高迁移率族蛋白

高迁移率族蛋白(highmobilitygroupprotein,HMG蛋白)广泛存在于真核生物细胞中,因其在聚丙稀凝胶电泳中的高迁移率而得名。HMG蛋白是真核细胞基因调控的动力体现者,是真核细胞内继组蛋白之后含量最为丰富的一组染色质蛋白质,它们在染色质的结构与功能及基因表达调控过程中均发挥着重要作用。HMG蛋白家族可分为HMGA、HMGB和HMGN三类亚家族。现对HMG蛋白家族的三类亚家族蛋白HMGA、HMGB和HMGN的结构与功能进行综述。     

胶质瘤来源外泌体通过高迁移率族蛋白B1促进胶质瘤干细胞形成

摘要 目的：研究胶质瘤来源外泌体中高迁移率族蛋白B1（HMGB1）对胶质瘤干细胞形成的影响及其意义。方法：使用外泌体提取试剂盒提取原代胶质母细胞瘤来源外泌体,通过透射电子显微镜、纳米粒度电位仪和Western blotting对外泌体进行鉴定;采用Western blotting检测外泌体中HMGB1的表达量;通过qRT-PCR、Western blotting、克隆球计数检测外泌体对胶质瘤干细胞形成的影响;siRNA敲低HMGB1的表达水平,并通过qRT-PCR、Western blotting、克隆球计数检测外泌体中HMGB1对胶质瘤干细胞形成的影响。结果：原代胶质瘤细胞可以分泌外泌体到肿瘤微环境并且外泌体中存在HMGB1;原代胶质瘤细胞来源外泌体可以上调邻近胶质瘤细胞干性相关分子CD133、OCT4、NANOG、SOX2的表达并促进干细胞克隆球的形成;通过siRNA敲低原代胶质瘤细胞HMGB1的表达后,外泌体中HMGB1的含量降低并且外泌体促进胶质瘤干细胞形成的作用减弱。结论：胶质瘤细胞来源外泌体可以通过HMGB1促进胶质瘤干细胞的形成。     

Sequence of a cDNA Encoding Turtle High Mobility Group 1 Protein

In order to understand sequence information about turtle HMG1 gene, a cDNA encoding HMG1 protein of the Chinese soft-shell turtle (Pelodiscus sinensis) was amplified by RT-PCR from kidney total RNA, and was cloned, sequenced and analyzed. The results revealed that the open reading frame (ORF) of turtle HMG1 cDNA is 606 bp long. The ORF codifies 202 amino acid residues, from which two DNA-binding domains and one polyacidic region are derived. The DNA-binding domains share higher amino acid identity with homologous sequences of chicken (96.5%) and mammals (74%) than homologous sequence of rainbow trout (67%). The polyacidic region shows 84.6% amino acid homology with the equivalent region of chicken HMG1 cDNA. Turtle HMG1 protein contains 3 Cys residues located at completely conserved positions. Conservation in sequence and structure suggests that the functions of turtle HMG1 cDNA may be highly conserved during evolution. To our knowledge, this is the first report of HMG1 cDNA sequence in any reptilian.From Genetika, Vol. 41, No. 7, 2005, pp. 925–930.Original English Text Copyright © 2005 by Jifang Zheng, Bi Hu, Duansheng Wu.The text was submitted by the authors in English.     

Biased Transmission of Sex Chromosomes in the Aphid Myzus persicae Is Not Associated with Reproductive Mode

Commonly, a single aphid species exhibits a wide range of reproductive strategies including cyclical parthenogenesis and obligate parthenogenesis. Sex determination in aphids is chromosomal; females have two X chromosomes, while males have one. X chromosome elimination at male production is generally random, resulting in equal representation of both X chromosomes in sons. However, two studies have demonstrated deviations from randomness in some lineages. One hypothesis to account for such deviations is that recessive deleterious mutations accumulate during bouts of asexual reproduction and affect male viability, resulting in overrepresentation of males with the least deleterious of the two maternal X chromosomes. This hypothesis results in a testable prediction: X chromosome transmission bias will increase with time spent in the asexual phase and should therefore be most extreme in the least sexual aphid life cycle class. Here we test this prediction in Myzus persicae. We used multiple heterozygous X-linked microsatellite markers to screen 1085 males from 95 lines of known life cycle. We found significant deviations from equal representation of X chromosomes in 15 lines; however, these lines included representatives of all life cycles. Our results are inconsistent with the hypothesis that deviations from randomness are attributable to mutation accumulation.     

