Osteosarcoma neutrophil extracellular trap network-associated gene recurrence and metastasis model.

Osteosarcoma (OS) is the most common malignancy in children and adolescents and has a high probability of recurrence and metastasis. A growing number of studies have shown that neutrophil extracellular traps (NETs) are strongly associated with cancer metastasis, but in osteosarcoma, genes associated with NETs that promote osteosarcoma recurrence and metastasis remain to be explored. We systematically investigated the gene expression patterns of NETs in OS samples from the GEO database. NETs molecular typing was evaluated based on NETs expression profiles, and the association between NETs molecular subtypes and immune microenvironment and metastatic features were explored. Ultimately, we constructed a signature model and column line graph associated with metastasis prediction and screened possible potential drugs for metastatic osteosarcoma. We established two different molecular subtypes of NETs, which showed significant differences in metastatic status, metastasis time, tumor immune microenvironment, and biological effects. We also constructed a NETs-related gene metastasis signature(NRGMS) to assess the expression pattern of NETs in patients to predict metastatic recurrence in osteosarcoma patients. We screened for TOMM40 and FH associated with metastatic recurrence in osteosarcoma patients. Overall, this study constructs a predictive model for osteosarcoma metastasis of NETs-related genes, which is expected to provide new insights into the metastasis of osteosarcoma.

A Positive Feedback Loop of E2F4-Mediated Activation of MNX1 Regulates Tumour Progression in Colorectal Cancer.

Purpose: Colorectal cancer (CRC) is the 3rd most prevalent malignant tumour globally. Although significant strides have been made in diagnosis and treatment, its prognosis at the moment remains unpromising. Therefore, there is an urgent and desperate need to identify novel biomarkers of CRC and evaluate its mechanism of tumourigenesis and development. Methods: JASPAR and RNAinter databases are used to analyze target genes associated with colorectal cancer. Western blotting, q-PCR and immunohistochemistry et, al. were used to detect the level of MNX1 in patients with colorectal cancer, and Chip-PCR was used to detect the targeted binding ability of E2F4 and MNX1. The cells and animal models overexpressed MNX1 and E2F4 were constructed by shRNA transfection. Results: Herein, MNX1 was highly expressed and linked to favourable overall survival curves in colorectal cancer. The functional assay revealed that MNX1 overexpression could promote proliferation, migration, and invasion of CRC cells. Based on the prediction of the JASPAR and RNAinter databases, the transcription factor, E2F4, was bound to the MNX1 promoter region. The Chromatin Immunoprecipitation (ChIP) assay verified the interactions between MNX1 and E2F4 in CRC. Additionally, we found that sh-E2F4 markedly downregulated the MNX1 levels and reduced CRC progression in vivo and in vitro, which reversed MNX1 overexpression. Conclusion: Therefore, our research discovered that E2F4-mediated abnormal MNX1 expression promotes CRC progression and could become a novel diagnostic or therapeutic target of CRC.

BMP6 participates in the pathogenesis of adolescent idiopathic scoliosis by regulating osteopenia.

Adolescent idiopathic scoliosis (AIS) is a complex disease characterized by three-dimensional structural deformities of the spine. Its pathogenesis is associated with osteopenia. Bone-marrow-derived mesenchymal stem cells (BMSCs) play an important role in bone metabolism. We detected 1919 differentially expressed mRNAs and 744 differentially expressed lncRNAs in BMSCs from seven patients with AIS and five patients without AIS via high-throughput sequencing. Multiple analyses identified bone morphogenetic protein-6 (BMP6) as a hub gene that regulates the abnormal osteogenic differentiation of BMSCs in AIS. BMP6 expression was found to be decreased in AIS and its knockdown in human BMSCs significantly altered the degree of osteogenic differentiation. Additionally, CAP1-217 has been shown to be a potential upstream regulatory molecule of BMP6. We showed that CAP1-217 knockdown downregulated the expression of BMP6 and the osteogenic differentiation of BMSCs. Simultaneously, knockout of BMP6 in zebrafish embryos significantly increased the deformity rate. The findings of this study suggest that BMP6 is a key gene that regulates the abnormal osteogenic differentiation of BMSCs in AIS via the CAP1-217/BMP6/RUNX2 axis.

