Learning the cellular activity representation based on gene regulatory networks for prediction of tumor response to drugs.

Predicting the response of tumor cells to anti-tumor drugs is critical to realizing cancer precision medicine. Currently, most existing methods ignore the regulatory relationships between genes and thus have unsatisfactory predictive performance. In this paper, we propose to predict anti-tumor drug efficacy via learning the activity representation of tumor cells based on a priori knowledge of gene regulation networks (GRNs). Specifically, the method simulates the cellular biosystem by synthesizing a cell-gene activity network and then infers a new low-dimensional activity representation for tumor cells from the raw high-dimensional expression profile. The simulated cell-gene network mainly comprises known gene regulatory networks collected from multiple resources and fuses tumor cells by linking them to hotspot genes that are over- or under-expressed in them. The resulting activity representation could not only reflect the shallow expression profile (hotspot genes) but also mines in-depth information of gene regulation activity in tumor cells before treatment. Finally, we build deep learning models on the activity representation for predicting drug efficacy in tumor cells. Experimental results on the benchmark GDSC dataset demonstrate the superior performance of the proposed method over SOTA methods with the highest AUC of 0.954 in the efficacy label prediction and the best R2 of 0.834 in the regression of half maximal inhibitory concentration (IC50) values, suggesting the potential value of the proposed method in practice.

Parameter-based analysis of clinical efficacy of combined bed nucleus of the stria terminalis-nucleus accumbens deep brain stimulation for treatment-resistant depression.

OBJECTIVE: Treatment-resistant depression (TRD) is a severely disabling psychiatric condition that responds poorly to conventional treatments. Deep brain stimulation (DBS) has been proposed for the treatment of patients with TRD in numerous studies. Several deep brain nuclei are considered as potential targets for TRD-DBS, but their clinical efficacy needs further validation. This study carried out dual-target combined stimulation of the bed nucleus of the stria terminalis (BNST) and nucleus accumbens (NAc) to investigate the effectiveness of the treatment for TRD patients. METHODS: An 8-contact DBS electrode was used in the study with a surgical path that crossed the BNST and NAc targets. Stimulation parameters and the corresponding severity of symptoms evaluated by the 17-item Hamilton Depression Rating Scale (HAMD-17) and other scales were obtained at each follow-up. The accuracy of electrode positions, the effect of combined stimulation, and the corresponding stimulation parameters were evaluated. Sweet spot prediction models were used to assess the effective stimulation sites in the treatment. RESULTS: The study included 23 TRD patients undergoing DBS at a single center from March 2021 to May 2023. At the last follow-up (range 4-24 months), 14 patients had responded to the treatment (HAMD-17 score improved ≥ 50%), 7 of whom had achieved clinical remission (HAMD-17 score ≤ 7). Electrode position analysis suggested that the BNST may be more important for the improvement of depressive symptoms than the NAc. Overlapped volumes of volume of tissue activated (VTA) and BNST were significantly correlated with absolute (ρleft = -0.377, p < 0.001; ρright = -0.251, p < 0.001) and percent (ρleft = -0.249, p < 0.001; ρright = -0.098, p = 0.102) changes in HAMD-17 score. The sweet spot model of HAMD-17 improvement also suggested that the VTA overlap with the dorsal side of BNST was associated with the impact on depressive symptoms (t = -4.10, p < 0.05). CONCLUSIONS: Combined BNST-NAc stimulation of TRD can effectively improve depressive symptoms, in which the BNST seems to have a dominant therapeutic effect. The results of this study not only help to optimize the DBS programming parameters, but also offer an opportunity to further understand the differences between the two targets. In the future, larger prospective cohorts are needed to verify the results of combined BNST-NAc DBS.

Comparative transcriptome profiling reveals the importance of GmSWEET15 in soybean susceptibility to Sclerotinia sclerotiorum.

