. CONCLUSION: EsPHT1;4 was critical for controlling the accumulation and translocation of Pi in plants, and can be subsequently used as an effective foreign gene for the improvement of P use efficiency of crops by genetic manipulation.

Transcriptome sequencing and functional analysis of Sedum lineare Thunb. upon salt stress.

Soil salinization is one major constraint to plant geographical distribution, yield, and quality, and as an ideal plant for the "greening" of flat-roofed buildings, Sedum lineare Thunb. has strong tolerance against a variety of environmental adversities including salinity with the underlying mechanism still remaining unknown. In this study, we performed de novo transcriptome sequencing on leaf and root samples of NaCl-treated S. lineare Thunb. and identified 584 differentially expressed genes (DEGs), which were further annotated by gene function classification and pathway assignments using the public data repositories. In addition to the increased gene expression level verified by qRT-PCR, the elevated activities of the corresponding enzymes were also demonstrated for peroxidase (POD), glutathione peroxidases (GPX), and cysteine synthase (CSase) in the NaCl-treated roots. Furthermore, two highly inducible genes without known functions related to salt tolerance were selected to be overexpressed and tested for their effects on salt tolerance in the model plant, Arabidopsis thaliana. Upon 150 mM NaCl treatment, 35S:SlCXE but not 35S:SlCYP72A transgenic Arabidopsis seedlings exhibited improved salt resistance as shown by the increased seed germination rates and longer primary roots of transgenic seedlings when compared to wild-type plants. Taken together, this work laid a foundation for a better understanding of the salt adaptation mechanism of S. lineare Thunb. and genes identified could serve as useful resources for the development of more salt-tolerant varieties of other species through genetic engineering.

SENP7 Potentiates cGAS Activation by Relieving SUMO-Mediated Inhibition of Cytosolic DNA Sensing.

Cyclic GMP-AMP (cGAMP) synthase (cGAS, a.k.a. MB21D1), a cytosolic DNA sensor, catalyzes formation of the second messenger 2'3'-cGAMP that activates the stimulator of interferon genes (STING) signaling. How the cGAS activity is modulated remains largely unknown. Here, we demonstrate that sentrin/SUMO-specific protease 7 (SENP7) interacted with and potentiated cGAS activation. The small ubiquitin-like modifier (SUMO) was conjugated onto the lysine residues 335, 372 and 382 of cGAS, which suppressed its DNA-binding, oligomerization and nucleotidyl-transferase activities. SENP7 reversed this inhibition via catalyzing the cGAS de-SUMOylation. Consistently, silencing of SENP7 markedly impaired the IRF3-responsive gene expression induced by cGAS-STING axis. SENP7-knockdown mice were more susceptible to herpes simplex virus 1 (HSV-1) infection. SENP7 was significantly up-regulated in patients with SLE. Our study highlights the temporal modulation of the cGAS activity via dynamic SUMOylation, uncovering a novel mechanism for fine-tuning the STING signaling in innate immunity.

Farrerol overcomes the invasiveness of lung squamous cell carcinoma cells by regulating the expression of inducers of epithelial mesenchymal transition.

In recent years Epithelial Mesenchymal Transition (EMT) has been proposed as a mechanism indispensable to acquisition of metastatic properties by tumor cells. In this study we tested the ability of Farrerol, a Chinese herb-derived compound to ablate the EMT in human lung squamous cell carcinoma cells. Human lung squamous cell carcinoma cells, Calu-1 were treated with various concentrations of Farrerol for 24 h to examine its effect on their viability by the MTT assay. Only those concentrations which showed least effect on the viability of Calu-1 cells were further used to evaluate the expression of epithelial and mesenchymal markers by western blotting. Furthermore the effect of such concentrations on the migration and invasion of Calu-1 cells was determined by wound healing and transwell invasion assays respectively. The results demonstrated that Farrerol treatment led to the downregulation of Slug and Zeb-1, transcriptional regulators of EMT with the concomitant increase and decrease in the expression of E-cadherin and vimentin respectively. These data were further supported by migration and invasion assays which demonstrated that Farrerol treatment caused inhibited the migration and invasion of Calu-1 lung squamous cell carcinoma cells. Taken together, our results indicate that Farrerol suppresses lung squamous cell carcinoma cell metastatic potential by modulating the expression of EMT proteins.