Research progress on the regulation of bone marrow stem cells by noncoding RNAs in adolescent idiopathic scoliosis.

Adolescent idiopathic scoliosis (AIS) is a common spinal deformity in young women, but its pathogenesis remains unclear. The primary pathogenic factors contributing to its development include genetics, abnormal bone metabolism, and endocrine factors. Bone marrow stem cells (BMSCs) play a crucial role in the pathogenesis of AIS by regulating its occurrence and progression. Noncoding RNAs (ncRNAs) are also involved in the pathogenesis of AIS, and their role in regulating BMSCs in patients with AIS requires further evaluation. In this review, we discuss the relevant literature regarding the osteogenic, chondrogenic, and lipogenic differentiation of BMSCs. The corresponding mechanisms of ncRNA-mediated BMSC regulation in patients with AIS, recent advancements in AIS and ncRNA research, and the importance of ncRNA translation profiling and multiomics are highlighted.

PECAM1 plays a role in the pathogenesis and treatment of bone metastases.

Bone is the third most common metastatic site for all primary tumors, the common primary focus of bone metastases include breast cancer, prostate cancer, and so on. And the median survival time of patients with bone metastases is only 2-3 years. Therefore, it is urgent to develop new targets to diagnose and treat bone metastases. Based on two data sets GSE146661 and GSE77930 associated with bone metastases, it was found that 209 genes differentially expressed in bone metastases group and control group. PECAM1 was selected as hub-gene for the follow-up research after constructing protein-protein interaction (PPI) network and enrichment analysis. Moreover, q-PCR analysis verified that the expression of PECAM1 decreased in bone metastatic tumor tissues. PECAM1 was believed to be possibly related to the function of osteoclasts, we knocked down the expression of PECAM1 with shRNA in lymphocytes extracted from bone marrow nailed blood. The results indicated that sh-PECAM1 treatment could promote osteoclast differentiation, and the sh-PECAM1-treated osteoclast culture medium could significantly promote the proliferation and migration of tumor cells. These results suggested that PECAM1 may be a potential biomarker for the diagnosis and treatment of bone metastases of tumor.

Metabolic syndrome and metastatic prostate cancer correlation study, a real-world study in a prostate cancer clinical research center, Xinjiang, China.

Objective: The aim of this study was to investigate the relevance of metabolic syndrome (MetS) and metabolic scores to the occurrence, progression and prognosis of metastatic prostate cancer (mPCA), assessing the definition of the variables of metabolic syndrome, and the potential mechanisms of MetS and mPCA. Methods: Data were obtained from the database of prostate cancer follow-up at the Urology Centre of the First Affiliated Hospital of Xinjiang Medical University (N=1303). After screening by inclusion and exclusion criteria, clinical data of 190 patients diagnosed with mPCA by pathology and imaging from January 2010 to August 2021 were finally included, including 111 cases in the MetS group and 79 cases in the Non-MetS group. Results: The MetS group was higher than the Non-MetS group: T stage, Gleasson score, initial PSA, tumor load, PSA after 7 months of ADT (P<0.05),with a shorter time to progression to CRPC stage(P<0.05)[where the time to progression to CRPC was relatively shorter in the high metabolic score subgroup of the MetS group than in the low subgroup (P<0.05)].Median survival time was significantly shorter in the MetS group than in the Non-MetS group (P<0.05),and there was a correlation with metabolic score, with the higher metabolic score subgroup having a lower survival time than the lower metabolic score subgroup (P<0.05). Conclusion: Those with mPCA combined with MetS had lower PSA remission rates, more aggressive tumors, shorter time to progression to CRPC and shorter median survival times than those with mPCA without MetS.Tumour progression and metabolic score showed a positive correlation, predicting that MetS may promote the progression of mPCA, suggesting that MetS may be a risk factor affecting the prognosis of mPCA.