Soybean sclerotinia stem rot (SSR) is a disease caused by Sclerotinia sclerotiorum that causes incalculable losses in soybean yield each year. Considering the lack of effective resistance resources and the elusive resistance mechanisms, we are urged to develop resistance genes and explore their molecular mechanisms. Here, we found that loss of GmSWEET15 enhanced the resistance to S. sclerotiorum, and we explored the molecular mechanisms by which gmsweet15 mutant exhibit enhanced resistance to S. sclerotiorum by comparing transcriptome. At the early stage of inoculation, the wild type (WT) showed moderate defense response, whereas gmsweet15 mutant exhibited more extensive and intense transcription reprogramming. The gmsweet15 mutant enriched more biological processes, including the secretory pathway and tetrapyrrole metabolism, and it showed stronger changes in defense response, protein ubiquitination, MAPK signaling pathway-plant, plant-pathogen interaction, phenylpropanoid biosynthesis, and photosynthesis. The more intense and abundant transcriptional reprogramming of gmsweet15 mutant may explain how it effectively delayed colonization by S. sclerotiorum. In addition, we identified common and specific differentially expressed genes between WT and gmsweet15 mutant after inoculation with S. sclerotiorum, and gene sets and genes related to gmsweet15_24 h were identified through Gene Set Enrichment Analysis. Moreover, we constructed the protein-protein interaction network and gene co-expression networks and identified several groups of regulatory networks of gmsweet15 mutant in response to S. sclerotiorum, which will be helpful for the discovery of candidate functional genes. Taken together, our results elucidate molecular mechanisms of delayed colonization by S. sclerotiorum after loss of GmSWEET15 in soybean, and we propose novel resources for improving resistance to SSR.

Structural and functional correlates of the response to deep brain stimulation at ventral capsule/ventral striatum region for treatment-resistant depression.

BACKGROUND: Though deep brain stimulation (DBS) shows increasing potential in treatment-resistant depression (TRD), the underlying neural mechanisms remain unclear. Here, we investigated functional and structural connectivities related to and predictive of clinical effectiveness of DBS at ventral capsule/ventral striatum region for TRD. METHODS: Stimulation effects of 71 stimulation settings in 10 TRD patients were assessed. The electric fields were estimated and combined with normative functional and structural connectomes to identify connections as well as fibre tracts beneficial for outcome. We calculated stimulation-dependent optimal connectivity and constructed models to predict outcome. Leave-one-out cross-validation was used to validate the prediction value. RESULTS: Successful prediction of antidepressant effectiveness in out-of-sample patients was achieved by the optimal connectivity profiles constructed with both the functional connectivity (R=0.49 at p<10-4; deviated by 14.4±10.9% from actual, p<0.001) and structural connectivity (R=0.51 at p<10-5; deviated by 15.2±11.5% from actual, p<10-5). Frontothalamic pathways and cortical projections were delineated for optimal clinical outcome. Similarity estimates between optimal connectivity profile from one modality (functional/structural) and individual brain connectivity in the other modality (structural/functional) significantly cross-predicted the outcome of DBS. The optimal structural and functional connectivity mainly converged at the ventral and dorsal lateral prefrontal cortex and orbitofrontal cortex. CONCLUSIONS: Connectivity profiles and fibre tracts following frontothalamic streamlines appear to predict outcome of DBS for TRD. The findings shed light on the neural pathways in depression and may be used to guide both presurgical planning and postsurgical programming after further validation.

The SsAtg1 Activating Autophagy Is Required for Sclerotia Formation and Pathogenicity in Sclerotinia sclerotiorum.