Retraction Note to: An assessment of transgenomics as a tool for gene discovery in Populus euphratica Oliv.

This article (Zhou et al. 2018) has been retracted by the authors because the sequence BIBAC 002A111F06 was incorrectly assigned to the wrong bacterial species. The BIBAC 002A111F06 sequence (GenBank Accession KC129717) reported in the paper was attributed to Populus euphratica Oliv. The BLAST search of this KC129717 sequence against the nr database at NCBI showed that it has very high similarity to a genomic sequence from the gram-negative bacteria Stenotrophomonas maltophilia. The bacterium associates with Populus euphratica Oliv. and DNA isolated from Populus euphratica Oliv. for the construction of the BIBAC clone library inlcuded DNA from Stenotrophomonas maltophilia. Therefore, the phenotype of the transgenic Arabidopsis line carrying the KC129717 sequence cannot be attributed to genes from Populus euphratica Oliv. The authors apologize for the confusion and misinterpretation of our data resulting from the incorrect sequence assignment. All authors agree to this retraction.

An assessment of transgenomics as a tool for gene discovery in Populus euphratica Oliv.

KEY MESSAGE: Transgenomics for gene discovery in Populus euphratica. Transgenomics, a member of the omics family of methodologies, is characterized as the introduction of DNA from one organism into another on a genome-wide scale followed by the identification of recipients with altered phenotypes. This strategy allows investigators to identify the gene(s) involved in these phenotypic changes. It is particularly promising for woody plants that have a long life cycle and for which molecular tools are limited. In this study, we constructed a large-insert binary bacterial artificial chromosome library of Populus euphratica, a stress-tolerant poplar species, which included 55,296 clones with average insert sizes of about 127 kb. To date, 1077 of the clones have been transformed into Arabidopsis thaliana via Agrobacterium by the floral dip method. Of these, 69 transgenic lines showed phenotypic changes represented by diverse aspects of plant form and development, 22 of which were reproducibly associated with the same phenotypic change. One of the clones conferring transgenic plants with increased salt tolerance, 002A1F06, was further analyzed and the 127,284 bp insert in this clone harbored eight genes that have been previously reported to be involved in stress resistance. This study demonstrates that transgenomics is useful in the study of functional genomics of woody plants and in the identification of novel gene(s) responsible for economically important traits. Thus, transgenomics can also be used for validation of quantitative trait loci mapped by molecular markers.

Distinct transgenic effects of poplar TDIF genes on vascular development in Arabidopsis.

KEY MESSAGE: Poplar CLE genes encoding TDIF motifs differentially regulate vascular cambial cell division and woody tissue organization in transgenic Arabidopsis. In Arabidopsis, CLE41 and CLE44 genes encode the tracheary element differentiation inhibitory factor (TDIF) peptide, which functions as a non-cell autonomous signal to regulate vascular development, and overexpression of AtCLE41/CLE44 generate similar phenotypic defects. In poplar, there are six CLE genes (PtTDIF1-4 and PtTDIF-like1-2) encoding two TDIF peptides (TDIF and TDIF-like peptide), which exhibit nearly same activities when exogenously applied to Arabidopsis seedlings. In this work, for each TDIF peptide, we chose two poplar CLE genes (PtTDIF2 and 3 for TDIF, and PtTDIF-like1-2 for TDIF-like peptide) to compare their in vivo effects in transgenic Arabidopsis. Our results showed that transgenic Arabidopsis lines overexpressing each individual PtTDIF gene exhibited dramatically distinct phenotypes associated with vascular development, demonstrating that TDIF motif is not the only functional determinant after genetic transformation. Moreover, we revealed that overexpressed poplar TDIFs enhanced the proliferation of (pro)cambial cells only in hypocotyls, but not in inflorescence stems by differentially regulating the transcriptional levels of WOX4 and WOX14 in these two tissues.

