Regulation of SmEXPA13 expression by SmMYB1R1-L enhances salt tolerance in Salix matsudana Koidz.

Expansins, cell wall proteins, play a significant role in plant stress resistance. Our previous study confirmed the expression of the expansin gene SmEXPA13 from Salix matsudana Koidz. enhanced salt tolerance of plants. This report presented an assay that the expression of SmEXPA13 was higher in the salt-resistant willow variety 9901 than in the salt-sensitive variety Yanjiang. In order to understand the possible reasons, a study of the regulation process was conducted. Despite being cloned from both varieties, SmEXPA13 and its promotor showed no significant differences in the structure and sequence. A transcription factor (TF), SmMYB1R1-L, identified through screening the yeast library of willow cDNA, was found to regulate SmEXPA13. Yeast one-hybrid (Y1H) assay confirmed that SmMYB1R1-L could bind to the MYB element at the -520 bp site on the SmEXPA13 promotor. A dual-luciferase reporter assay also demonstrated that SmMYB1R1-L could greatly activate SmEXPA13 expression. The willow calli with over-expression of SmMYB1R1-L exhibited better physiological performance than the wild type under salt stress. Further testing the expression of SmMYB1R1-L displayed it significantly higher in 9901 willow than that in Yanjiang under salt stress. In conclusion, the high accumulation of SmMYB1R1-L in 9901 willow under salt stress led to the high expression of SmEXPA13, resulting in variations in salt stress resistance among willow varieties. The SmMYB1R1-L/SmEXPA13 cascade module in willow offers a new perspective on plant resistance mechanisms.

Hepatic and portal vein segmentation with dual-stream deep neural network.

BACKGROUND: Liver lesions mainly occur inside the liver parenchyma, which are difficult to locate and have complicated relationships with essential vessels. Thus, preoperative planning is crucial for the resection of liver lesions. Accurate segmentation of the hepatic and portal veins (PVs) on computed tomography (CT) images is of great importance for preoperative planning. However, manually labeling the mask of vessels is laborious and time-consuming, and the labeling results of different clinicians are prone to inconsistencies. Hence, developing an automatic segmentation algorithm for hepatic and PVs on CT images has attracted the attention of researchers. Unfortunately, existing deep learning based automatic segmentation methods are prone to misclassifying peripheral vessels into wrong categories. PURPOSE: This study aims to provide a fully automatic and robust semantic segmentation algorithm for hepatic and PVs, guiding subsequent preoperative planning. In addition, to address the deficiency of the public dataset for hepatic and PV segmentation, we revise the annotations of the Medical Segmentation Decathlon (MSD) hepatic vessel segmentation dataset and add the masks of the hepatic veins (HVs) and PVs. METHODS: We proposed a structure with a dual-stream encoder combining convolution and Transformer block, named Dual-stream Hepatic Portal Vein segmentation Network, to extract local features and long-distance spatial information, thereby extracting anatomical information of hepatic and portal vein, avoiding misdivisions of adjacent peripheral vessels. Besides, a multi-scale feature fusion block based on dilated convolution is proposed to extract multi-scale features on expanded perception fields for local features, and a multi-level fusing attention module is introduced for efficient context information extraction. Paired t-test is conducted to evaluate the significant difference in dice between the proposed methods and the comparing methods. RESULTS: Two datasets are constructed from the original MSD dataset. For each dataset, 50 cases are randomly selected for model evaluation in the scheme of 5-fold cross-validation. The results show that our method outperforms the state-of-the-art Convolutional Neural Network-based and transformer-based methods. Specifically, for the first dataset, our model reaches 0.815, 0.830, and 0.807 at overall dice, precision, and sensitivity. The dice of the hepatic and PVs are 0.835 and 0.796, which also exceed the numeric result of the comparing methods. Almost all the p-values of paired t-tests on the proposed approach and comparing approaches are smaller than 0.05. On the second dataset, the proposed algorithm achieves 0.749, 0.762, 0.726, 0.835, and 0.796 for overall dice, precision, sensitivity, dice for HV, and dice for PV, among which the first four numeric results exceed comparing methods. CONCLUSIONS: The proposed method is effective in solving the problem of misclassifying interlaced peripheral veins for the HV and PV segmentation task and outperforming the comparing methods on the relabeled dataset.