Sclerotinia sclerotiorum is a necrotrophic phytopathogenic fungus that produces sclerotia. Sclerotia are essential components of the survival and disease cycle of this devastating pathogen. In this study, we analyzed comparative transcriptomics of hyphae and sclerotia. A total of 1959 differentially expressed genes, 919 down-regulated and 1040 up-regulated, were identified. Transcriptomes data provide the possibility to precisely comprehend the sclerotia development. We further analyzed the differentially expressed genes (DEGs) in sclerotia to explore the molecular mechanism of sclerotia development, which include ribosome biogenesis and translation, melanin biosynthesis, autophagy and reactivate oxygen metabolism. Among these, the autophagy-related gene SsAtg1 was up-regulated in sclerotia. Atg1 homologs play critical roles in autophagy, a ubiquitous and evolutionarily highly conserved cellular mechanism for turnover of intracellular materials in eukaryotes. Therefore, we investigated the function of SsAtg1 to explore the function of the autophagy pathway in S. sclerotiorum. Deficiency of SsAtg1 inhibited autophagosome accumulation in the vacuoles of nitrogen-starved cells. Notably, ΔSsAtg1 was unable to form sclerotia and displayed defects in vegetative growth under conditions of nutrient restriction. Furthermore, the development and penetration of the compound appressoria in ΔSsAtg1 was abnormal. Pathogenicity analysis showed that SsAtg1 was required for full virulence of S. sclerotiorum. Taken together, these results indicate that SsAtg1 is a core autophagy-related gene that has vital functions in nutrient utilization, sclerotia development and pathogenicity in S. sclerotiorum.

Does economic agglomeration improve agricultural green total factor productivity? Evidence from China's Yangtze river delta.

Economic agglomeration plays an important role in China's social transformation process of industry feeding agriculture and urban supporting rural areas, and is one of the core weapons to promote agricultural economic growth and green and efficient development. Based on panel data of 41 cities in the Yangtze River Delta (YRD) from 2010 to 2020, this paper constructs an "environment-resource-energy-economy" agricultural input-output system, taking into account carbon emissions and surface pollution, and provides a more comprehensive accounting of agricultural green total factor productivity (AGTFP), the non-linear effects of economic agglomeration on AGTFP and shock responses were empirically analyzed using a panel threshold model and a panel vector autoregression (VAR), respectively. The findings show that: (1) during the period 2010-2020, the AGTFP in the YRD showed an overall rising trend with regional spatial agglomeration characteristics. (2) Economic agglomeration has a triple threshold effect on AGTFP, which was a weak facilitative effect in the early stage, inhibited by the siphoning effect of resource loss and arable land fragmentation in the growth stage, promoted by the radiation effect of external increasing return to scale in the form of sharing, matching and learning in the maturity stage, and finally tends to decline. (3) The shock response of economic agglomeration to AGTFP showed a continuous positive shock, peaking in the first period and then gradually converging to zero. (4) The heterogeneity analysis demonstrated that economic agglomeration has a considerable impact on boosting AGTFP in non-metropolitan areas and cities on the outskirts of YRD. In the future, China should effectively play a positive role in economic agglomeration on AGTFP and enhance the mutual coordination of economic agglomeration and agricultural green development in the process of urban cluster economic growth.

A systematic review of brain morphometry related to deep brain stimulation outcome in Parkinson's disease.

While the efficacy of deep brain stimulation (DBS) is well-established in Parkinson's Disease (PD), the benefit of DBS varies across patients. Using imaging features for outcome prediction offers potential in improving effectiveness, whereas the value of presurgical brain morphometry, derived from the routinely used imaging modality in surgical planning, remains under-explored. This review provides a comprehensive investigation of links between DBS outcomes and brain morphometry features in PD. We systematically searched PubMed and Embase databases and retrieved 793 articles, of which 25 met inclusion criteria and were reviewed in detail. A majority of studies (24/25), including 1253 of 1316 patients, focused on the outcome of DBS targeting the subthalamic nucleus (STN), while five studies included 57 patients receiving globus pallidus internus (GPi) DBS. Accumulated evidence showed that the atrophy of motor cortex and thalamus were associated with poor motor improvement, other structures such as the lateral-occipital cortex and anterior cingulate were also reported to correlated with motor outcome. Regarding non-motor outcomes, decreased volume of the hippocampus was reported to correlate with poor cognitive outcomes. Structures such as the thalamus, nucleus accumbens, and nucleus of basalis of Meynert were also reported to correlate with cognitive functions. Caudal middle frontal cortex was reported to have an impact on postsurgical psychiatric changes. Collectively, the findings of this review emphasize the utility of brain morphometry in outcome prediction of DBS for PD. Future efforts are needed to validate the findings and demonstrate the feasibility of brain morphometry in larger cohorts.

