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"She, Min"
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Endoplasmic reticulum-associated N-glycan degradation of cold-upregulated glycoproteins in response to chilling stress in Arabidopsis
N-glycosylation has a great impact on glycoprotein structure, conformation, stability, solubility, immunogenicity and enzyme activity. Structural characterization of N-glycoproteome has been challenging but can provide insights into the extent of protein folding and surface topology. We describe a highly sensitive proteomics method for large-scale identification and quantification of glycoproteins in Arabidopsis through 15N-metabolic labeling, selective enrichment of glycopeptides, data-dependent MS/MS analysis and automated database searching.
In-house databases of Arabidopsis glycoproteins and glycopeptides containing Asn-X-Ser/Thr/Cys motifs were constructed by reducing 20% and 90% of the public database size, respectively, to enable a rapid analysis of large datasets for comprehensive identification and quantification of glycoproteins and heterogeneous N-glycans in a complex mixture.
Proteome-wide analysis identified c. 100 stress-related N-glycoproteins, of which the endoplasmic reticulum (ER) resident proteins were examined to be up-regulated. Quantitative measurements provided a molecular signature specific to glycoproteins for determining the degree of plant stress at low temperature.
Structural N-glycoproteomics following time-course cold treatments revealed the stressresponsive degradation of high-mannose type N-glycans in ER in response to chilling stress, which may aid in elucidating the cellular mechanisms of protein relocation, transport, trafficking, misfolding and degradation under stress conditions.
Journal Article
Region-selective and site-specific glycation of influenza proteins surrounding the viral envelope membrane
Analysis of protein modifications is critical for quality control of therapeutic biologics. However, the identification and quantification of naturally occurring glycation of membrane proteins by mass spectrometry remain technically challenging. We used highly sensitive LC MS/MS analyses combined with multiple enzyme digestions to determine low abundance early-stage lysine glycation products of influenza vaccines derived from embryonated chicken eggs and cultured cells. Straightforward sequencing was enhanced by MS/MS fragmentation of small peptides. As a result, we determined a widespread distribution of lysine modifications attributed by the region-selectivity and site-specificity of glycation toward influenza matrix 1, hemagglutinin and neuraminidase. Topological analysis provides insights into the site-specific lysine glycation, localizing in the distinct structural regions of proteins surrounding the viral envelope membrane. Our finding highlights the proteome-wide discovery of lysine glycation of influenza membrane proteins and potential effects on the structural assembly, stability, receptor binding and enzyme activity, demonstrating that the impacts of accumulated glycation on the quality of products can be directly monitored by mass spectrometry-based structural proteomics analyses.
Journal Article
النساء في الكومونات الشعبي /
لم يكن التسلط على المرأة أمرا جديدا في المجتمع الصيني، فقد كان متوارثا، تستطيع الاستدلال عليه من الأمثال الشعبية التي تتناول مكانة المرأة ضمنه، كالقول الكونفوشي : أن تكون المرأة عديمة الموهبة، فتلك فضيلة، أو كحقيقة أن بعضهن كبرن دون أن يتمكنَّ من الخروج من بيوتهن على الإطلاق إلا صوب بيوت أزواجهن أو القبر.
Book
Autophagy induction contributes to the resistance to methotrexate treatment in rheumatoid arthritis fibroblast-like synovial cells through high mobility group box chromosomal protein 1
Background
Rheumatoid arthritis fibroblast-like synovial cells (RA-FLS) show resistance to methotrexate (MTX) treatment. To better understand the mechanisms of this resistance, RA-FLS and osteoarthritis fibroblast-like synovial cells (OA-FLS) were isolated and exposed to MTX. We analyzed the autophagy induced by MTX in vitro and its relationship to apoptosis.
Methods
Cell viability was evaluated using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, and apoptosis was detected by flow cytometry and Western blot analysis. Autophagy was determined by transmission electron microscopy as well as Western blot analysis. The expression levels of Beclin-1, LC3, Akt, p-Akt, mammalian target of rapamycin (mTOR), p-mTOR, high mobility group box chromosomal protein 1 (HMGB1), and an 85 kDa caspase cleaved fragment of poly(ADP-ribose) polymerase were measured by Western blotting.
Results
MTX-induced apoptosis was increased in OA-FLS compared with RA-FLS. However, MTX stimulated the autophagy response in RA-FLS by inducing autophagosome formation, but not in OA-FLS. In RA-FLS, transfection with Beclin-1 small interfering RNA inhibited autophagy and increased susceptibility to MTX, which induces cell death. MTX upregulated autophagy through its ability to enhance the expression of HMGB1 and Beclin-1 rather than through the Akt/mTOR pathway.
Conclusions
Autophagy induction contributes to resistance to MTX treatment in fibroblasts from patients with rheumatoid arthritis.