Lymphoma Recognition in Histology Image of Gastric Mucosal Biopsy with Prototype Learning.

Lymphomas are a group of malignant tumors developed from lymphocytes, which may occur in many organs. Therefore, accurately distinguishing lymphoma from solid tumors is of great clinical significance. Due to the strong ability of graph structure to capture the topology of the micro-environment of cells, graph convolutional networks (GCNs) have been widely used in pathological image processing. Nevertheless, the softmax classification layer of the graph convolutional models cannot drive learned representations compact enough to distinguish some types of lymphomas and solid tumors with strong morphological analogies on H&E-stained images. To alleviate this problem, a prototype learning based model is proposed, namely graph convolutional prototype network (GCPNet). Specifically, the method follows the patch-to-slide architecture first to perform patch-level classification and obtain image-level results by fusing patch-level predictions. The classification model is assembled with a graph convolutional feature extractor and prototype-based classification layer to build more robust feature representations for classification. For model training, a dynamic prototype loss is proposed to give the model different optimization priorities at different stages of training. Besides, a prototype reassignment operation is designed to prevent the model from getting stuck in local minima during optimization. Experiments are conducted on a dataset of 183 Whole slide images (WSI) of gastric mucosa biopsy. The proposed method achieved superior performance than existing methods.Clinical relevance- The work proposed a new deep learning framework tailored to lymphoma recognition on pathological image of gastric mucosal biopsy to differentiate lymphoma, adenocarcinoma and inflammation.

The complete mitochondrial genome of Zelkova schneideriana Hand-Mazz (Ulmaceae) and phylogenetic analysis.

Zelkova schneideriana Hand-Mazz is a second-class key protected wild plant in China. Here, the complete mitochondrial genome of Zelkova schneideriana Hand-Mazz was sequenced using Nanopore Sequel and Illumina NovaSeq platform. The mitochondrial genome was assembled into three circular-mapping molecules with the genome sizes of 154,640 bp, 192,388 bp and 146,907 bp, including 36 protein-coding genes, 19 tRNA genes, and 3 rRNA genes. Phylogenetic analysis indicated that Zelkova schneideriana Hand-Mazz is close with Hemiptelea davidii, a species in same Ulmaceae Mirb.

Expression, Purification, and Preliminary Protection Study of Dehydrin PicW1 From the Biomass of Picea wilsonii.

Dehydrins (DHNs) belong to group II of late embryogenesis-abundant (LEA) proteins, which are up-regulated in most plants during cold, drought, heat, or salinity stress. Despite the importance of dehydrins for the plants to resist abiotic stresses, it is necessary to obtain plant-derived dehydrins from different biomass. Generally, dehydrin PicW1 from Picea wilsonii is involved in Kn-type dehydrin with five K-segments, which has a variety of biological activities. In this work, Picea wilsonii dehydrin PicW1 was expressed in Escherichia coli and purified by chitin-affinity chromatography and size-exclusion chromatography, which showed as a single band by SDS-PAGE. A cold-sensitive enzyme of lactate dehydrogenase (LDH) is used to explore the protective activities of other proteins. Temperature stress assays showed that PicW1 had an effective protective effect on LDH activity, which was better than that of bovine serum albumin (BSA). This study provides insights into the purification and protective activity of K5 DHNs for the advancement of dehydrin structure and function from biomass.

Genome sequencing and phylogenetic analysis of allotetraploid Salix matsudana Koidz.

Polyploidy is a common phenomenon among willow species. In this study, genome sequencing was conducted for Salix matsudana Koidz (also named Chinese willow), an important greening and arbor tree species, and the genome of this species was compared with those of four other tree species in Salicaceae. The total genome sequence of S. matsudana was 655.72 Mb in size, with repeated sequences accounting for 45.97% of the total length. In total, 531.43 Mb of the genome sequence could be mapped onto 38 chromosomes using the published genetic map as a reference. The genome of S. matsudana could be divided into two groups, the A and B genomes, through homology analysis with the genome of Populus trichocarpa, and the A and B genomes contained 23,985 and 25,107 genes, respectively. 4DTv combined transposon analysis predicted that allotetraploidy in S. matsudana appeared ~4 million years ago. The results from this study will help reveal the evolutionary history of S. matsudana and lay a genetic basis for its breeding.