Predicting the onset of freezing of gait in Parkinson's disease.

BACKGROUND: Freezing of gait is a debilitating symptom of Parkinson's disease associated with high risks of falls and poor quality of life. While productive therapy for FoG is still underway, early prediction of FoG could help high-risk PD patients to take preventive measures. In this study, we predicted the onset of FoG in de novo PD patients using a battery of risk factors from patients enrolled in PPMI cohort. METHODS: Baseline characteristics were compared between subjects who developed FoG (68 patients, 37.2%, pre-FoG group) during the five-year follow up and subjects who did not (115 patients, 62.8%, non-FoG group). A multivariate logistic regression model was built based on backward stepwise selection of factors that were associated with FoG onset in the univariate analysis. ROC curves were used to assess sensitivity and specificity of the predictive model. RESULTS: At baseline, age, PIGD score, cognitive functions, autonomic functions, sleep behavior, fatigue and striatal DAT uptake were significantly different in the pre-FoG group relative to the non-FoG group. However, there was no difference in genetic characteristics between the two patient sets. Univariate analysis showed several motor and non-motor factors that correlated with FoG, including PIGD score, MDS-UPDRS part II score, SDMT score, HVLT Immediate/Total Recall, MOCA, Epworth Sleepiness Scale, fatigue, SCOPA-AUT gastrointestinal score, SCOPA-AUT urinary score and CSF biomarker Abeta42. Multivariate logistic analysis stressed that high PIGD score, fatigue, worse SDMT performance and low levels of Abeta42 were independent risk factors for FoG onset in PD patients. CONCLUSIONS: Combining motor and non-motor features including PIGD score, poor cognitive functions and CSF Abeta can identify PD patients with high risk of FoG onset.

Efficacy, safety and prognostic factors of camrelizumab plus carboplatin and pemetrexed chemotherapy in advanced lung adenocarcinoma patients.

WHAT IS KNOWN AND OBJECTIVE: Camrelizumab is a recently developed PD-1 inhibitor in China applied in treating different cancers including lung cancer. This study is designed to evaluate the efficacy, safety and prognostic factors for camrelizumab plus carboplatin and pemetrexed (CP) chemotherapy in treating patients with advanced lung adenocarcinoma. METHODS: Of 51 advanced lung adenocarcinoma patients with negative driver genes who received camrelizumab plus CP chemotherapy were recruited. These patients received four cycles of camrelizumab plus CP chemotherapy in a 21-day cycle. Then, camrelizumab, pemetrexed or camrelizumab plus pemetrexed was administered as maintenance therapy. RESULTS AND DISCUSSION: The rates of complete response, partial response, stable disease and progressive disease were 2.0%, 56.8%, 19.6% and 5.9%, respectively; while treatment response of 15.7% of patients was missing or not evaluable. The objective response and disease control rates were 58.8% and 78.4%, respectively. With a median follow-up period of 14.9 months (the follow-up duration ranged from 3.9 months to 24.3 months), 41 (83.4%) cases of disease progression and 22 (43.1%) cases of death were recorded. The median progression-free survival (PFS) was 10.5 months (95% confidence interval (CI): 8.4-12.6 months) with a 1-year PFS rate of 36.3% and a 2-year PFS rate of 7.5%. In addition, the median overall survival (OS) was 18.7 months (95% CI: 16.4-21.0 months) with a 1-year OS rate of 79.1% and a 2-year OS rate of 30.4%. In consideration of safety, the most frequent adverse events were peripheral neuropathy (37.3%), neutropenia (37.3%), alopecia (35.3%), etc. and most of them were grade 1-2 and could be controlled. WHAT IS NEW AND CONCLUSION: Camrelizumab plus CP chemotherapy achieves favourable efficacy and tolerable adverse events in advanced lung adenocarcinoma patients.

Impact of Dairy Imports on Raw Milk Production Technology Progress in China.

