Publications by authors named "Jie Zhai"

29 Publications

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Fatal outcome of malignant phyllodes tumor of the breast in pregnancy: a case and literature review.

Gland Surg 2021 Jan;10(1):371-377

Department of Breast Surgical Oncology, National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China.

Phyllodes tumor of the breast (PTB) is a rare fibroepithelial breast neoplasm that accounts for less than 1% of breast tumors. Only a few cases related to pregnancy have been reported. It is not known how pregnancy affects the diagnosis, treatment and prognosis of breast tumors. Here we report a case of a 38-year-old female patient with a small, mobile palpable lump in the left breast for about 15 years. it was considered a benign lesion and no surgical treatment was performed at the beginning. The left breast mass became larger suddenly during pregnancy, However, she did not see the doctor and receive any treatment in time. The lump was resected one year after labor and confirmed to be malignant phyllodes of the breast by histopathology and immunohistochemistry. Unfortunately, local recurrence occurred within six months after the first operation, and lung metastasis occurred eight months later. And this patient finally died 13 months after the operation due to tumor progression. This is the first report of pregnancy-related malignant PTB, with local recurrence and distant metastasis in a short period. This case report highlights a situation: the patient should be diagnosed early and treated in time when she has a previous breast fibroadenoma, but suddenly increased during pregnancy.
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http://dx.doi.org/10.21037/gs-20-538DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7882345PMC
January 2021

Identification of distinct immune landscapes using an automated nine-color multiplex immunofluorescence staining panel and image analysis in paraffin tumor tissues.

Sci Rep 2021 Feb 25;11(1):4530. Epub 2021 Feb 25.

Department of Translational Molecular Pathology, The University of Texas MD Anderson Cancer Center, Unit 9512130 Holcombe Blvd, Houston, TX, 77030, USA.

Immune profiling is becoming a vital tool for identifying predictive and prognostic markers for translational studies. The study of the tumor microenvironment (TME) in paraffin tumor tissues such as malignant pleural mesothelioma (MPM) could yield insights to actionable targets to improve patient outcome. Here, we optimized and tested a new immune-profiling method to characterize immune cell phenotypes in paraffin tissues and explore the co-localization and spatial distribution between the immune cells within the TME and the stromal or tumor compartments. Tonsil tissues and tissue microarray (TMA) were used to optimize an automated nine-color multiplex immunofluorescence (mIF) panel to study the TME using eight antibodies: PD-L1, PD-1, CD3, CD8, Foxp3, CD68, KI67, and pancytokeratin. To explore the potential role of the cells into the TME with this mIF panel we applied this panel in twelve MPM cases to assess the multiple cell phenotypes obtained from the image analysis and well as their spatial distribution in this cohort. We successful optimized and applied an automated nine-color mIF panel to explore a small set of MPM cases. Image analysis showed a high degree of cell phenotype diversity with immunosuppression patterns in the TME of the MPM cases. Mapping the geographic cell phenotype distribution in the TME, we were able to identify two distinct, complex immune landscapes characterized by specific patterns of cellular distribution as well as cell phenotype interactions with malignant cells. Successful we showed the optimization and reproducibility of our mIF panel and their incorporation for comprehensive TME immune profiling into translational studies that could refine our ability to correlate immunologic phenotypes with specific patterns of cells distribution and distance analysis. Overall, this will improve our ability to understand the behavior of cells within the TME and predict new treatment strategies to improve patient outcome.
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http://dx.doi.org/10.1038/s41598-021-83858-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7907283PMC
February 2021

Effect of n-3 polyunsaturated fatty acid supplementation on muscle mass and function with aging: A meta-analysis of randomized controlled trials.

Prostaglandins Leukot Essent Fatty Acids 2021 Feb 12;165:102249. Epub 2021 Jan 12.

Institute of Nutrition & Health, Qingdao University, Qingdao, China; School of Public Health, Qingdao University, Qingdao, China. Electronic address:

The results of randomized controlled trials (RCTs) investigating supplemental n-3 polyunsaturated fatty acids (PUFA) on muscle mass and function have been inconsistent. The present study aimed to quantitatively evaluate the effect of n-3 PUFA supplementation on indicators of muscle mass and function in healthy subjects. A systematic literature search was conducted up to July 2020 with databases of PubMed and Web of science. The random-effects model was implemented to calculate the weighted mean difference of net change of indicators regarding muscle mass and function. A total of nine studies (thirteen treatment groups) with 2067 participants were included for data analysis. The summary estimate showed that n-3 PUFA supplementation significantly increased the grip strength (1.17 kg; 95% CI: 0.27, 2.08 kg). Non-significant effect was observed with respect to muscle mass parameters, including fat mass (-0.67 kg; 95% CI: -2.20, 0.87 kg) and lean mass (0.33 kg; 95% CI: -0.35, 1.00 kg). Regarding muscle function indicators, there were non-significant effects on walking speed (-0.01 m•s; 95% CI: -0.03, 0.01 m•s), time up and go test (-0.25 s; 95% CI: -0.55, 0.04 s), respectively. The findings of this study indicated that supplementation with n-3 PUFA might have beneficial effects to improve muscle mass and function in healthy participants. However, there was no significant improvement in the subjects' muscle mass. Whether n-3 PUFA supplementation has favorable effects in participants with sarcopenia are warranted to be further investigated.
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http://dx.doi.org/10.1016/j.plefa.2021.102249DOI Listing
February 2021

Changes in oxidation-antioxidation function on the thymus of chickens infected with reticuloendotheliosis virus.

BMC Vet Res 2020 Dec 11;16(1):483. Epub 2020 Dec 11.

College of Veterinary Medicine, Northeast Agricultural University, 150030, Harbin, People's Republic of China.

Background: Reticuloendotheliosis virus (REV) is a retrovirus that causes severe immunosuppression in poultry. Animals grow slowly under conditions of oxidative stress. In addition, long-term oxidative stress can impair immune function, as well as accelerate aging and death. This study aimed to elucidate the pathogenesis of REV from the perspective of changes in oxidative-antioxidative function following REV infection.

