Evolution of the interleukin-1 gene family in mammals

The phylogeny of interleukin-1 family genes shows that human interleukin-1 (IL-1) is more closely related to IL-1 of the bovine than to IL-1 of the mouse, whereas human interleukin-1 (IL-1) is more closely related to IL-1 of the mouse than to IL-1 of the bovine. The IL-1 receptor antagonist (IL-1) shows homology to the C-terminal region of both IL-1 and IL-1. In the C-terminal region, the IL-1 genes of human and mouse have diverged more from each other at nonsynonymous sites than have either IL-1 or IL-1; because the same pattern is not seen at synonymous sites, it must be due not to a difference in mutation rate but rather to a greater degree of functional constraint on this region in the IL-1 and IL-1 proteins than in the IL-1 protein. But synonymous sites in IL-1 of mouse have evolved more rapidly than in IL-1 of human, indicating a higher rate of mutation in the former gene. In the N-terminal region of the protein, nonsynonymous sites have evolved at similar rates in IL-1 and IL-1. The first exon of the IL-1 gene, which encodes the leader peptide, shows evidence of homology with the first exon of IL-1, which is not translated. Thus, it seems likely that IL-1 evolved by duplication of an IL-1 gene and loss of expression of exons 2–4. Correspondence to: A.L. Hughes     

猕猴桃AcSWEET基因家族的鉴定与表达分析

SWEET（Sugars will eventually be exported transporters）是近年来在植物中发现的一组糖转运蛋白,在植物生长、发育和非生物及生物胁迫响应等多种生理过程中发挥着重要作用。本研究利用生物信息学方法对猕猴桃（Actinidia chinensis Planch.）AcSWEET基因家族进行了鉴定,共获得29个AcSWEET基因,并对其氨基酸数量、相对分子量、等电点、不稳定系数、亚细胞定位、亲水指数进行了分析。结果显示：29个基因编码的氨基酸数目为680~906个;分子量范围为7.531~101.266 kDa,等电点在6.95~9.90,多数蛋白为定位于细胞膜的疏水性蛋白,具有1~2个MtN3结构域或PQ-loop结构域。此外,AcSWEET基因的外显子数量在4~6个,系统进化分析结果表明猕猴桃AcSWEET基因家族被分为4个亚族,同一亚族基因具有相似的内含子、外显子以及保守基序。表达模式分析结果表明,这些基因在果实不同发育时期具有表达特异性。推测AcSWEET26、AcSWEET7、AcSWEET15和AcSWEET13可能参与猕猴桃的蔗糖转运和积累。     

Ricerche embriologiche e cariologiche sul genere “Euphorbia„

Riassunto

L'A. stabilisce che in Euphorbia Paralias L., in E. falcata L. var. acuminata (Lam.) Fior, E. pubescens Vahl ed E. Pithyusa L. var. ovalifolia Fiori lo sviluppo del gametofito femminile è di tipo Normale S-nucleato; in Euphorbia Characias L. il gametofito femminile può svilupparsi secondo il tipo Normale e il tipo Scilla con una frequenza di sviluppo del primo rispetto al secendo del valore approssimativo percentuale del 7%; in Euphorbia amygdaloides L. il gametofito si sviluppa sempre secondo lo schema del tipo Scilla. Stabilisce inoltre che il numero aploide dei cromosomi è n=S in Euphorbia Paralias L., n = 18 in E. falcala var. acuminata (Lam.) Fiori, n = 7 in E. pubescens Vahl, n = 18 in E. Pithyusa L. var. ovalifolia Fiori, n = 10 in E. Characias L. ed n = 9 in E. amygdaloides L. Fenomeni di associazione secondaria dei cromosomi del corredo aploide potrebbero indicare che il numero aploide n = 7 di Euphorbia pubcscens Vahl sia derivato dal numero aploide 6. Si prospetta quindi la possibilità dell'esistenza nel genere Euphorbia di una serie poliploide secondaria con termini 7 - 14 - 21 - 28, rispettivamente diploidi, tetraploidi, esaploidi ed ottoploidi.     