Expression of the ripAA Gene in the Soilborne Pseudomonas mosselii Can Promote the Control Efficacy against Tobacco Bacterial Wilt.

The environmental bacterium Pseudomonas mosselii produces antagonistic secondary metabolites with inhibitory effects on multiple plant pathogens, including Ralstonia solanacearum, the causal agent of bacterial wilt. In this study, an engineered P. mosselii strain was generated to express R. solanacearum ripAA, which determines the incompatible interactions with tobacco plants. The ripAA gene, together with its native promoter, was integrated into the P. mosselii chromosome. The resulting strain showed no difference in antimicrobial activity against R. solanacearum. Promoter-LacZ fusion and RT-PCR experiments demonstrated that the ripAA gene was transcribed in culture media. Compared with that of the wild type, the engineered strain reduced the disease index by 9.1% for bacterial wilt on tobacco plants. A transcriptome analysis was performed to identify differentially expressed genes in tobacco plants, and the results revealed that ethylene- and jasmonate-dependent defense signaling pathways were induced. These data demonstrates that the engineered P. mosselii expressing ripAA can improve biological control against tobacco bacterial wilt by the activation of host defense responses.

The Clinical and Radiological Outcomes of Subtrochanteric Osteotomy in Crowe Type IV Hip Dysplasia: A Comparison of Three Different Stem Designs.

AIMS: The purpose of this study was to determine whether there are differences in clinical and radiographic outcomes among three different stem designs for subtrochanteric osteotomy in Crowe type IV developmental dysplasia of the hip (DDH). METHODS: A retrospective analysis of prospectively collected data was undertaken from a consecutive series of 37 Crowe type IV DDHs treatment of noncemented total hip arthroplasty with chevron subtrochanteric osteotomy in 30 patients. Patients are divided into three groups, including Ribbed group (using Link Ribbed stem; n = 14), Synergy group (using Synergy stem; n = 9), and Link Classic Uncemented (LCU) group (using LCU stem; n = 14), according to the design of the stem. The clinical and radiographic outcomes were evaluated. RESULTS: All patients were followed for 36 months. The time of bone union of the LCU stem was significantly longer than that of the Synergy stem (P = 0.02) and the Ribbed stem (P > 0.05); the time of bone union of the Ribbed stem was longer than that of the Synergy stem (P > 0.05). The length of stem in the distal femur of the Ribbed stem (P = 0.000) and the Synergy stem (P = 0.001) is significantly longer than that of the LCU stem. There were three hips with malunion, stem loosening, and varus alignment, which were observed in the LCU stem. None of these were observed in Ribbed and Synergy stems. In total hip arthroplasty with a noncemented stem combined with subtrochanteric femoral osteotomy for Crowe IV DDH, 89.2% hips (33/37) can achieve good and excellent clinical outcomes. There were three hips (1 hip in the Ribbed stem and two in the LCU stem) with fair clinical outcomes and one hip (LCU stem) with poor clinical outcomes. CONCLUSIONS: Although Ribbed, Synergy, and LCU stems have similar clinical outcomes, the LCU stem has a tendency to a varus position, longer union time, malunion, and stem loosening, when compared with the Ribbed and Synergy stems. We recommend against adoption of the LCU stem for Crowe IV DDH with subtrochanteric femoral osteotomy. LEVEL OF EVIDENCE: Level III, therapeutic study.

[The Best Time of Minimal Residual Disease Monitoring for Predicting Survival and Prognosis in Children with T-Cell Acute Lymphoblastic Leukemia].