急性肺损伤大鼠肺组织高迁移率族蛋白1的表达及其意义

目的动态观察高迁移率族蛋白1（HGMB1）在失血性休克复合内毒素注射致急性肺损伤（ALl）大鼠肺组织的表达情况,初步探讨HMGB1在ALI发病机制中的作用。方法采取失血性休克复合内毒素注射手段建立ALl大鼠动物模型,采用RT-PCR方法,检测肺组织HMGB1mRNA的表达情况。结果正常大鼠肺组织有少量HMGBlmRNA表达,遭受失血性休克复合内毒素注射打击后,HMGB1mRNA表达迅速升高,至ALI24h达最高峰,随后有所下降,ALl各组大鼠表达水平与正常对照组比较差异均有统计学意义（P〈0．01）。结论正常大鼠肺组织有一定水平HMGBlmRNA的表达,遭受失血性休克及内毒素注射打击后,HMGBlmRNA表达异常增高,可引起过度炎症反应,从而促进ALI的发生与发展。     

高迁移率族蛋白B1在炎症性疾病中的研究进展

高迁移率族蛋白B1(high mobility group box 1 protein,HMGB1)是广泛存在于真核细胞核内的非组蛋白染色体结合蛋白,因其在聚丙烯酰胺凝胶电泳(PAGE)中迁移速度快而得名。近年来的研究表明,HMGB1作为一种重要的晚期炎症介质,在多种急慢性炎症中均有表达。本文就高迁移率族蛋白B1的结构、释放、致炎作用、与炎症性疾病的关系以及炎症时对高迁移率族蛋白B1的干预措施等方面研究近况做一综述。     

High SEPT9_i1 Protein Expression Is Associated with High-Grade Prostate Cancers

Septins are a family of GTP-binding cytoskeleton proteins expressed in many solid tumors. Septin 9 (SEPT9) in particular was found overexpressed in diverse carcinomas. Herein, we studied the expression of SEPT9 isoform 1 protein (SEPT9_i1) in human prostate cancer specimens. We utilized immunohistochemical staining to study the expression of SEPT9_i1 protein. Staining level was analyzed in association with clinical characteristics and the pathological Gleason grade and score. Fifty human prostate cancer specimens (42 primary tumors and 8 metastatic lesions) were stained by SEPT9_i1 antibody and analyzed. SEPT9_i1 protein was expressed in prostate cancer cells but absent in normal epithelial cells. The intensity of staining was correlated proportionally to pretreatment prostate-specific antigen (PSA) blood levels and Gleason score (P < 0.05). SEPT9_i1 was highly expressed in all metastatic lesions. A significant assocation between SEPT9_i1 expression and high Gleason score on multivariate linear regression analysis was found. We conclude that SEPT9_i1 is expressed in high-grade prostate tumors suggesting it has a significant role in prostate tumorigenesis and that it could serve as a molecular marker for prostate tumor progression.     

Cellular Mechanisms of High Mobility Group 1 (HMGB-1) Protein Action in the Diabetic Retinopathy