Methods: A total of 80 one-day-old specific pathogen free (SPF) chickens were randomly divided into a control group (Group C) and an REV-infected group (Group I). The chickens in Group I received intraperitoneal injections of REV with 10/0.1 mL TCID. Thymus was collected on day 1, 3, 7, 14, 21, 28, 35, and 49 for histopathology and assessed the status of oxidative stress.

Results: In chickens infected with REV, the levels of HO and MDA in the thymus increased, the levels of TAC, SOD, CAT, and GPx1 decreased, and there was a reduction in CAT and Gpx1 mRNA expression compared with the control group. The thymus index was also significantly reduced. Morphological analysis showed that REV infection caused an increase in the thymic reticular endothelial cells, inflammatory cell infiltration, mitochondrial swelling, and nuclear damage.

Conclusions: These results indicate that an increase in oxidative stress enhanced lipid peroxidation, markedly decreased antioxidant function, caused thymus atrophy, and immunosuppression in REV-infected chickens.
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http://dx.doi.org/10.1186/s12917-020-02708-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7731740PMC
December 2020

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Mol Ther Oncolytics 2020 Jun 30;17:153-168. Epub 2020 Mar 30.

Department of Breast Surgical Oncology, National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100021, People's Republic of China.

A plethora of previous studies have been focused on the role of indoleamine 2,3-dioxygenase 1 (IDO1) in cancer immunity; however, the alternative way of targeting tryptophan 2,3-dioxygenase (TDO2) in cancer immunotherapy has been largely ignored. In particular, the specific role of TDO2 in breast cancer remains unclear. In the present study, we systematically explored and validated the expression and prognostic value of TDO2 in breast cancer using large-scale transcriptome data. We observed overexpression of TDO2 in many types of cancer tissues compared with adjacent normal tissues. TDO2 overexpression was revealed to be positively correlated with malignancy and tumor grade in breast cancer. TDO2 expression was higher in estrogen-negative breast cancer and triple-negative breast cancer, and it was correlated with worse outcome in breast cancer patients. TDO2 expression was correlated with immune infiltrates and tryptophan metabolism-related genes (IDO1 and kynureninase [KYNU]). Therefore, our results indicated that TDO2 plays a pivotal role in regulating the immune microenvironment and tryptophan metabolism in breast cancer, and it predicts poor prognosis in breast cancer, which suggests that TDO2 might be a promising novel immunotherapy target for breast cancer. Additionally, we established the concept that tryptophan-catabolizing enzymes (IDO1, IDO2, TDO2, and KYNU) may function through co-regulating the immunological microenvironment, and thus immunotherapy targeting IDO1 alone might be insufficient.
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http://dx.doi.org/10.1016/j.omto.2020.03.013DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7178007PMC
June 2020

Regulatory mechanism of microRNA-155 in chicken embryo fibroblasts in response to reticuloendotheliosis virus infection.

Vet Microbiol 2020 Mar 10;242:108610. Epub 2020 Feb 10.

Heilongjiang Key Laboratory for Laboratory Animals and Comparative Medicine, College of Veterinary Medicine, Northeast Agricultural University, NO. 59 Mucai Street, Harbin 150030, People's Republic of China. Electronic address:

Reticuloendotheliosis virus (REV) infection of multiple avian species can lead to a number of diseases such as runting syndrome, immunosuppression and oncogenesis, causing major economic losses. MicroRNAs play important roles in post-transcriptional regulation, effectively inhibiting protein synthesis, and participating in many biological processes in cells, including proliferation, differentiation, apoptosis, lipometabolism, virus infection and replication, and tumorigenesis. Based on our previous high-throughput sequencing results, we explore the regulatory mechanisms of microRNA-155(miR-155) in chicken embryo fibroblasts (CEFs) in response to REV infection. Our results revealed expression of miR-155 in CEFs after REV infection upregulated in a time- and dose-dependent manner, indicating miR-155 plays a role in REV infection in CEFs indeed. After transfected with miR-155-mimic and miR-155-inhibitor, we found overexpression of miR-155 targeted caspase-6 and FOXO3a to inhibit apoptosis and accelerate cell cycle, thus improving viability of REV-infected CEFs. This result also verified the protective role of miR-155 in the viability of CEFs in the presence of REV. Knockdown of miR-155 also supported these above conclusions. Our findings uncover a new mechanism of REV pathogenesis in CEFs, and also provide a theoretical basis for uncovering new effective treatment and prevention methods for RE based on miR-155.
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http://dx.doi.org/10.1016/j.vetmic.2020.108610DOI Listing
March 2020

Representation learning for clinical time series prediction tasks in electronic health records.

BMC Med Inform Decis Mak 2019 12 17;19(Suppl 8):259. Epub 2019 Dec 17.

Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, 528 Zhangheng Road, Shanghai, 201203, China.

Background: Electronic health records (EHRs) provide possibilities to improve patient care and facilitate clinical research. However, there are many challenges faced by the applications of EHRs, such as temporality, high dimensionality, sparseness, noise, random error and systematic bias. In particular, temporal information is difficult to effectively use by traditional machine learning methods while the sequential information of EHRs is very useful.

Method: In this paper, we propose a general-purpose patient representation learning approach to summarize sequential EHRs. Specifically, a recurrent neural network based denoising autoencoder (RNN-DAE) is employed to encode inhospital records of each patient into a low dimensional dense vector.

Results: Based on EHR data collected from Shuguang Hospital affiliated to Shanghai University of Traditional Chinese Medicine, we experimentally evaluate our proposed RNN-DAE method on both mortality prediction task and comorbidity prediction task. Extensive experimental results show that our proposed RNN-DAE method outperforms existing methods. In addition, we apply the "Deep Feature" represented by our proposed RNN-DAE method to track similar patients with t-SNE, which also achieves some interesting observations.