OBJECTIVE: To investigate the optimal time of monitoring minimal residual disease (MRD) for predicting survival and prognosis in children with T-cell acute lymphoblastic leukemia (T-ALL) after treated by CCLG-ALL2008 chemotherapy. METHODS: 96 children with T-ALL receiving CCLG-ALL2008 chemotherapy treated in our hospital from January 2015 to January 2020 were retrospectively summarized. The follow-up time was 9.0-65.0 months, with a median of 43.5 months. Kaplan-Meier survival curve was used to detect the overall event-free survival (EFS) and overall survival (OS) of the patients. The clinical data, MRD levels after 15 d, 33 d and 90 d chemotherapy between EFS group and relapse group, as well as OS group and death group were compared by using univariate analysis. Multivariate Logistic regression analysis was used to screen the main risk factors affecting EFS and OS of the patients. The patients were divided into low, moderate and high-risk according to the MRD level after 15 d, 33 d and 90 d, the differences of EFS and OS between each groups were compared again. RESULTS: By the end of follow-up, 50 patients recurred and other 46 patients non-recurred; 40 patients died and 56 patients survived, the EFS was (49.5±6.3)% and OS was (61.5±5.9)%. Univariate analysis showed that the initial WBC count in EFS group (n=46) was significantly lower than that in relapse group (n=50), and MRD levels after 33 d and 90 d were significantly less also (P<0.05). Prednisone response in OS group (n=56) was better than that in death group (n=40), and central nerve invasion rate was lower, MRD level after 33 and 90 d were lower (P<0.05). Logistic regression analysis showed that MRD level after 90 d was the main risk factor affecting EFS of the patients; prednisone reaction, central nerve invasion and MRD level after 90 d were the main risk factors affecting OS of the patients (P<0.05). There were no differences of EFS or OS between the groups according to the MRD levels after 15 and 33 d (P>0.05), however for 90 d, EFS and OS of the patients in high-risk group were significantly lower than those in medium-risk group, and those in medium-risk group were lower than those in low-risk group (P<0.05). CONCLUSION: The MRD level after 90 days CCLG-ALL2008 chemotherapy may be the best time to predict the survival and prognosis in T-ALL children.

Unsupervised Abstract Reasoning for Raven's Problem Matrices.

Raven's Progressive Matrices (RPM) is highly correlated with human intelligence, and it has been widely used to measure the abstract reasoning ability of humans. In this paper, to study the abstract reasoning capability of deep neural networks, we propose the first unsupervised learning method for solving RPM problems. Since the ground truth labels are not allowed, we design a pseudo target based on the prior constraints of the RPM formulation to approximate the ground-truth label, which effectively converts the unsupervised learning strategy into a supervised one. However, the correct answer is wrongly labelled by the pseudo target, and thus the noisy contrast will lead to inaccurate model training. To alleviate this issue, we propose to improve the model performance with negative answers. Moreover, we develop a decentralization method to adapt the feature representation to different RPM problems. Extensive experiments on three datasets demonstrate that our method even outperforms some of the supervised approaches. Our code is available at https://github.com/visiontao/ncd.

The role of endocrine hormones in the pathogenesis of adolescent idiopathic scoliosis.

Adolescent idiopathic scoliosis (AIS) is a common spinal deformity characterized by changes in the three-dimensional structure of the spine. It usually initiates during puberty, the peak period of human growth when the secretion of numerous hormones is changing, and it is more common in females than in males. Accumulating evidence shows that the abnormal levels of many hormones including estrogen, melatonin, growth hormone, leptin, adiponectin and ghrelin, may be related to the occurrence and development of AIS. The purpose of this review is to provide a summary and critique of the research published on each hormone over the past 20 years, and to highlight areas for future study. It is hoped that the presentation will help provide a better understanding of the role of endocrine hormones in the pathogenesis of AIS.

The LysR-Type Transcriptional Regulator CrgA Negatively Regulates the Flagellar Master Regulator flhDC in Ralstonia solanacearum GMI1000.