China's dairy product import volume and output continue to grow rapidly, and to a certain extent, it will form a substitute for the Chinese dairy market. Therefore, it is necessary to study the impact of the import of dairy products on the technological progress of raw milk production in China. Using the data from 2005 to 2017, this paper uses the DEA model and the input-output model to analyze the impact of China's dairy product imports on the technological progress of raw milk production. The model results show that: (1) there are differences in the technological content of dairy products from different importing countries; (2) The total technological content of imported dairy products hinders the improvement of the technological progress index of small, medium and large-scale production of raw milk in China, and has the most prominent negative impact on the technological progress of large-scale raw milk production in China; (3) The technological content of dairy products imports from New Zealand, Australia, Germany, the Netherlands and other countries can help improve the technological progress index of China's moderate-scale production of raw milk, while importing countries from the United States, Canada and other countries hinder it.

Gene-gene interaction and new onset of major depressive disorder: Findings from a Chinese freshmen nested case-control study.

BACKGROUND: The gene-gene interaction is known to be the genetic cause of major depressive disorder (MDD). Several genes have been found to be related to MDD. The objectives of this study were to verify the susceptibility genes of MDD in a sample of university students in China, and to investigate possible gene-gene interactions in relation to the risk of MDD. METHODS: 7,627 Chinese Han freshmen were enrolled at baseline survey in 2018. After a 2-year follow-up, 170 new onset MDD cases and 680 controls with DNA samples reserved were sequenced and genotyped for 4 selected Single Nucleotide Polymorphisms (SNPs) in a nested case-control study (ratio of 1:4). Chi-square test was used to identify the relationships between SNPs and risk of MDD. Generalized multifactor dimensionality reduction (GMDR) was used to analyze the gene-gene interactions. RESULTS: The 2-year incidence of MDD in Chinese college students was 3.75% (95% CI: 3.24%, 4.34%). There was no statistical difference in MDD incidences between males (3.74%, 95% CI: 3.12%, 4.49%) and females (3.77%, 95% CI: 2.97%, 4.78%) (p>0.05). TMEM161B (rs768705) was positively associated with new onset MDD (χ2 = 0.75, p = 0.023). The AG genotype of rs768705 was significant (OR=1.640, 95%CI:1.414-2.358). The gene-gene interaction between TMEM161B (rs768705) and LHPP (rs35936514) was statistically significant in this nested case-control study (p = 0.011). The CV consistency was 9/10 and the testing accuracy was 0.5274. LIMITATIONS: The results could not be inferred to other ethnics. CONCLUSIONS: This study provided evidence that combined rs768705 (TMEM161B) and rs35936514 (LHPP) may modulate the risk of MDD.

RCAN1 Inhibits BACE2 Turnover by Attenuating Proteasome-Mediated BACE2 Degradation.

Amyloid-ß protein (Aß) is the main component of neuritic plaques, the pathological hallmark of Alzheimer's disease (AD). ß-site APP cleaving enzyme 1 (BACE1) is a major ß-secretase contributing to Aß generation. ß-site APP cleaving enzyme 2 (BACE2), the homolog of BACE1, is not only a θ-secretase but also a conditional ß-secretase. Previous studies showed that regulator of calcineurin 1 (RCAN1) is markedly increased by AD and promotes BACE1 expression. However, the role of RCAN1 in BACE2 regulation remains elusive. Here, we showed that RCAN1 increases BACE2 protein levels. Moreover, RCAN1 inhibits the turnover of BACE2 protein. Furthermore, RCAN1 attenuates proteasome-mediated BACE2 degradation, but not lysosome-mediated BACE2 degradation. Taken together, our work indicates that RCAN1 inhibits BACE2 turnover by attenuating proteasome-mediated BACE2 degradation. It advances our understanding of BACE2 regulation and provides a potential mechanism of BACE2 dysregulation in AD.

Site-specific proteolytic cleavage prevents ubiquitination and degradation of human REV3L, the catalytic subunit of DNA polymerase ζ.