Conclusion: We propose an effective unsupervised RNN-DAE method to summarize patient sequential information in EHR data. Our proposed RNN-DAE method is useful on both mortality prediction task and comorbidity prediction task.
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http://dx.doi.org/10.1186/s12911-019-0985-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6916209PMC
December 2019

Mechanism of trastuzumab resistance caused by HER-2 mutation in breast carcinomas.

Cancer Manag Res 2019 2;11:5971-5982. Epub 2019 Jul 2.

Department of Breast Surgical Oncology, National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100021, People's Republic of China.

Trastuzumab is an effective drug for the treatment of Her2-positive breast cancer. But, primary or secondary resistances to trastuzumab have become an important factor influencing the curative effect. The mechanisms of trastuzumab resistance are somewhat complex. The present work aims to explore the mechanism of trastuzumab resistance caused by HER-2 mutation in breast carcinomas. Firstly, the HER2 wild type (WT) and HER2 mutant (HER2 Q429R, HER2 Q429H and HER2 T798M are the commonest 3 types of mutations) MCF7 cell lines were established. Cell proliferation inhibition was then assessed by the Cell Counting Kit-8 assay and BrdU assay. Transwell invasion assays were also conducted to investigate the metastatic potential influenced by the HER2 mutation. Furthermore, Western blotting and co-immunoprecipitation were conducted to detect protein levels and the physical interaction of HER2 and trastuzumab. The results showed that the mutant MCF7 cells were less sensitive to trastuzumab than the WTMCF7 cells. The mutation of HER2 almost had no influence on the expression of HER2 and the interaction of HER2 and trastuzumab. Finally, the mutation of HER2 weakened the inhibition of trastuzumab in the PI3K/AKT pathways. In addition, the inhibition of PI3K/AKT signaling-pathway increased the trastuzumab-sensitivity of HER2-mutant MCF7 cells. Dysregulation of the PI3K-AKT signaling-pathway was a key mechanism inducing the trastuzumab-resistance to HER2 mutant breast cancer cells.
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http://dx.doi.org/10.2147/CMAR.S194137DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6618040PMC
July 2019

LINC01355 suppresses breast cancer growth through FOXO3-mediated transcriptional repression of CCND1.

Cell Death Dis 2019 06 26;10(7):502. Epub 2019 Jun 26.

Department of Breast Surgical Oncology, National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China.

Previously, several protein-coding tumor suppressors localized at 1p36 have been reported. In the present work, we focus on functional long non-coding RNAs (lncRNAs) embedded in this locus. Small interfering RNA was used to identify lncRNA candidates with growth-suppressive activities in breast cancer. The mechanism involved was also explored. LINC01355 were downregulated in breast cancer cells relative to non-malignant breast epithelial cells. Overexpression of LINC01355 significantly inhibited proliferation, colony formation, and tumorigenesis of breast cancer cells. LINC01355 arrested breast cancer cells at the G0/G1 phase by repressing CCND1. Moreover, LINC01355 interacted with and stabilized FOXO3 protein, leading to transcriptional repression of CCND1. Importantly, LINC01355-mediated suppression of breast cancer growth was reversed by knockdown of FOXO3 or overexpression of CCND1. Clinically, LINC01355 was downregulated in breast cancer specimens and correlated with more aggressive features. There was a negative correlation between LINC01355 and CCND1 expression in breast cancer samples. LINC01355 acts as a tumor suppressor in breast cancer, which is ascribed to enhancement of FOXO3-mediated transcriptional repression of CCND1. Re-expression of LINC01355 may provide a potential therapeutic strategy to block breast cancer growth and progression.
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http://dx.doi.org/10.1038/s41419-019-1741-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6594972PMC
June 2019

Recent Advances in Understanding FOXN3 in Breast Cancer, and Other Malignancies.

Front Oncol 2019 31;9:234. Epub 2019 May 31.

Department of Breast Surgical Oncology, National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China.

FOXN3 (forkhead box N3; CHES1: check point suppressor 1) belongs to the forkhead box (FOX) protein family. FOXN3 displays transcriptional inhibitory activity, and is involved in cell cycle regulation and tumorigenesis. FOXN3 is a tumor suppresser and alterations in FOXN3 are found in of a variety of cancers including melanoma, osteosarcoma, and hepatocellular carcinoma. While the roles of FOXN3 role in some cancers have been explored, its role in breast cancer remains unclear. Here we describe current state of knowledge of FOXN3 functions, and focus on its roles (known and potential) in breast cancer.
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http://dx.doi.org/10.3389/fonc.2019.00234DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6555274PMC
May 2019

Changes in apoptosis, proliferation and T lymphocyte subtype on thymic cells of SPF chickens infected with reticuloendotheliosis virus.

Mol Immunol 2019 07 29;111:87-94. Epub 2019 Apr 29.

College of Veterinary Medicine, Northeast Agricultural University, Harbin 150030, PR China; Heilongjiang Key Laboratory of Laboratory Animals and Comparative Medicine, Harbin 150030, PR China. Electronic address:

Reticuloendotheliosis virus (REV), an avian retrovirus is able to infect a variety of birds and can cause immunosuppression. The aim of this study was to investigate the relationship of thymic lymphocytes apoptosis, proliferation and T cell subtype with immunosuppression. In this study, a hundred and twenty one-day old SPF chickens were randomly divided into control groups (group C) and a REV infection groups (group I). The chickens of group I received intraperitoneal injections of REV with 10/0.1 ml TCID. On day 14, 21, 28 and 35 post-inoculation, the chickens of C group and I group were sacrificed by cardiac puncture blood collection, and the thymic lymphocytes was sterile collected. The proliferation ability of lymphocytes was tested by Cell Counting Kit-8. Flow cytometry was performed to detect apoptosis, cell cycle stage and the change in T cell subtype. The RNA genome copy numbers of REV virus were detected using real-time PCR. Real-time PCR and western blotting were performed to analyze the expression of CyclinD1 and Bcl-2. Our results showed that REV genome copy number steadily declined, the proliferation potential of thymic lymphocytes was inhibited, lymphocytes apoptosed, the ratio of CD4+/CD8+ decreased and the expression of CyclinD1 and Bcl-2 were firstly inhibited, then rapidly recovered. Thus, immunosuppression lead by REV is closely related to the change of T cell subtype, apoptosis, and proliferation of thymic lymphocytes.
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http://dx.doi.org/10.1016/j.molimm.2019.04.003DOI Listing
July 2019

Expression signatures and roles of microRNAs in inflammatory breast cancer.