Population genetics theory of concerted evolution and its application to the immunoglobulin V gene tree

Summary The previous simple model for treating concerted evolution of multigene families has been revised to be compatible with various new observations on the immunoglobulin variable region family and other families. In the previous model, gene conversion and unequal crossing-over were considered, and it was assumed that genes are randomly arranged on the chromosome; neither subdivision nor correlation of gene identity and chromosomal distance were considered. Although this model satisfactorily explains the observed amino acid diversity within and between species, it fails to predict the very ancient branching of the mouse immunoglobulin heavy chain V-gene family. By incorporating subdivided structure and genetic correlation with chromosomal distance into the simple model, the data of divergence may be satisfactorily explained, as well as the rate of nucleotide substitution and the amino acid diversity. The rate at which a V-gene is duplicated or deleted by conversion or by unequal crossing-over is estimated by the new model to be on the order of 10^–6 per year. The model may be applicable to other multigene families, such as those coding for silkmoth chorion or mammalian kallikrein.Contribution no. 1560 from the National Institute of Genetics, Mishima, 411 Japan     

Development and validation of an antibody-dependent cell-mediated cytotoxicity-reporter gene assay

Humanized monoclonal antibodies (mAbs) are the fastest growing class of biological therapeutics that are being developed for various medical indications, and more than 30 mAbs are already approved and in the market place. Antibody-dependent cell-mediated cytotoxicity (ADCC) is an important biological function attributed to the mechanism of action of several therapeutic antibodies, particularly oncology targeting mAbs. The ADCC assay is a complicated and highly variable assay. Thus, the use of an ADCC assay as a lot release test or a stability test for clinical trial batches of mAbs has been a substantial challenge to install in quality control laboratories. We describe here the development and validation of an alternate approach, an ADCC-reporter gene assay that is based on the key attributes of the PBMC-based ADCC assay. We tested the biological relevance of this assay using an anti-CD20 based model and demonstrated that this ADCC-reporter assay correlated well with standard ADCC assays when induced with the drugable human isotypes [IgG1, IgG2, IgG4, IgG4S > P (S228P) and IgG4PAA (S228P, F234A, L235A)] and with IgG1 isotype variants with varying amounts of fucosylation. This data demonstrates that the ADCC-reporter gene assay has performance characteristics (accuracy, precision and robustness) to be used not only as a potency assay for lot release and stability testing for antibody therapeutics, but also as a key assay for the characterization and process development of therapeutic molecules.     

Development and validation of an antibody-dependent cell-mediated cytotoxicity-reporter gene assay

Humanized monoclonal antibodies (mAbs) are the fastest growing class of biological therapeutics that are being developed for various medical indications, and more than 30 mAbs are already approved and in the market place. Antibody-dependent cell-mediated cytotoxicity (ADCC) is an important biological function attributed to the mechanism of action of several therapeutic antibodies, particularly oncology targeting mAbs. The ADCC assay is a complicated and highly variable assay. Thus, the use of an ADCC assay as a lot release test or a stability test for clinical trial batches of mAbs has been a substantial challenge to install in quality control laboratories. We describe here the development and validation of an alternate approach, an ADCC-reporter gene assay that is based on the key attributes of the PBMC-based ADCC assay. We tested the biological relevance of this assay using an anti-CD20 based model and demonstrated that this ADCC-reporter assay correlated well with standard ADCC assays when induced with the drugable human isotypes [IgG1, IgG2, IgG4, IgG4S > P (S228P) and IgG4PAA (S228P, F234A, L235A)] and with IgG1 isotype variants with varying amounts of fucosylation. This data demonstrates that the ADCC-reporter gene assay has performance characteristics (accuracy, precision and robustness) to be used not only as a potency assay for lot release and stability testing for antibody therapeutics, but also as a key assay for the characterization and process development of therapeutic molecules.     