Diabetic retinopathy is one of the main microvascular complications of diabetes and remains one of the leading causes of blindness worldwide. Recent studies have revealed an important role of inflammatory and proangiogenic high mobility group 1 (HMGB-1) cytokine in diabetic retinopathy. To elucidate cellular mechanisms of HMGB-1 activity in the retina, we performed this study. The histological features of diabetic retinopathy include loss of blood-vessel pericytes and endothelial cells, as well as abnormal new blood vessel growth. To establish the role of HMGB-1 in vulnerability of endothelial cells and pericytes, cultures of these cells, or co-cultures with glial cells, were treated with HMGB-1 and assessed for survival after 24 hours. The expression levels of the cytokines, chemokines, and cell adhesion molecules in glial and endothelial cells were tested by quantitative RT-PCR to evaluate changes in these cells after HMGB-1 treatment. Animal models of neovascularization were also used to study the role of HMGB-1 in the retina. We report that pericyte death is mediated by HMGB-1-induced cytotoxic activity of glial cells, while HMGB-1 can directly mediate death of endothelial cells. We also found that HMGB-1 affects endothelial cell activity. However, we did not observe a difference in the levels of neovascularization between HMGB-1-treated eyes compared to the control eyes, nor in the levels of proangiogenic cytokine VEGF-A expression between glial cells treated with HMGB-1 and control cells. Our data also indicate that HMGB-1 is not involved in retinal neovascularization in the oxygen-induced retinopathy model. Thus, our data suggest that retinal pericyte and endothelial injury and death in diabetic retinopathy may be due to HMGB-1-induced cytotoxic activity of glial cells as well as the direct effect of HMGB-1 on endothelial cells. At the same time, our findings indicate that HMGB-1 plays an insignificant role in retinal and choroidal neovascularization.     

Potentiation of NMDA Receptor-Dependent Cell Responses by Extracellular High Mobility Group Box 1 Protein

Background

Extracellular high mobility group box 1 (HMGB1) protein can operate in a synergistic fashion with different signal molecules promoting an increase of cell Ca²⁺ influx. However, the mechanisms responsible for this effect of HMGB1 are still unknown.

Principal Findings

Here we demonstrate that, at concentrations of agonist per se ineffective, HMGB1 potentiates the activation of the ionotropic glutamate N-methyl-D-aspartate receptor (NMDAR) in isolated hippocampal nerve terminals and in a neuroblastoma cell line. This effect was abolished by the NMDA channel blocker MK-801. The HMGB1-facilitated NMDAR opening was followed by activation of the Ca²⁺-dependent enzymes calpain and nitric oxide synthase in neuroblastoma cells, resulting in an increased production of NO, a consequent enhanced cell motility, and onset of morphological differentiation. We have also identified NMDAR as the mediator of HMGB1-stimulated murine erythroleukemia cell differentiation, induced by hexamethylenebisacetamide. The potentiation of NMDAR activation involved a peptide of HMGB1 located in the B box at the amino acids 130–139. This HMGB1 fragment did not overlap with binding sites for other cell surface receptors of HMGB1, such as the advanced glycation end products or the Toll-like receptor 4. Moreover, in a competition assay, the HMGB1_(130–139) peptide displaced the NMDAR/HMGB1 interaction, suggesting that it comprised the molecular and functional site of HMGB1 regulating the NMDA receptor complex.

Conclusion

We propose that the multifunctional cytokine-like molecule HMGB1 released by activated, stressed, and damaged or necrotic cells can facilitate NMDAR-mediated cell responses, both in the central nervous system and in peripheral tissues, independently of other known cell surface receptors for HMGB1.     

Urinary Levels of High Mobility Group Box-1 Are Associated with Disease Activity in Antineutrophil Cytoplasmic Autoantibody-Associated Vasculitis

Background

High mobility group box-1 (HMGB1), a kind of pro-inflammatory mediator, is associated with inflammatory conditions and tissue damage. Our previous study demonstrated that the circulating levels of HMGB1 correlated with disease activity of antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV). In the current study, we aimed to measure urinary levels of HMGB1 in AAV patients, correlated them to clinical activity index and analysed the immunohistochemical HMGB1 staining in kidney specimens.

Methods

50 patients with AAV in active stage and 56 patients with AAV in remission were recruited. The urinary levels of HMGB1 were determined by enzyme-linked immunosorbent assay. Moreover, renal biopsy specimens from 27 patients with active AAV were randomly collected to evaluate the deposition of HMGB1.

Results

Urinary HMGB1 levels in AAV patients in active stage were significantly higher than those in AAV patients in remission and healthy controls (1.46 [0.56-3.43] versus 0.38 [0.10-1.35] mg/μmolCr, P=0.001; 1.46 [0.56-3.43] versus 0.48 [0.40-0.60] mg/μmolCr, P=0.000, respectively). Further analysis found that urinary levels of HMGB1 correlated with erythrocyte sedimentation rate (r=0.354, p=0.012), C-reactive protein (r=0.289, p=0.042), and Birmingham Vasculitis Activity Score (r=0.350, p=0.013). Renal tissue of active AAV patients showed HMGB1 was mainly expressed in the cytoplasm and the extracellular space. The percentage of HMGB1-negative nuclei in renal tissue of patients with active AAV was significantly higher than that in normal controls (60.6±20.2 % versus 2.7±0.6 %, p<0.01).