Cancer Cell Int 2019 31;19:23. Epub 2019 Jan 31.

1Department of Breast Surgical Oncology, National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100021 China.

Inflammatory breast cancer (IBC) is an infrequent but aggressive manifestation of breast cancer, which accounts for 2-4% of all breast cancer cases but responsible for 7-10% of breast cancer-related deaths, and with a 20-30% 10-year overall survival compared with 80% for patients with non-IBC with an unordinary phenotype, whose molecular mechanisms are still largely unknown to date. Discovering and identifying novel bio-markers responsible for diagnosis and therapeutic targets is a pressing need. MicroRNAs are a class of small non-coding RNAs that are capable to post-transcriptionally regulate gene expression of genes by targeting mRNAs, exerting vital and tremendous affects in numerous malignancy-related biological processes, including cell apoptosis, metabolism, proliferation and differentiation. In this study, we review present and high-quality evidences regarding the potential applications of inflammatory breast cancer associated microRNAs for diagnosis and prognosis of this lethal disease.
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http://dx.doi.org/10.1186/s12935-018-0709-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6357482PMC
January 2019

Immunological therapy: A novel thriving area for triple-negative breast cancer treatment.

Cancer Lett 2019 02 9;442:409-428. Epub 2018 Nov 9.

Department of Breast Surgical Oncology, National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100021, China. Electronic address:

Triple-negative breast cancer (TNBC) refers to cancers that are low in expression of the estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2). TNBC tends to behave more aggressively than other types of breast cancer. Unlike other breast cancer subtypes (ie, ER-positive, HER2-positive subtypes), there are no approved targeted treatments available, other than the administration of chemotherapy. Immunotherapy is a new kind of treatment approach for TNBC when compared with the surgical treatment, chemotherapy, endocrine therapy, and molecular targeting therapy. The present article reviews the research progresses of immunotherapy for TNBC in recent years. The full text structure covers molecular classification of TNBC, active immunotherapy of TNBC, passive immunotherapy of TNBC, oncolytic immunotherapy and the prospect of immunotherapy for TNBC.
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http://dx.doi.org/10.1016/j.canlet.2018.10.042DOI Listing
February 2019

Integrative Analyses of Transcriptome Sequencing Identify Functional miRNAs in the Chicken Embryo Fibroblasts Cells Infected With Reticuloendotheliosis Virus.

Front Genet 2018 29;9:340. Epub 2018 Aug 29.

Department of Pathophysiology, College of Veterinary Medicine, Northeast Agricultural University, Harbin, China.

In this study, we found a much higher proportion of reticuloendotheliosis virus (REV) infected chicken embryo fibroblasts (CEF) were in active cell division phase than that of control cells which indicated that REV can affect the fate of CEF. So, we performed high-throughput sequencing and transcriptomic analysis to identify functional miRNAs, in order to figure out the possible mechanism in the interaction of REV with CEF. In total, 50 differentially expressed miRNAs (DEmiRNAs) were identified. Then target genes of DEmiRNAs were predicted and identified by transcriptome profile results. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment were conducted to analyze the identified target genes of miRNAs which showed that metabolism, cell cycle, and apoptosis were the most related pathways involved in infection of REV. We analyzed the genes related to cell cycle which indicated that CyclinD1-CDK6 complex played an important role in regulating the transition of the cell cycle from G1 phase to S phase during REV infection. Fluorescence microscope identification showed that REV inhibited the apoptosis of CEF which was in accordance with transcriptome results. A novel miRNA, named novel-72 was found, KEGG analysis was conducted to predict the biological function of its target genes which showed that those target genes were significantly enriched in mTOR signaling pathway and functioned to promote cell cycle and cell growth during the REV infection. In conclusion, REV could induce the up-regulation of cell metabolism, cell cycle and mTOR signaling pathway while inhibit apoptosis of the cell.
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http://dx.doi.org/10.3389/fgene.2018.00340DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6128223PMC
August 2018

c-Jun and Camk2a contribute to the drug resistance of induction docetaxel/cisplatin/5-fluorouracil in hypopharyngeal carcinoma.

Int J Clin Exp Pathol 2018 1;11(9):4605-4613. Epub 2018 Sep 1.

Department of Otorhinolaryngology Head and Neck Surgery, Beijing Tongren Hospital, Capital Medical University Beijing, China.

Hypopharyngeal carcinoma (HPC) is a subtype of head and neck squamous cell carcinoma, and prognosis has improved significantly over the past three decades. Induction docetaxel/cisplatin/5 fluorouracil (TPF) chemotherapy is regarded as the standard of treatment for locoregionally advanced HPC. However, patients who do not respond to cisplatin suffer, rather than benefit, from chemotherapy treatment. The goal of this study was to identify molecules involved in TPF resistance and to clarify their molecular mechanisms. Using the FaDu cell line as the cell model, the TPF IC50 was identified, and c-Jun, IL6, Camk2a, c-fos knockdown using siRNAs resulted in a significant declined TPF IC50. Retrospective analysis of the expression status of c-Jun, IL6, Camk2a, and c-fos by immunohistochemistry staining in sectioned HPC tissues from TPF-sensitive and TPF-insensitive patients shows that Camk2a and c-Jun were associated with the clinical pathogenesic features in HPC. The experiments also indicate that both Camk2a and c-Jun were responsive to TPF treatment. This study identified Camk2a and c-Jun as candidate genes that confer induction TPF resistance, which would help in the discovery of potential therapeutic markers and in developing a personalized and precise treatment approach for HPC patients.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6962968PMC
September 2018

Analysis of alternative splicing in chicken embryo fibroblasts in response to reticuloendotheliosis virus infection.