Ferrerol overcomes the invasiveness of lung squamous cell carcinoma cells by regulating the expression of inducers of Epithelial Mesenchymal Transition.

In recent years Epithelial Mesenchymal Transition (EMT) has been proposed as a mechanism indispensable to acquisition of metastatic properties by tumor cells. In this study we tested the ability of Ferrerol, a Chinese herb-derived compound to ablate the EMT in human lung squamous cell carcinoma cells. Human lung squamous cell carcinoma cells, Calu-1 were treated with various concentrations of Ferrerol for 24 h to examine its effect on their viability by the MTT assay. Only those concentrations which showed least effect on the viability of Calu-1 cells were further used to evaluate the expression of epithelial and mesenchymal markers by western blotting. Furthermore the effect of such concentrations on the migration and invasion of Calu-1 cells was determined by wound healing and transwell invasion assays respectively. The results demonstrated that Ferrerol treatment led to the downregulation of Slug and Zeb-1, transcriptional regulators of EMT with the concomitant increase and decrease in the expression of E-cadherin and vimentin respectively. These data were further supported by migration and invasion assays which demonstrated that Ferrerol treatment caused inhibited the migration and invasion of Calu-1 lung squamous cell carcinoma cells. Taken together, our results indicate that Ferrerol suppresses lung squamous cell carcinoma cell metastatic potential by modulating the expression of EMT proteins.

La1+xBa1-xGa3O7+0.5x Oxide Ion Conductor: Cationic Size Effect on the Interstitial Oxide Ion Conductivity in Gallate Melilites.

Substitution of La3+ for Ba2+ in LaBaGa3O7 melilite yields a new interstitial-oxide-ion conducting La1+xBa1-xGa3O7+0.5x solid solution, which only extends up to x = 0.35, giving a maximum interstitial oxygen content allowed in La1+xBa1-xGa3O7+0.5x as about half of those allowed in La1+x(Sr/Ca)1-xGa3O7+0.5x. La1.35Ba0.65Ga3O7.175 ceramic displays bulk conductivity ∼1.9 × 10-3 S/cm at 600 °C, which is lower than those of La1.35(Sr/Ca)0.65Ga3O7.175, showing the reduced mobility for the oxygen interstitials in La1+xBa1-xGa3O7+0.5x than in La1+x(Sr/Ca)1-xGa3O7+0.5x. Rietveld analysis of neutron powder diffraction data reveals that the oxygen interstitials in La1.35Ba0.65Ga3O7.175 are located within the pentagonal tunnels at the Ga level between two La/Ba cations along the c-axis and stabilized via incorporating into the bonding environment of a three-linked GaO4 among the five GaO4 tetrahedra forming the pentagonal tunnels, similar to the Sr and Ca counterparts. Both static lattice atomistic simulation and density functional theory calculation show that LaBaGa3O7 has the largest formation energy for oxygen interstitial defects among La1+xM1-xGa3O7+0.5x (M = Ba, Sr, Ca), consistent with the large Ba2+ cations favoring interstitial oxygen defects in melilite less than the small cations Sr2+ and Ca2+. The cationic-size control of the ability to accommodate the oxygen interstitials and maintain high mobility for the oxygen interstitials in La1+xM1-xGa3O7+0.5x (M = Ba, Sr, Ca) gallate melilites is understood in terms of local structural relaxation to accommodate and transport the oxygen interstitials. The accommodation and migration of the interstitials in the melilite structure require the tunnel-cations being able to adapt to the synergic size expansion for the interstitial-containing tunnel and contraction for the tunnels neighboring the interstitial-containing tunnel and continuous tunnel-size expansion and contraction. However, the large oxygen bonding separation requirement of the large Ba2+ along the tunnel not only suppresses the ability to accommodate the interstitials in the tunnels neighboring the Ba2+-containing tunnel but also reduces the mobility of the oxygen interstitials among the pentagonal tunnels.