Genome-wide profiling of alternative splicing genes in hybrid poplar (P.alba×P.glandulosa cv.84K) leaves.

Alternative splicing (AS) is a post-transcriptional process common in plants and essential for regulation of environmental fitness of plants. In the present study, we focus on the AS events in poplar leaves to understand their effects on plant growth and development. The hybrid poplar (P.alba×P.glandulosa cv.84K) leaves were collected for RNA extraction. The extracted RNA was sequenced using on an Illumina HiSeq™ 2000 platform. Using the Populus trichocarpa genome as the reference, a total of 3810 AS genes were identified (9225 AS events), which accounted for 13.51% of all the expressed genes. Intron retention was the most common AS event, accounting for 43.86% of all the AS events, followed by alternative 3' splice sites (23.75%), alternative 5' splice sites (23.71%), and exon skipping (8.68%). Chromosomes 10 had the most condensed AS events (33.67 events/Mb) and chromosome 19 had the least (12.42 events/Mb). Association analysis showed that AS in the poplar leaves was positively correlated with intron length, exon number, exon length, and gene expression level, and was negatively correlated with GC content. AS genes in the poplar leaves were associated mainly with inositol phosphate metabolism and phosphatidylinositol signaling system pathways that would be significant on wooden plant production.

Cavitation characteristics of flowing low and high boiling-point perfluorocarbon phase-shift nanodroplets during focused ultrasound exposures.

This work investigated and compared the dynamic cavitation characteristics between low and high boiling-point phase-shift nanodroplets (NDs) under physiologically relevant flow conditions during focused ultrasound (FUS) exposures at different peak rarefactional pressures. A passive cavitation detection (PCD) system was used to monitor cavitation activity during FUS exposure at various acoustic pressure levels. Root mean square (RMS) amplitudes of broadband noise, spectrograms of the passive cavitation detection signals, and normalized inertial cavitation dose (ICD) values were calculated. Cavitation activity of low-boiling-point perfluoropentane (PFP) NDs and high boiling-point perfluorohexane (PFH) NDs flowing at in vitro mean velocities of 0-15 cm/s were compared in a 4-mm diameter wall-less vessel in a transparent tissue-mimicking phantom. In the static state, both types of phase-shift NDs exhibit a sharp rise in cavitation intensity during initial FUS exposure. Under flow conditions, cavitation activity of the PFH NDs reached the steady state less rapidly compared to PFP NDs under the lower acoustic pressure (1.35 MPa); at the higher acoustic pressure (1.65 MPa), the RMS amplitude increased more sharply during the initial FUS exposure period. In particular, the RMS-time curves of the PFP NDs shifted upward as the mean flow velocity increased from 0 to 15 cm/s; the RMS amplitude of the PFH ND solution increased from 0 to 10 cm/s and decreased at 15 cm/s. Moreover, amplitudes of the echo signal for the low boiling-point PFP NDs were higher compared to the high boiling-point PFH NDs in the lower frequency range, whereas the inverse occurred in the higher frequency range. Both PFP and PFH NDs showed increased cavitation activity in the higher frequency under the flow condition compared to the static state, especially PFH NDs. At 1.65 MPa, normalized ICD values for PFH increased from 0.93 ± 0.03 to 0.96 ± 0.04 and from 0 to 10 cm/s, then decreased to 0.86 ± 0.05 at 15 cm/s. This work contributes to our further understanding of cavitation characteristics of phase-shift NDs under physiologically relevant flow conditions during FUS exposure. In addition, the results provide a reference for selecting suitable phase-shift NDs to enhance the efficiency of cavitation-mediated ultrasonic applications.

Codon usage by chloroplast gene is bias in Hemiptelea davidii.