Avian Pathol 2018 Dec 20;47(6):585-594. Epub 2018 Sep 20.

a Laboratory Pathological Physiology, College of Veterinary Medicine , Northeast Agricultural University , Harbin , People's Republic of China.

Alternative splicing (AS) plays a significant role in regulation of genomic expression at the transcriptional level and is involved in many important biological functions of cells, thus a gene can be spliced into distinct transcript variants then translated to many different kinds of protein. Reticuloendotheliosis virus (REV) is a kind of retrovirus that can infect multiple avian species, leading to runting syndrome, immunosuppression and oncogenesis. In this present study, we analyzed AS in REV-infected chicken embryo fibroblasts (CEFs) which were inoculated with the second generation of REV (group VB) and compared with normal CEFs (group C) by high-throughput RNA sequencing technology. A total of 6,939 genes which were alternatively spliced were detected, among them, skipped exon (SE) was the most common pattern. Moreover, 5,607 AS genes were detected as differentially expressed; compared with group C, group VB has 2,825 genes upregulated significantly and 2,782 genes downregulated significantly. These 5,607 differentially expressed AS genes are involved in many important biological processes. Many of them are involved in apoptosis and tumourigenesis. We also proved, by agarose gel electrophoresis, that AS events predicted by our study are authentic and AS is closely related with apoptosis and tumourigenesis in REV-infected CEFs. Our study provides the best analysis to date of the potential link between AS and CEFs in response to REV infection. Research highlights Transcriptomics analysis of REV-infected CEFs using high-throughput sequencing. Potential link between alternative splicing and CEFs in response to REV infection. Skipped exon is the most common spliced pattern in REV-infected CEFs. Differentially expressed genes mainly involved in apoptosis and tumourigenesis.
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http://dx.doi.org/10.1080/03079457.2018.1511047DOI Listing
December 2018

An uncommon granulocytic sarcoma of the breast: a case report and literature review.

Onco Targets Ther 2018 25;11:3685-3690. Epub 2018 Jun 25.

Department of Breast Surgical Oncology, National Cancer Center/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, People's Republic of China,

Granulocytic sarcoma (GS) is an uncommon extramedullary manifestation of acute myeloid leukemia. GS is often likely to be clinically misdiagnosed as another type of primary breast cancer due to its rarity. We report an uncommon case of breast GS in a patient and review the relevant literature.
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http://dx.doi.org/10.2147/OTT.S149149DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6026916PMC
June 2018

Competing endogenous RNA network analysis of CD274, IL‑10 and FOXP3 co‑expression in laryngeal squamous cell carcinoma.

Mol Med Rep 2018 Mar 19;17(3):3859-3869. Epub 2017 Dec 19.

Department of Otorhinolaryngology Head and Neck Surgery, Beijing Tongren Hospital, Capital Medical University, Beijing 100730, P.R. China.

Laryngeal squamous cell carcinoma (LSCC) is one of the most common types of head and neck malignant tumor; however, there is a lack of effective molecular targets for therapy. The present study detected the expression of three immunity‑associated molecules [forkhead box (FOX)3, interleukin (IL)‑10 and cluster of differentiation (CD)274] in 133 LSCC samples using immunohistochemistry (IHC); subsequently, the association between their expression and the clinical characteristics of LSCC were analyzed. Spearman's rank correlation method, Kaplan‑Meier and Cox regression model were used to analyze the correlations of the three proteins and their clinical significance. StarBase and miRTarBase databases were used to establish the competitive endogenous (ce)RNA network of the three molecules. IHC demonstrated that the positive expression rates of FOXP3, IL‑10 and CD274 were 68.4, 73.7 and 58.6% in 133 LSCC samples, respectively. In addition, it was identified that the expression of the three proteins was closely correlated with the clinical characteristics of LSCC, including lymph node metastasis and prognosis (P<0.05). There was also a significant association of co‑expression between any two proteins (P<0.001). Furthermore, the expression levels of FOXP3, IL‑10 and CD274 were negatively associated with the survival rate of patients with LSCC (P<0.05). The results of a Cox regression model suggested that the three proteins were prognostic risk factors for LSCC (P<0.05). The ceRNA network revealed that 10 microRNAs (miRs; including miR‑16‑5p and miR‑214‑3p), 123 long non‑coding RNAs (including X‑inactive specific transcript, H19 and metastasis associated lung adenocarcinoma transcript 1) and 408 circular RNAs (including ATP‑binding cassette subfamily C member 1 hsa_circ_001569 and ISY1 splicing factor homolog hsa_circ_001859) may regulate the expression of FOXP3, IL‑10 and CD274. The data generated from the present study may increase the understanding of the immune escape mechanisms of LSCC and may be beneficial for the development of a specific immunotherapy.
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http://dx.doi.org/10.3892/mmr.2017.8307DOI Listing
March 2018

TIPE2 expression is increased in peripheral blood mononuclear cells from patients with rheumatoid arthritis.

Oncotarget 2017 Oct 23;8(50):87472-87479. Epub 2017 Sep 23.

Department of Orthopedic Surgery, Peking Union Medical College Hospital, Peking Union Medical College, Chinese Academy of Medical Science, Beijing, China.

We investigated the changes in mRNA and protein expression of tumor necrosis factor-α-induced protein 8-like 2 (TIPE2) and PEST-containing nuclear protein (PCNP) in peripheral blood lymphocytes from 54 patients with rheumatoid arthritis (RA) and the spleens of model mice with collagen-induced arthritis (CIA) to generate new ideas for clinical diagnosis and treatment. Expression levels of both TIPE2 and PCNP were higher in RA patients and CIA mice than in their respective controls. They were also higher in the 32 patients with active RA than in the 22 with inactive RA ( < 0.001 for both). After comprehensively treating patients with active RA with anti-inflammatory and antirheumatic drugs for 6 months, they were stable, and there was no difference in TIPE2 levels between the treated patients and those with inactive RA ( = 0.85). In addition, TIPE2 mRNA levels in peripheral blood correlated positively with PCNP (R = 0.744, = 0.001). The DAS28 score correlated positively with peripheral blood TIPE2 levels in the RA patients (R = 0.945, = 0.001). These findings suggest TIPE2 expression increases with the severity of RA.
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http://dx.doi.org/10.18632/oncotarget.21267DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5675647PMC
October 2017

Minichromosome maintenance (MCM) protein 4 overexpression is a potential prognostic marker for laryngeal squamous cell carcinoma.