The invasion and colonization of host plants by the destructive pathogen Ralstonia solanacearum rely on its cell motility, which is controlled by multiple factors. Here, we report that the LysR-type transcriptional regulator CrgA (RS_RS16695) represses cell motility in R. solanacearum GMI1000. CrgA possesses common features of a LysR-type transcriptional regulator and contains an N-terminal helix-turn-helix motif as well as a C-terminal LysR substrate-binding domain. Deletion of crgA results in an enhanced swim ring and increased transcription of flhDC In addition, the ΔcrgA mutant possesses more polar flagella than wild-type GMI1000 and exhibits higher expression of the flagellin gene fliC Despite these alterations, the ΔcrgA mutant did not have a detectable growth defect in culture. Yeast one-hybrid and electrophoretic mobility shift assays revealed that CrgA interacts directly with the flhDC promoter. Expressing the ß-glucuronidase (GUS) reporter under the control of the crgA promoter showed that crgA transcription is dependent on cell density. Soil-soaking inoculation with the crgA mutant caused wilt symptoms on tomato (Solanum lycopersicum L. cv. Hong yangli) plants earlier than inoculation with the wild-type GMI1000 but resulted in lower disease severity. We conclude that the R. solanacearum regulator CrgA represses flhDC expression and consequently affects the expression of fliC to modulate cell motility, thereby conditioning disease development in host plants.IMPORTANCERalstonia solanacearum is a widely distributed soilborne plant pathogen that causes bacterial wilt disease on diverse plant species. Motility is a critical virulence attribute of R. solanacearum because it allows this pathogen to efficiently invade and colonize host plants. In R. solanacearum, motility-defective strains are markedly affected in pathogenicity, which is coregulated with multiple virulence factors. In this study, we identified a new LysR-type transcriptional regulator (LTTR), CrgA, that negatively regulates motility. The mutation of the corresponding gene leads to the precocious appearance of wilt symptoms on tomato plants when the pathogen is introduced using soil-soaking inoculation. This study indicates that the regulation of R. solanacearum motility is more complex than previously thought and enhances our understanding of flagellum regulation in R. solanacearum.

Functional Characterization of RNA Silencing Suppressor P0 from Pea Mild Chlorosis Virus.

To counteract host antiviral RNA silencing, plant viruses encode numerous viral suppressors of RNA silencing (VSRs). P0 proteins have been identified as VSRs in many poleroviruses. However, their suppressor function has not been fully characterized. Here, we investigated the function of P0 from pea mild chlorosis virus (PMCV) in the suppression of local and systemic RNA silencing via green fluorescent protein (GFP) co-infiltration assays in wild-type and GFP-transgenic Nicotiana benthamiana (line 16c). Amino acid deletion analysis showed that N-terminal residues Asn 2 and Val 3, but not the C-terminus residues from 230-270 aa, were necessary for PMCV P0 (P0PM) VSR activity. P0PM acted as an F-box protein, and triple LPP mutation (62LPxx79P) at the F-box-like motif abolished its VSR activity. In addition, P0PM failed to interact with S-phase kinase-associated protein 1 (SKP1), which was consistent with previous findings of P0 from potato leafroll virus. These data further support the notion that VSR activity of P0 is independent of P0-SKP1 interaction. Furthermore, we examined the effect of P0PM on ARGONAUTE1 (AGO1) protein stability, and co-expression analysis showed that P0PM triggered AGO1 degradation. Taken together, our findings suggest that P0PM promotes degradation of AGO1 to suppress RNA silencing independent of SKP1 interaction.

The Ralstonia solanacearum effector RipI induces a defence reaction by interacting with the bHLH93 transcription factor in Nicotiana benthamiana.