The base composition of the chloroplast genes is of great interest because they play a highly significant role in the evolutionary development of the plants. Evaluation of the 48 chloroplast protein-coding genes of Hemiptelea davidii showed that the average GC content was about 37.32%, while at the third codon base position alone the average GC content was only 27.80%. The 48 genes were classified into five groups based on the gene function and each group displayed specific codon characteristics. Based on the relative synonymous codon usage analysis, a total of 30 high-frequency codons and 11 optimal codons were identified, most of them ended with A or T. Neutrality plot, ENC-plot and PR2-plot analyses showed that the codon usage bias of the chloroplast genes of H. davidii was greatly influenced by natural selection pressures. Meanwhile, the frequency of codon usage of chloroplast genes among different plant species displayed similarities, with some synonymous codons were preferred to be used in H. davidii. In this study, the codon usage pattern of the chloroplast protein coding genes of H. davidii provides us with a better understanding of the expression of chloroplast genes, and may advice the future molecular breeding programmes.

In situ localization of alkaline phosphatase activity in tumor cells by an aggregation-induced emission fluorophore-based probes.

In situ detection of certain specific enzyme activities in cells is deeply attached to tumor diagnosis. Conventional enzyme-responsive fluorescent probes have difficulty detecting targeted enzymes in situ in cells due to the low detection accuracy caused by the spread of fluorescence probes. In order to solve this problem, we have designed and synthesized an enzyme-responsive, water-soluble fluorescent probe with AIE characteristics, which could aggregate and precipitate to produce in situ fluorescence when reacting with the targeted enzyme in cells. The AIE fluorophore (TPEQH) was utilized to design the enzyme-responsive, fluorescent probe (TPEQHA) by introducing a phosphate group on to it, which could be specifically decomposed by the targeted enzyme, namely alkaline phosphatase (ALP). In tumor cells, TPEQH was highly produced due to the interaction of phosphate on the TPEQHA and the overexpressed ALP. Water-insoluble TPEQH then precipitated and release fluorescence in situ, thereby successfully detecting the ALP. Furthermore, the expression level of ALP could be determined by the fluorescence intensity of TPEQH with higher accuracy due to the inhibition of TPEQH leak, which demonstrated a potential application of in suit ALP detection in both clinical diagnosis and scientific research of tumor.

Over-expression of PttEXPA8 gene showed various resistances to diverse stresses.

Expansins play a pivotal role in plant adaptation to environmental stress via cell wall loosening. To evaluate the roles of expansin in response to different environmental stress conditions, the expansin gene PttEXPA8 from Populus tomentosa was transformed into tobacco. Analysis of physiological indices demonstrated the transgenic plants with improved resistance to heat, drought, salt, cold, and cadmium stress but to different extents. In mature plants, PttEXPA8 exerted the greatest effect on heat stress, with a response index value of 137.46%, followed by drought, cadmium, cold, and salt stress with response index values of 101.04%, 70.61%, 69.95%, and 54.68%, respectively. Over-expression of PttEXPA8 resulted in differential responses in physiological indices to the stresses. Soluble sugar content showed the highest response to the stresses, with an average response index value of 29.29%, whereas the absolute response index value for malondialdehyde content, relative electrolyte leakage, chlorophyll content, and superoxide dismutase activity ranged from 11.01% to 19.21%. The present results provide insight into the roles of expansin in stress resistance in Populus.

Complete mitochondrial genome of Hemiptelea davidii (Ulmaceae) and phylogenetic analysis.

Hemiptelea davidii (Hance) Planch is a potential valuable forest tree in arid sandy environments. Here, the complete mitochondrial genome of H. davidii was assembled using a combination of the PacBio Sequel data and the Illumina Hiseq data. The mitochondrial genome is 460,941 bp in length, including 37 protein-coding genes, 19 tRNA genes, and three rRNA genes. The GC content of the whole mitochondrial genome is 44.84%. Phylogenetic analyses indicated that H. davidii is close with Cannabis and Morus species.

Comparative analysis of expansin gene codon usage patterns among eight plant species.