J BUON 2017 Sep-Oct;22(5):1272-1277

Department of Otorhinolaryngology, Head and Neck Surgery, Beijing Tongren Hospital, Capital Medical University, Beijing, 100730, China.

Purpose: The minichromosomal maintenance (MCM) proteins are involved in the initiation and DNA replication. The role of MCM4 remains to be elucidated. The purpose of this study was to investigate the effects of MCM4 in laryngeal squamous cell carcinoma (LSCC) cell growth and apoptosis.

Methods: LSCC cell line UMSCC 5 was used in this study. The small interfering RNA (siRNA) of MCM 4 gene was used to identify the effects of MCM4 on the proliferation and apoptosis using methylimidazole tetrazolium (MTT) assay and flow-cytometry, respectively. Confirmed LSCC and adjacent non-tumor tissues were collected from 34 patients who were willing to participate in the study, from 2010 through 2015, from 163 patients undergoing treatment in the Department of Otorhinolaryngology/Head and Neck Surgery of Beijing Tongren Hospital in Capital Medical University of P.R. China. Immunohistochemical staining of MCM4 expression in the resected tissues was performed to analyze the correlation between its expression and the clinicopathological characteristics.

Results: The results showed that siRNA of MCM4 could significantly inhibit LSCC cell line UMSCC 5 proliferation and induce apoptosis. MCM4 mRNA was higher expressed in carcinoma tissues than in adjacent normal tissues. MCM4 expression was correlated with male gender, smoking history and poor differentiation.

Conclusions: We noticed a significant role for MCM4 overexpression in human LSCC tissues and their corresponding adjacent non-neoplastic tissues and found that siRNA of MCM4 can significantly decrease the proliferation of cancer cells. It is suggested that MCM4 profiling could potentially be used to predict response to treatment and prognosis in LSCC.
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July 2019

SLC7A11, a component of cysteine/glutamate transporter, is a novel biomarker for the diagnosis and prognosis in laryngeal squamous cell carcinoma.

Oncol Rep 2017 Nov 20;38(5):3019-3029. Epub 2017 Sep 20.

Department of Pathology, Beijing Key Laboratory of Head and Neck Molecular Diagnostic Pathology, Beijing Tongren Hospital, Capital Medical University, Beijing 100730, P.R. China.

Solute carrier family 7, membrane 11 (SLC7A11) or (xCT) is a component of the cysteine-glutamate transporter, which plays a critical role in glutathione homeostasis which is important to protect cells from oxidative stress. SLC7A11 is distributed in various tissues and participates in the occurrence of a number of diseases, particularly in the pathogenesis of malignant tumors, but its role in laryngeal cancer development has not yet been clearly defined. The objective of the present study was to investigate the role of SLC7A11 in laryngeal squamous cell carcinoma (LSCC). We conducted immunohistochemistry and RT-PCR to evaluate the protein and mRNA levels of SLC7A11 in LSCC and in control tissues, respectively. The knockdown experiments were conducted with SLC7A11 short hairpin RNA (shRNA) lentivirus, and the protein and mRNA levels of SLC7A11 were assessed by RT-PCR and western blotting. The functional study of SLC7A11 in vitro was conducted by MTT assay, and the effects on the cell cycle were detected using flow cytometry. Immunohistochemical results revealed that the expression levels of SLC7A11, Ki-67 and p53 in LSCC tissues were higher than those in laryngeal dysplasia tissues. The Spearman rank correlation analysis revealed that the expression of SLC7A11 was positively correlated with the expression of p53 and Ki-67. Cox regression analysis and Kaplan-Meier plots confirmed that the expression levels of SLC7A11 were a prognostic factor for overall survival (OS) rates and postoperative recurrence of LSCC. Moreover, the functional study of SLC7A11 in vitro revealed that knockdown of SLC7A11 using shRNA inhibited cell proliferation by inducing cell cycle arrest at the G1 phase. Immunohistochemical and RT-PCR results and knockdown experiments of SLC7A11 revealed that SLC7A11 was involved in the progression of LSCC, and may provide clinical information for the evaluation of OS rates and postoperative recurrence of LSCC. Collectively, these observations suggest that SLC7A11 may be a vital biomarker for the diagnosis and prognosis in human LSCC, and targeting SLC7A11 appears to be a potentially significant method for LSCC treatment.
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http://dx.doi.org/10.3892/or.2017.5976DOI Listing
November 2017

A meta-analysis: Is there any association between MiR-608 rs4919510 polymorphism and breast cancer risks?

PLoS One 2017 22;12(8):e0183012. Epub 2017 Aug 22.

Department of Breast Surgical Oncology, China National Cancer Center/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Chaoyangqu, Panjiayuan, Beijing, P. R. China.

Object: To combine the data from previously conducted studies about the associations between miR-608 rs4919510 polymorphism (C>G) and breast cancer risks.

Methods: According to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines, we conducted a systematic review of the related literatures searched from PubMed, Embase, Cochrane Library, Web of Science, and China National Knowledge Internet (CNKI) (time: ~ December 2016). Using DerSimonian-Laird random-effects models [Pooling Model: Mantel Haenszel (MH)], odd ratios (ORs) with 95% confidence intervals (95% CIs) were estimated in the allele model, homozygote model, heterozygote model, dominant model and recessive model. Heterogeneity was analyzed using Labbr plots and I2 statistic. Publication bias was analyzed using contour-enhanced funnel plots.