REV3L, the catalytic subunit of DNA polymerase ζ (Pol ζ), is indispensable for translesion DNA synthesis, which protects cells from deleterious DNA lesions resulting from various intrinsic and environmental sources. However, REV3L lacks a proofreading exonuclease activity and consequently bypasses DNA lesions at the expense of increased mutations, which poses a severe threat to genome stability. Here we report a site-specific proteolytic event of human REV3L. We show that REV3L is cleaved by a threonine aspartase, Taspase1 (TASP1), to generate an N-terminal 70-kDa fragment (N70) and a polypeptide carrying the C-terminal polymerase catalytic domain in human cells. Strikingly, such a post-translational cleavage event plays a vital role in controlling REV3L stability by preventing ubiquitination and proteasome-mediated degradation of REV3L. Indicative of the biological importance of the above REV3L post-translational processing, cellular responses to UV and cisplatin-induced DNA lesions are markedly impaired in human HCT116 cell derivatives bearing defined point mutations in the endogenous REV3L gene that compromise REV3L cleavage. These findings establish a new paradigm in modulating the abundance of REV3L through site-specific proteolysis in human cells.

Inhibition of biofilm formation and exopolysaccharide synthesis of Enterococcus faecalis by phenyllactic acid.

This study aimed to evaluate the inhibitory activity of phenyllactic acid (PLA) against the biofilm formation of Enterococcus faecalis and to explore its potential molecular mechanism. The MIC value of PLA that inhibited the growth of E. faecalis R612-Z1 in BHI broth was 5 mg/mL. PLAs at subinhibitory concentrations of 1.25 and 2.50 mg/mL were found to inhibit biofilm formation by a crystal violet staining assay. The cell swimming and swarming motilities of E. faecalis were reduced in the presence of PLA. An apparent decrease in the thickness of PLA-treated biofilms was observed through confocal laser scanning microscopy analysis. The exopolysaccharide production in E. faecalis biofilms was inhibited by EPS quantification assay and scanning electron microscopy (SEM). qRT-PCR analyses showed that PLA down-regulated the transcription of Ebp pili genes (ebpABC) and Epa polysaccharide genes (epaABE). PLA inhibited the biofilm formation by interfering with cell mobility and EPS production of E. faecalis. In addition, PLA at concentrations of 10.0 mg/mL can effectively control the bacterial cells in a three-day-old mature biofilm of E. faecalis grown on 24-well flat-bottom polystyrene plates and stainless-steel surfaces. Thus, PLA is potentially an effective agent to control E. faecalis biofilms.

The nature of small molecules adsorbed on defective carbon nanotubes.

In this work, we perform a comprehensive theoretical study on adsorption of representative 10-electron molecules H2O, CH4 and NH3 onto defective single-walled carbon nanotubes. Results of adsorption energy and charge transfer reveal the existence of both chemical adsorption (CA) and physical adsorption (PA). While PA processes are common for all molecules, CA could be further achieved by the polar molecule NH3, whose lone-pair electrons makes it easier to be bonded with the defective nanotube. Our systematic work could contribute to the understanding on intermolecular interactions and the design of future molecular detectors.

UBA6 and Its Bispecific Pathways for Ubiquitin and FAT10.

Questions have been raised since the discovery of UBA6 and its significant coexistence with UBE1 in the ubiquitin-proteasome system (UPS). The facts that UBA6 has the dedicated E2 enzyme USE1 and the E1-E2 cascade can activate and transfer both ubiquitin and ubiquitin-like protein FAT10 have attracted a great deal of attention to the regulational mechanisms of the UBA6-USE1 cascade and to how FAT10 and ubiquitin differentiate with each other. This review recapitulates the latest advances in UBA6 and its bispecific UBA6-USE1 pathways for both ubiquitin and FAT10. The intricate networks of UBA6 and its interplays with ubiquitin and FAT10 are briefly reviewed, as are their individual and collective functions in diverse physiological conditions.

Antimicrobial Activity of Phenyllactic Acid Against Enterococcus faecalis and Its Effect on Cell Membrane.