Conclusion

Urinary levels of HMGB1 may be associated with the disease activity in AAV patients.     

Birth of Cone Bipolar Cells,but Not Rod Bipolar Cells,Is Associated with Existing RGCs

Retinal ganglion cells (RGCs) play important roles in retinogenesis. They are required for normal retinal histogenesis and retinal cell number balance. Developmental RGC loss is typically characterized by initial retinal neuronal number imbalance and subsequent loss of retinal neurons. However, it is not clear whether loss of a specific non-RGC cell type in the RGC-depleted retina is due to reduced cell production or subsequent degeneration. Taking advantage of three knockout mice with varying degrees of RGC depletion, we re-examined bipolar cell production in these retinas from various aspects. Results show that generation of the cone bipolar cells is correlated with the existing number of RGCs. However, generation of the rod bipolar cells is unaffected by RGC shortage. Results report the first observation that RGCs selectively influence the genesis of subsequent retinal cell types.     

Role of the Acidic Tail of High Mobility Group Protein B1 (HMGB1) in Protein Stability and DNA Bending

High mobility group box (HMGB) proteins are abundant nonhistone proteins found in all eukaryotic nuclei and are capable of binding/bending DNA. The human HMGB1 is composed of two binding motifs, known as Boxes A and B, are L-shaped alpha-helix structures, followed by a random-coil acidic tail that consists of 30 Asp and Glu residues. This work aimed at evaluating the role of the acidic tail of human HMGB1 in protein stability and DNA interactions. For this purpose, we cloned, expressed and purified HMGB1 and its tailless form, HMGB1ΔC, in E. coli strain. Tryptophan fluorescence spectroscopy and circular dichroism (CD) experiments clearly showed an increase in protein stability promoted by the acidic tail under different conditions, such as the presence of the chemical denaturant guanidine hydrochloride (Gdn.HCl), high temperature and low pH. Folding intermediates found at low pH for both proteins were denatured only in the presence of chemical denaturant, thus showing a relatively high stability. The acidic tail did not alter the DNA-binding properties of the protein, although it enhanced the DNA bending capability from 76° (HMGB1ΔC) to 91° (HMGB1), as measured using the fluorescence resonance energy transfer technique. A model of DNA bending in vivo was proposed, which might help to explain the interaction of HMGB1 with DNA and other proteins, i.e., histones, and the role of that protein in chromatin remodeling.     

Somatic Chromosomes of the Rabbit

The number of chromosomes of the rabbit was determined at 44, and both the X and the Y chromosomes have submedially situated centromeres. X is of medium size while Y is one of the smallest chromosomes of the complement. For practical purposes, the chromosomes have been divided into groups according to their appearance.     

EF-1[alpha] Is Associated with a Cytoskeletal Network Surrounding Protein Bodies in Maize Endosperm Cells

By using indirect immunofluorescence and confocal microscopy, we documented changes in the distribution of elongation factor-1[alpha] (EF-1[alpha]), actin, and microtubules during the development of maize endosperm cells. In older interphase cells actively forming starch grains and protein bodies, the protein bodies are enmeshed in EF-1[alpha] and actin and are found juxtaposed with a multidirectional array of microtubules. Actin and EF-1[alpha] appear to exist in a complex, because we observed that the two are colocalized, and treatment with cytochalasin D resulted in the redistribution of EF-1[alpa]. These data suggest that EF-1[alpha] and actin are associated in maize endosperm cells and may help to explain the basis of the correlation we found between the concentration of EF-1[alpha] and lysine content. The data also support the hypothesis that the cytoskeleton plays a role in storage protein deposition. The distributions of EF-1[alpha] actin, and microtubules change during development. We observed that in young cells before the accumulation of starch and storage protein, EF-1[alpha], actin, and microtubules are found mainly in the cell cortex or in association with nuclei.