Results: We included 5 eligible studies with 7948 patients. The ORs and their 95% CIs in the 5 genetic models mentioned above were 1.009 (95% CI: 0.922, 1.104; p = 0.847), 1.098 (95% CI: 0.954, 1.264; p = 0.194), 1.076 (95% CI: 0.956, 1.211; p = 0.227), 1.043 (95% CI: 0.880, 1.236; p = 0.628), 1.007 (95% CI: 0.906, 1.118; p = 0.899), respectively.

Conclusion: In the present meta-analysis, no relationships between miR-608 rs4919510 polymorphism (C>G) and the risk of breast cancer were found. More studies are warranted to further validate the conclusion.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0183012PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5568721PMC
October 2017

Microarray gene expression analysis of chemosensitivity for docetaxel, cisplatin and 5-fluorouracil (TPF) combined chemotherapeutic regimen in hypopharyngeal squamous cell carcinoma.

Chin J Cancer Res 2017 Jun;29(3):204-212

Department of Otorhinolaryngology Head and Neck Surgery, Beijing Tongren Hospital, Capital Medical University, Beijing 100730, China.

Objective: To screen out a set of candidate genes which could help to determine whether patients with hypopharyngeal squamous cell carcinoma (HSCC) could benefit from docetaxel, cisplatin and 5-fluorouracil (TPF) induction chemotherapy.

Methods: Gene-expression profiles in 12 TPF-sensitive patients were compared to 9 resistant controls by microarray analysis. Subsequently, expression levels of potential biomarkers in chemosensitive cell line FaDu after TPF treatment were observed by quantitative real-time polymerase chain reaction (qRT-PCR).

Results: Through microarray analysis, 1,579 differentially expressed genes were identified, of which 815 were up-regulated in TPF chemotherapy-responsive tissues whereas 764 were down-regulated. Gene ontology (GO) analysis suggested these genes participating in physiological processes including transcription and its regulation, cellular signal transduction and metabolic process. Additionally, Kyoto Encyclopedia of Genes and Genomes (KEGG) database revealed that MAPK and Jat/STAT signaling pathways occupied important roles in TPF chemotherapeutic sensitivity. Moreover, cell culture experiments revealed the expression alternations of , , , and exposed to TPF treatment by qRT-PCR, whilst providing an insight into the mechanism underlying TPF chemotherapeutic response in HSCC.

Conclusions: These results provided a battery of genes related to TPF chemotherapeutic sensitivity and might act as molecular targets in HSCC treatment. Moreover, these candidate biomarkers could contribute to HSCC individualized treatment.
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http://dx.doi.org/10.21147/j.issn.1000-9604.2017.03.06DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5497207PMC
June 2017

Web service reputation evaluation based on QoS measurement.

ScientificWorldJournal 2014 13;2014:373902. Epub 2014 Apr 13.

Department of Computer Science and Engineering, East China University of Science and Technology, Shanghai 200237, China.

In the early service transactions, quality of service (QoS) information was published by service provider which was not always true and credible. For better verification the trust of the QoS information was provided by the Web service. In this paper, the factual QoS running data are collected by our WS-QoS measurement tool; based on these objectivity data, an algorithm compares the difference of the offered and measured quality data of the service and gives the similarity, and then a reputation evaluation method computes the reputation level of the Web service based on the similarity. The initial implementation and experiment with three Web services' example show that this approach is feasible and these values can act as the references for subsequent consumers to select the service.
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http://dx.doi.org/10.1155/2014/373902DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4009137PMC
June 2015

Novel web service selection model based on discrete group search.

ScientificWorldJournal 2014 15;2014:460593. Epub 2014 Apr 15.

Department of Computer Science and Engineering, East China University of Science and Technology, Shanghai 200237, China.

In our earlier work, we present a novel formal method for the semiautomatic verification of specifications and for describing web service composition components by using abstract concepts. After verification, the instantiations of components were selected to satisfy the complex service performance constraints. However, selecting an optimal instantiation, which comprises different candidate services for each generic service, from a large number of instantiations is difficult. Therefore, we present a new evolutionary approach on the basis of the discrete group search service (D-GSS) model. With regard to obtaining the optimal multiconstraint instantiation of the complex component, the D-GSS model has competitive performance compared with other service selection models in terms of accuracy, efficiency, and ability to solve high-dimensional service composition component problems. We propose the cost function and the discrete group search optimizer (D-GSO) algorithm and study the convergence of the D-GSS model through verification and test cases.
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http://dx.doi.org/10.1155/2014/460593DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4032750PMC
January 2015

Dynamic trans-acting factor colocalization in human cells.

Cell 2013 Oct 24;155(3):713-24. Epub 2013 Oct 24.

Department of Genetics, Stanford University School of Medicine, Stanford, CA 94305, USA.

Different trans-acting factors (TFs) collaborate and act in concert at distinct loci to perform accurate regulation of their target genes. To date, the cobinding of TF pairs has been investigated in a limited context both in terms of the number of factors within a cell type and across cell types and the extent of combinatorial colocalizations. Here, we use an approach to analyze TF colocalization within a cell type and across multiple cell lines at an unprecedented level. We extend this approach with large-scale mass spectrometry analysis of immunoprecipitations of 50 TFs. Our combined approach reveals large numbers of interesting TF-TF associations. We observe extensive change in TF colocalizations both within a cell type exposed to different conditions and across multiple cell types. We show distinct functional annotations and properties of different TF cobinding patterns and provide insights into the complex regulatory landscape of the cell.
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http://dx.doi.org/10.1016/j.cell.2013.09.043DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4079469PMC
October 2013

Application of stopped-flow kinetics methods to investigate the mechanism of action of a DNA repair protein.

J Vis Exp 2010 Mar 31(37). Epub 2010 Mar 31.

Molecular Biology and Biochemistry Department, Wesleyan University.