3-Phenyllactic acid (PLA) was reported to have an effective antimicrobial activity. This study evaluated the antimicrobial activity of PLA against foodborne Enterococcus faecalis and its effect on cell membrane. The minimum concentration of PLA to inactivate E. faecalis in brain heart infusion broth was 5 mg/mL. PLA solutions of 5 and 10 mg/mL can inactivate E. faecalis population ≥6 log CFU/mL within 60 and 30 min, respectively. The cell membranes of most E. faecalis cells were damaged after PLA treatment according to the images of scanning electron microscopy and transmission electron microscopy. The differences in the regions of cell membrane protein, fatty acid, and polysaccharide were revealed by Fourier transform infrared spectroscopy, which further indicated cell membrane damages. The cell membrane permeability was increased when the concentration of PLA treatment was increased in the membrane permeability assays. Finally, almost all bacterial cells were damaged after treatment with 10 mg/mL PLA for 30 min, further confirmed by flow cytometry analysis. This study concluded that PLA is effective in inactivating E. faecalis cells through the leakage of intracellular components caused by cell membrane damage.

Molecular Characterization of the Complete Genome of Three Basal-BR Isolates of Turnip mosaic virus Infecting Raphanus sativus in China.

Turnip mosaic virus (TuMV) infects crops of plant species in the family Brassicaceae worldwide. TuMV isolates were clustered to five lineages corresponding to basal-B, basal-BR, Asian-BR, world-B and OMs. Here, we determined the complete genome sequences of three TuMV basal-BR isolates infecting radish from Shandong and Jilin Provinces in China. Their genomes were all composed of 9833 nucleotides, excluding the 3'-terminal poly(A) tail. They contained two open reading frames (ORFs), with the large one encoding a polyprotein of 3164 amino acids and the small overlapping ORF encoding a PIPO protein of 61 amino acids, which contained the typically conserved motifs found in members of the genus Potyvirus. In pairwise comparison with 30 other TuMV genome sequences, these three isolates shared their highest identities with isolates from Eurasian countries (Germany, Italy, Turkey and China). Recombination analysis showed that the three isolates in this study had no "clear" recombination. The analyses of conserved amino acids changed between groups showed that the codons in the TuMV out group (OGp) and OMs group were the same at three codon sites (852, 1006, 1548), and the other TuMV groups (basal-B, basal-BR, Asian-BR, world-B) were different. This pattern suggests that the codon in the OMs progenitor did not change but that in the other TuMV groups the progenitor sequence did change at divergence. Genetic diversity analyses indicate that the PIPO gene was under the highest selection pressure and the selection pressure on P3N-PIPO and P3 was almost the same. It suggests that most of the selection pressure on P3 was probably imposed through P3N-PIPO.

Functional Analysis of the Maize C-Repeat/DRE Motif-Binding Transcription Factor CBF3 Promoter in Response to Abiotic Stress.

The ZmCBF3 gene is a member of AP2/ERF transcription factor family, which is a large family of plant-specific transcription factors that share a well-conserved DNA-binding domain. To understand the regulatory mechanism of ZmCBF3 gene expression, we isolated and characterized the ZmCBF3 promoter (PZmCBF3). Three deletion fragments of PZmCBF3 were generated, C1-C3, from the translation start codon at position -1079, -638, and -234, and fused to the GUS reporter gene. Each deletion construct was analyzed by Agrobacterium-mediated stable transformation and expression in Arabidopsis thaliana. GUS expression assays indicated that the PZmCBF3 exhibited root-specific expression activity. A 234-bp fragment upstream of the ZmCBF3 gene conferred a high level of GUS activity in Arabidopsis. Some cis-acting elements involved in the down-regulation of gene expression were detected in the promoter, encompassing positions -1079 to -234. PZmCBF3 was activated by cold stress. The MYCCONSENSUSAT elements (CANNTG) were responsible for the ability of PZmCBF3 to respond to cold stress. The results of the present study suggest that PZmCBF3 might play a role in cold tolerance in maize.