Journal Article
حول (الاستعمار وكل الرجعيين نمور من ورق) /
يتناول كتاب (ماوتسي يونج) وهو صاحب سيرة طويلة عبر ما يقرب من سبعين عاما، نشأ في ريف الصين لأب فلاح فقير، استهوته الماركسية فانتمى إليها، ثم صار أحد نجومها في الحزب الشيوعي الصيني، ثم صار رئيسا للحزب، ثم استقل بعرش الصين وجلس عليه حوالي ثلاثين عاما، قاد فيها الصين برؤيته الخاصة فصنع منها دولة قوية في وقت وجيز، ولا يزال كتابه الأحمر مرجعا أساسيا للفكر الصيني والسياسة الصينية، بل إن دواوين شعره هي الأكثر مبيعا في الصين، ولا تزال سيرته مقصد كثير من المطلعين.
Book
Significance of the Balance between Regulatory T (Treg) and T Helper 17 (Th17) Cells during Hepatitis B Virus Related Liver Fibrosis
Hepatitis B virus-related liver fibrosis (HBV-LF) always progresses from inflammation to fibrosis. However, the relationship between these two pathological conditions is not fully understood. Here, it is postulated that the balance between regulatory T (Treg) cells and T helper 17 (Th17) cells as an indicator of inflammation may predict fibrosis progression of HBV-LF.
The frequencies and phenotypes of peripheral Treg and Th17 cells of seventy-seven HBeAg-positive chronic hepatitis B (CHB) patients who underwent liver biopsies and thirty healthy controls were determined by flow cytometry. In the periphery of CHB patients, both Treg and Th17 frequencies were significantly increased and correlated, and a lower Treg/Th17 ratio always indicated more liver injury and fibrosis progression. To investigate exact effects of Treg and Th17 cells during HBV-LF, a series of in vitro experiments were performed using purified CD4(+), CD4(+)CD25(+), or CD4(+)CD25(-) cells from the periphery, primary human hepatic stellate cells (HSCs) isolated from healthy liver specimens, human recombinant interleukin (IL)-17 cytokine, anti-IL-17 antibody and HBcAg. In response to HBcAg, CD4(+)CD25(+) cells significantly inhibited cell proliferation and cytokine production (especially IL-17 and IL-22) by CD4(+)CD25(-) cells in cell-contact and dose-dependent manners. In addition, CD4(+) cells from CHB patients, compared to those from HC subjects, dramatically promoted proliferation and activation of human HSCs. Moreover, in a dramatically dose-dependent manner, CD4(+)CD25(+) cells from CHB patients inhibited, whereas recombinant IL-17 response promoted the proliferation and activation of HSCs. Finally, in vivo evidence about effects of Treg/Th17 balance during liver fibrosis was obtained in concanavalin A-induced mouse fibrosis models via depletion of CD25(+) or IL-17(+) cells, and it's observed that CD25 depletion promoted, whereas IL-17 depletion, alleviated liver injury and fibrosis progression.
The Treg/Th17 balance might influence fibrosis progression in HBV-LF via increase of liver injury and promotion of HSCs activation.
Journal Article
Associations among microbial enterotype, brain structure, and working memory: A combined structural and diffusion MRI study
•Three enterotypes showed differences in cortical thickness of the prefrontal cortex.•Three enterotypes differed in mean diffusivity of the cerebral peduncle and cingulum.•Prefrontal cortical thickness mediated the enterotype-working memory association.
Enterotype analysis classifies individuals based on gut microbial community composition using clustering techniques. Despite evidence suggesting the important role of enterotype in affecting brain function and working memory, little is known about the brain structural substrates.
We collected fecal samples and utilized 16S rDNA amplicon sequencing to identify three enterotypes (Bacteroides, Prevotella, and Ruminococcaceae) among 511 healthy young adults through unsupervised clustering. Structural and diffusion MRI techniques were adopted to assess gray matter morphology and white matter integrity. Inter-enterotype differences in brain structure were tested, followed by correlation and mediation analyses to investigate the potential relationships among enterotype, brain structure, and working memory.
The three enterotypes exhibited significant differences in cortical thickness of the prefrontal cortex and mean diffusivity of the cerebral peduncle and cingulum. Moreover, prefrontal cortical thickness was correlated with working memory and further acted as a significant mediator of the association between enterotype and working memory.
Our findings may contribute to the growing literature on the microbiota-brain-cognition relationship, setting the stage for future longitudinal and interventional research.
Journal Article
Highly sensitive characterization of non-human glycan structures of monoclonal antibody drugs utilizing tandem mass spectrometry
Glycosylation is an important attribute of monoclonal antibodies (mAbs) for assessing manufacturing quality. Analysis of non-human glycans containing terminal galactose-α1,3-galactose and
N
-glycolylneuraminic acid is essential due to the potential immunogenicity and insufficient efficacy caused by mAb expression in non-human mammalian cells. Using parallel sequencing of isobaric glycopeptides and isomeric glycans that were separated by reversed-phase and porous graphitic carbon LC, we report a highly sensitive LC MS/MS method for the comprehensive characterization of low-abundance non-human glycans and their closely related structural isomers. We demonstrate that the straightforward use of high-abundance diagnostic ions and complementary fragments under the positive ionization low-energy collision-induced dissociation is a universal approach to rapidly discriminate branch-linkage structures of biantennary glycans. Our findings reveal the structural diversity of non-human glycans and sulfation of α-galactosylated glycans, providing both an analytical method and candidate structures that could potentially be used in the crucial quality control of therapeutic mAb products.
Journal Article