Expansins are essential components of plant cell wall and play an important role in plant growth, development, and stress resistance via loosening function. To understand the codon usage pattern of expansin genes, we gained the sequence data of expansin genes from eight plant species. Statistics analysis showed obvious codon characteristics between monocot and dicot plants. Comparably, expansin genes in monocot plants had really higher GC content, more high-frequency codons, and more optimal codons than that in dicot plants. Several monocot plants performed somehow as dicot plants in a few characters. Codon information of expansin genes might contribute to the understanding of the relationship and evolution clues between monocot and dicot plants. It further gained insight into the improvement of the gene expression and roles.

Transgenic creeping bentgrass plants expressing a Picea wilsonii dehydrin gene (PicW) demonstrate improved freezing tolerance.

Agrostis stolonifera L. 'Penn A-4' is a common creeping bentgrass species that is widely used in urban landscaping and golf courses. To prolong the green stage of this grass, a dehydrin gene PicW isolated from Wilson's spruce (Picea wilsonii) was transformed into plants of 'Penn A-4' cultivar via a straightforward stolon node infection system. A putative transgenic plant was obtained and its tolerance to low-temperature stress was evaluated. When the transgenic line was subjected to a freezing (- 5 °C) treatment, it showed better viability and more robust physiology than wild type, as evidenced by higher soluble sugar and proline contents, and lower relative electrical conductivity and malondialdehyde content. The transgenic line also showed tolerance to a chilling treatment (5 °C), although its performance was not significantly different from that of wild-type plants. Overall, the research here clearly revealed the explicit role of PicW in increasing freezing tolerance of grass at the whole-plant level, and demonstrated that the straightforward stolon node transformation method could be well used to genetically modify turfgrass. The obtained transgenic line might be as genetic resource for breeding program and practiced to grow in cold temperate zones.

PicW2 from Picea wilsonii: preparation, purification, crystallization and X-ray diffraction analysis.

Low temperature is a major limiting factor for plant growth and development. Dehydrin proteins are generally induced in response to low-temperature stress. In previous research, a full-length dehydrin gene, PicW2, was isolated from Picea wilsonii and its expression was associated with hardiness to cold. In order to gain insight into the mechanism of low-temperature tolerance by studying its three-dimensional crystal structure, prokaryotically expressed PicW2 dehydrin protein was purified using chitosan-affinity chromatography and gel filtration, and crystallized using the vapour-diffusion method. The crystal grew in a condition consisting of 0.1 M HEPES pH 8.0, 25%(w/v) PEG 3350 using 4 mg ml-1 protein solution at 289 K. X-ray diffraction data were collected from a crystal at 100 K to 2.82 Šresolution. The crystal belonged to space group C121, with unit-cell parameters a = 121.55, b = 33.26, c = 73.39 Å, α = γ = 90.00, ß = 109.01°. The asymmetric unit contained one molecule of the protein, with a corresponding Matthews coefficient of 2.87 Å3 Da-1 and a solvent content of 57.20%. Owing to a lack of structures of homologous dehydrin proteins, molecular-replacement trials failed. Data collection for selenium derivatization of PicW2 and crystal structure determination is currently in progress.

Correction: Transcriptome analysis of Phytolacca americana L. in response to cadmium stress.

[This corrects the article DOI: 10.1371/journal.pone.0184681.].

WHITE PANICLE3, a Novel Nucleus-Encoded Mitochondrial Protein, Is Essential for Proper Development and Maintenance of Chloroplasts and Mitochondria in Rice.