Ralstonia solanacearum releases a set of effectors into plant cells that modify the host defence reaction. The role of the effector protein RipI during infection has not been elucidated. In this study, we demonstrated that transient overexpression of RipI induces the hypersensitive response (HR), up-regulating the HR marker gene hin1, in Nicotiana benthamiana. Deletion of R. solanacearum ripI led to increased virulence in tomato (Solanum lycopersicum) plants. Through yeast two-hybrid and pull-down assays, we identified an interaction between the N. benthamiana transcription factor bHLH93 and RipI, both of which could be localized in the nucleus of Arabidopsis protoplasts. Silencing of bHLH93 markedly attenuated the RipI-induced HR and induced expression of the PDF1.2 defence gene. These data demonstrate that the R. solanacearum effector RipI induces a host defence reaction by interacting with the bHLH93 transcription factor.

Ralstonia solanacearum elicitor RipX Induces Defense Reaction by Suppressing the Mitochondrial atpA Gene in Host Plant.

RipX of Ralstonia solanacearum is translocated into host cells by a type III secretion system and acts as a harpin-like protein to induce a hypersensitive response in tobacco plants. The molecular events in association with RipX-induced signaling transduction have not been fully elucidated. This work reports that transient expression of RipX induced a yellowing phenotype in Nicotiana benthamiana, coupled with activation of the defense reaction. Using yeast two-hybrid and split-luciferase complementation assays, mitochondrial ATP synthase F1 subunit α (ATPA) was identified as an interaction partner of RipX from N. benthamiana. Although a certain proportion was found in mitochondria, the YFP-ATPA fusion was able to localize to the cell membrane, cytoplasm, and nucleus. RFP-RipX fusion was found from the cell membrane and cytoplasm. Moreover, ATPA interacted with RipX at both the cell membrane and cytoplasm in vivo. Silencing of the atpA gene had no effect on the appearance of yellowing phenotype induced by RipX. However, the silenced plants improved the resistance to R. solanacearum. Moreover, qRT-PCR and promoter GUS fusion experiments revealed that the transcript levels of atpA were evidently reduced in response to expression of RipX. These data demonstrated that RipX exerts a suppressive effect on the transcription of atpA gene, to induce defense reaction in N. benthamiana.

Unsupervised Online Video Object Segmentation With Motion Property Understanding.

Unsupervised video object segmentation aims to automatically segment moving objects over an unconstrained video without any user annotation. So far, only few unsupervised online methods have been reported in the literature, and their performance is still far from satisfactory because the complementary information from future frames cannot be processed under online setting. To solve this challenging problem, in this paper, we propose a novel unsupervised online video object segmentation (UOVOS) framework by construing the motion property to mean moving in concurrence with a generic object for segmented regions. By incorporating the salient motion detection and the object proposal, a pixel-wise fusion strategy is developed to effectively remove detection noises, such as dynamic background and stationary objects. Furthermore, by leveraging the obtained segmentation from immediately preceding frames, a forward propagation algorithm is employed to deal with unreliable motion detection and object proposals. Experimental results on several benchmark datasets demonstrate the efficacy of the proposed method. Compared to state-of-the-art unsupervised online segmentation algorithms, the proposed method achieves an absolute gain of 6.2%. Moreover, our method achieves better performance than the best unsupervised offline algorithm on the DAVIS-2016 benchmark dataset. Our code is available on the project website: https://www.github.com/visiontao/uovos.

Ghrelin up-regulates cartilage-specific genes via the ERK/STAT3 pathway in chondrocytes of patients with adolescent idiopathic scoliosis.