Transient kinetic analysis is indispensable for understanding the workings of biological macromolecules, since this approach yields mechanistic information including active site concentrations and intrinsic rate constants that govern macromolecular function. In case of enzymes, for example, transient or pre-steady state measurements identify and characterize individual events in the reaction pathway, whereas steady state measurements only yield overall catalytic efficiency and specificity. Individual events such as protein-protein or protein-ligand interactions and rate-limiting conformational changes often occur in the millisecond timescale, and can be measured directly by stopped-flow and chemical-quench flow methods. Given an optical signal such as fluorescence, stopped-flow serves as a powerful and accessible tool for monitoring reaction progress from substrate binding to product release and catalytic turnover(1,2). Here, we report application of stopped-flow kinetics to probe the mechanism of action of Msh2-Msh6, a eukaryotic DNA repair protein that recognizes base-pair mismatches and insertion/deletion loops in DNA and signals mismatch repair (MMR)(3-5). In doing so, Msh2-Msh6 increases the accuracy of DNA replication by three orders of magnitude (error frequency decreases from approximately 10(-6) to 10(-9) bases), and thus helps preserve genomic integrity. Not surprisingly, defective human Msh2-Msh6 function is associated with hereditary non-polyposis colon cancer and other sporadic cancers(6-8). In order to understand the mechanism of action of this critical DNA metabolic protein, we are probing the dynamics of Msh2-Msh6 interaction with mismatched DNA as well as the ATPase activity that fuels its actions in MMR. DNA binding is measured by rapidly mixing Msh2-Msh6 with DNA containing a 2-aminopurine (2-Ap) fluorophore adjacent to a G:T mismatch and monitoring the resulting increase in 2-aminopurine fluorescence in real time. DNA dissociation is measured by mixing pre-formed Msh2-Msh6 G:T(2-Ap) mismatch complex with unlabeled trap DNA and monitoring decrease in fluorescence over time(9). Pre-steady state ATPase kinetics are measured by the change in fluorescence of 7-diethylamino-3-((((2-maleimidyl)ethyl)amino)carbonyl) coumarin)-labeled Phosphate Binding Protein (MDCC-PBP) on binding phosphate (Pi) released by Msh2-Msh6 following ATP hydrolysis(9,10). The data reveal rapid binding of Msh2-Msh6 to a G:T mismatch and formation of a long-lived Msh2-Msh6 G:T complex, which in turn results in suppression of ATP hydrolysis and stabilization of the protein in an ATP-bound form. The reaction kinetics provide clear support for the hypothesis that ATP-bound Msh2-Msh6 signals DNA repair on binding a mismatched base pair in the double helix. F
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http://dx.doi.org/10.3791/1874DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3168206PMC
March 2010

Saccharomyces cerevisiae Msh2-Msh6 DNA binding kinetics reveal a mechanism of targeting sites for DNA mismatch repair.

Proc Natl Acad Sci U S A 2010 Jan 22;107(2):680-5. Epub 2009 Dec 22.

Molecular Biology and Biochemistry Department, Wesleyan University, 205 Hall-Atwater Laboratories, Middletown, CT 06459, USA.

The DNA mismatch repair system (MMR) identifies replication errors and damaged bases in DNA and functions to preserve genomic integrity. MutS performs the task of locating mismatched base pairs, loops and lesions and initiating MMR, and the fundamental question of how this protein targets specific sites in DNA is unresolved. To address this question, we examined the interactions between Saccharomyces cerevisiae Msh2-Msh6, a eukaryotic MutS homolog, and DNA in real time. The reaction kinetics reveal that Msh2-Msh6 binds a variety of sites at similarly fast rates (k (ON) approximately 10(7) M(-1) s(-1)), and its selectivity manifests in differential dissociation rates; e.g., the protein releases a 2-Aminopurine:T base pair approximately 90-fold faster than a G:T mismatch. On releasing the 2-Ap:T site, Msh2-Msh6 is able to move laterally on DNA to locate a nearby G:T site. The long-lived Msh2-Msh6.G:T complex triggers the next step in MMR--formation of an ATP-bound clamp--more effectively than the short-lived Msh2-Msh6.2-Ap:T complex. Mutation of Glu in the conserved Phe-X-Glu DNA binding motif stabilizes Msh2-Msh6(E339A).2-Ap:T complex, and the mutant can signal 2-Ap:T repair as effectively as wild-type Msh2-Msh6 signals G:T repair. These findings suggest a targeting mechanism whereby Msh2-Msh6 scans DNA, interrogating base pairs by transient contacts and pausing at potential target sites, and the longer the pause the greater the likelihood of MMR.
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http://dx.doi.org/10.1073/pnas.0908302107DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2818965PMC
January 2010

Mechanism of MutS searching for DNA mismatches and signaling repair.

J Biol Chem 2008 Dec 14;283(52):36646-54. Epub 2008 Oct 14.

Department of Chemistry and Curriculum in Applied Sciences and Engineering, University of North Carolina, Chapel Hill, North Carolina 27599, USA.

DNA mismatch repair is initiated by the recognition of mismatches by MutS proteins. The mechanism by which MutS searches for and recognizes mismatches and subsequently signals repair remains poorly understood. We used single-molecule analyses of atomic force microscopy images of MutS-DNA complexes, coupled with biochemical assays, to determine the distributions of conformational states, the DNA binding affinities, and the ATPase activities of wild type and two mutants of MutS, with alanine substitutions in the conserved Phe-Xaa-Glu mismatch recognition motif. We find that on homoduplex DNA, the conserved Glu, but not the Phe, facilitates MutS-induced DNA bending, whereas at mismatches, both Phe and Glu promote the formation of an unbent conformation. The data reveal an unusual role for the Phe residue in that it promotes the unbending, not bending, of DNA at mismatch sites. In addition, formation of the specific unbent MutS-DNA conformation at mismatches appears to be required for the inhibition of ATP hydrolysis by MutS that signals initiation of repair. These results provide a structural explanation for the mechanism by which MutS searches for and recognizes mismatches and for the observed phenotypes of mutants with substitutions in the Phe-Xaa-Glu motif.
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http://dx.doi.org/10.1074/jbc.M805712200DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2606009PMC
December 2008