Mitochondria and chloroplasts are interacting organelles that play important roles in plant development. In addition to a small number proteins encoded by their own genomes, the majority of mitochondrial and chloroplast proteins are encoded in the cell nucleus and imported into the organelle. As a consequence, coordination between mitochondria, chloroplasts, and the nucleus is of crucial importance to plant cells. Variegated mutants are chloroplast-defective mutants and are considered to be ideal models for studying the intercommunication between these organelles. Here, we report the isolation of WHITE PANICLE3 (WP3), a nuclear gene involved in variegation, from a naturally occurring white panicle rice mutant. Disrupted expression of WP3 in the mutant leads to severe developmental defects in both chloroplasts and mitochondria, and consequently causes the appearance of white-striped leaves and white panicles in the mutant plants. Further investigation showed that WP3 encodes a protein most likely targeted to mitochondria and is specifically expressed in rice panicles. Interestingly, we demonstrate that the recessive white-panicle phenotype in the wp3 mutant is inherited in a typical Mendelian manner, while the white-striped leaf phenotype in wp3 is maternally inherited. Our data collectively suggest that the nucleus-encoded mitochondrial protein, WP3, plays an essential role in the regulation of chloroplast development in rice panicles by maintaining functional mitochondria. Therefore, the wp3 mutant is an excellent model in which to explore the communication between the nucleus, mitochondria, and chloroplasts in plant cells.

Over-expression of the poplar expansin gene PtoEXPA12 in tobacco plants enhanced cadmium accumulation.

As important cell wall proteins in plants, expansins are involved in a serious of abiotic stress resistance such as drought, heat, salt, even heavy metals. To understand the role of expansins in cadmium (Cd) stress, we analyzed the expression patterns of 36 expansin genes in Populus tomentosa. A Cd-induced expansin gene, PtoEXPA12, was identified, cloned, and transformed into tobacco plants. After treatment with Cd, the transgenic plants showed stronger symptoms of Cd toxicity as to the wild-type tobacco plants. Further physiological tests showed that the transformants had higher relative electrolyte leakage and superoxide dismutase activity, more malondialdehyde and H2O2 content, and lower chlorophyll content in Cd stress. Cd content measurement showed it is 1.40-2.07-fold higher and 1.29-1.38-fold higher separately in roots and shoots of transgenic plants than those in wild-type plants, while the transfer coefficient value kept invariably even decreased. Therefore, PtoEXPA12 was really involved in Cd uptake and accumulation, and led to Cd toxicity of cells. It would be a potentially applicable part in phytoremediation system.

Transgenic tobacco plants expressing a P1B-ATPase gene from Populus tomentosa Carr. (PtoHMA5) demonstrate improved cadmium transport.

Heavy metal ATPase (HMA) plays an important role in phytoremediation via long-distance transportation from root to shoot. In this report, we identified a heavy metal ATPase gene, PtoHMA5, from Populus tomentosa Carr. Its encoded peptide consists of 967 amino acids and has eight trans-membrane motifs inside. Tobacco plants were transformed with this gene via Agrobacterium tumefaciens-mediated method. After exposure to 50mg/LCdCl2 for 10d, the transgenic lines displayed higher cadmium accumulation in leaves than did the wild-type plants with an absolute increase of 25.04%, while the transfer coefficient increased by 16.01%-43.25%. Physiological testing including assessment of relative electrolytic leakage (REL), malondialdehyde (MDA) content, and chlorophyll content revealed that the transgenic lines were seriously affected when compared with the wild-type plants. In summary, PtoHMA5 is really involved in cadmium transport from root to shoot but is not associated with the removal of cadmium toxicity.

Diversity in cadmium accumulation and resistance associated with various metallothionein genes (type III) in Phytolacca americana L.

Metallothioneins (MTs) are known for their heavy metal deoxidation during phytoremediation. To estimate their roles in the cadmium (Cd) hyperaccumulator Phytolacca americana L., three MT genes, PaMT3-1, PaMT3-2 and PaMT3-3, belonging to the MT3 subfamily were cloned. They separately encoded 63, 65 and 65 amino acids, containing12, 10 and 11 cysteines (Cys), respectively. Each gene was individually transformed and expressed in Escherichia coli cells. A Cd-resistance assay showed that the recombinant strains had enhanced survival rates, especially those containing PaMT3-1 and PaMT3-3. Additionally, the recombinant strains were high Cd accumulators, with the recombinant PaMT3-1's maximum accumulation being 2.16 times that of the empty vector strains. The numbers of cysteines and the structures of MT proteins were associated with the Cd enrichment and resistance capabilities. PaMT3-1 could be an effective gene resource in future plant Cd remediation-related breeding programs.