Adolescent idiopathic scoliosis (AIS) is a severe spinal deformity that often occurs during puberty. The occurrence of AIS is suggested to be related to abnormal development of cartilage. Our previous study found increased serum ghrelin levels in AIS patients that may linked to the development of AIS. However, whether ghrelin affects cartilage in AIS patients is unclear. We used quantitative real-time PCR (qRT-PCR) and immunohistochemistry to detect the expression of cartilage-specific genes and the ghrelin receptor, growth hormone secretagogue receptor (GHSR). The mRNA and protein levels of collagen II (COLII), SOX9, AGGRECAN (ACAN) and GHSR were higher in AIS patients than in controls. In addition, the protein levels of GHSR downstream signaling pathway members p-STAT3 (Ser727), and p-ERK1/2 were increased. Furthermore, we treated chondrocytes from AIS patients with 100 nM ghrelin, the cell proliferation assay and Western blotting showed that ghrelin promotes chondrocyte proliferation and enhances COLII, SOX9, ACAN, p-ERK1/2 and p-STAT3 expression, respectively. Interestingly, all these observed alterations were abolished by ghrelin + [D-Lys3]-GHRP-6 (a ghrelin receptor inhibitor) treatment. And after U0126 (an inhibitor of ERK1/2 phosphorylation) treatment, ERK1/2 and STAT3 (Ser727) phosphorylation was simultaneously suppressed indicating that ERK1/2 is an upstream pathway protein of STAT3 (Ser727). In conclusion, ghrelin plays an important role in upregulating cartilage-specific genes on AIS primary chondrocytes by activating ERK/STAT3 signaling pathway.

The RSc0454-Encoded FAD-Linked Oxidase Is Indispensable for Pathogenicity in Ralstonia solanacearum GMI1000.

Ralstonia solanacearum is the causal agent of bacterial wilt disease. Here, we report that a large FAD-linked oxidase encoded by RSc0454 in GMI1000 is required for pathogenicity. The FAD-linked oxidase encoded by RSc0454 is composed of 1,345 amino acids, including DUF3683, lactate dehydrogenase (LDH), and succinate dehydrogenase (SDH) domains. The RSc0454 protein showed both LDH and SDH activities. To investigate its role in pathogenicity, a deletion mutant of the RSc0454 gene was constructed in GMI1000, which was impaired in its ability to cause bacterial wilt disease in tomato. A single DUF3683, LDH, or SDH domain was insufficient to restore bacterial pathogenicity. Mutagenesis of the RSc0454 gene did not affect growth rate but caused cell aggregation at the bottom of the liquid nutrient medium, which was reversed by exogenous applications of lactate, fumarate, pyruvate, and succinate. qRT-PCR and promoter LacZ fusion experiments demonstrated that RSc0454 gene transcription was induced by lactate and fumarate (both substrates of LDH). Compared with the downregulation of the succinate dehydrogenase gene sdhBADC and the lactate dehydrogenase gene ldh, RSc0454 gene transcription was enhanced in planta. This suggests that the oxidase encoded by RSc0454 was involved in a redox balance, which is in line with the different living conditions of R. solanacearum.

Response regulator VemR regulates the transcription of flagellar rod gene flgG by interacting with σ54 factor RpoN2 in Xanthomonas citri ssp. citri.

Xanthomonas citri ssp. citri, a polar flagellated bacterium, causes citrus canker disease worldwide. In this study, we found that the X. citri ssp. citri response regulator VemR plays a regulatory role in flagellum-derived cell motility. Deletion of the vemR gene resulted in a reduction in cell motility, as well as reductions in virulence and exopolysaccharide production. Reverse transcription-polymerase chain reaction (RT-PCR) demonstrated that vemR is transcribed in an operon together with rpoN2 and fleQ. In the vemR mutant, the flagellar distal rod gene flgG was significantly down-regulated. Because flgG is also rpoN2 dependent, we speculated that VemR and RpoN2 physically interact, which was confirmed by yeast two-hybrid and maltose-binding protein (MBP) pull-down assays. This suggested that the transcription of flgG is synergistically controlled by VemR and RpoN2. To confirm this, we constructed a vemR and rpoN2 double mutant. In this mutant, the reductions in cell motility and flgG transcription were unable to be restored by the expression of either vemR or rpoN2 alone. In contrast, the expression of both vemR and rpoN2 together in the double mutant restored the wild-type phenotype. Together, our data demonstrate that the response regulator VemR functions as an RpoN2 cognate activator to positively regulate the transcription of the rod gene flgG in X. citri ssp. citri.