大豆miR-171基因家族的进化与功能分析

运用生物信息学方法在miRBase搜索大豆miR-171(gma-miR-171)基因家族的序列,分析gma-miR-171序列的进化特征并预测其靶基因。结果表明,在miRBase中共搜索到21条gma-miR-171基因家族序列。序列分析发现,gma-miR-171基因家族序列保守性较差,只有2个碱基完全保守。对gma-miR-171基因进行定位,21个成员分散在12条染色体上,其中Chr06上gma-miR-171基因最多,共4个。进化分析表明,位于同一条染色体上的gma-miR-171基因没有表现出较近的亲缘关系。靶基因预测获得14个gma-miR-171基因家族的靶基因,包括蛋白激酶、磷酸酶、输出蛋白、转录因子等,说明gma-miR-171基因家族在大豆中具有广泛的调控功能。     

人工microRNA干扰DREB亚族A-5组转录抑制子基因增强了拟南芥对低温和高盐胁迫的耐受性

转录抑制子是一类与DNA的特异位点结合并抑制其附近基因转录的蛋白质,主要通过与转录激活子或基本转录复合物发生作用以及引起染色质重排等3种方式来抑制目标基因的转录.DREB类转录因子的A-5组中共有6个转录抑制子,其功能和作用机制还有待进一步研究.通过设计人工microRNA-A5(amiRNA-A5)使其较特异地作用这...     

朱红大杜鹃MADS-box基因家族的全基因组鉴定与特征分析

MADS-box是在植物生长发育过程中扮演重要角色的一类转录因子,特别是花器官形成、开花时间控制及果实发育与成熟等过程。本研究基于朱红大杜鹃（Rhododendron griersonianum Balf. f. et Forrest）全基因组测序数据,利用生物信息学方法从中鉴定出 81 个 MADS-box 基因,并进行了分析。根据系统发育关系和蛋白结构将 MADS-box分为两类：Type-Ⅰ型包含 24 个基因,Type-Ⅱ型包含57个基因,这些基因不均匀地分布于12条染色体上;81个MADS-box 基因中,存在 6 对片段复制和 1 对串联复制基因,且它们经历了纯化选择作用;同时,基因启动子区域含有光响应、植物生长、激素反应和胁迫响应等顺式作用元件。综上,本研究鉴定了朱红大杜鹃的MADS-box家族转录因子,为深入研究其 MADS-box 蛋白的生物学功能提供了基础。     

SPECIFIC CHANGES IN THE PANCREACTIC EXPRESSION OF THE INTERLEUKIN 1 FAMILY OF GENES DURING EXPERIMENTAL ACUTE PANCREATITIS

Interleukin 1β (IL-1β) is produced in large amounts during acute pancreatitis and is believed to play a primary role in determining pancreatitis severity and the degree of pancreatic tissue destruction. This study was undertaken to characterize intrapancreatic production of IL-1β and the remainder of the IL-1 family of genes during sterile acute pancreatitis. Moderate or severe necrotizing pancreatitis was induced by the intraperitoneal injection of a cholecystokinin analogue or the feeding of a choline deficient diet, respectively. Animals were killed during the progression of pancreatitis with severity scored by histological grading and serum amylase concentration. The expression of IL-1β, IL-1 Receptor 1 (IL-1R1), Il-1R2, IL-1R antagonist (IL-1Ra), and ICE mRNA within the pancreas was examined by quantitative differential RT-PCR. Corresponding intrapancreatic and serum proteins were measured by enzyme-linked immunosorbent assay (ELISA). There was constitutive expression of pancreatic IL-1R1, IL-1R2, IL-1Ra, and ICE but not IL-1β. As pancreatitis developed, mRNA for IL-1β, IL-1Ra, and ICE increased in parallel with the degree of pancreatitis severity (allP<0.001 vs baseline) while mRNA for both receptors remained stable (P=NS). Intrapancreatic and systemic IL-1β and IL-1Ra protein also increased as pancreatitis developed (bothP<0.001) with tissue levels being continuously greater than serum. This study demonstrated that sterile, endotoxin-free acute pancreatitis induces the upregulation of specific members of the IL-1 family of genes including production of large amounts of IL-1β and its receptor antagonist within the pancreatic parenchyma. These changes are indicative of pancreatitis severity and are not model dependent.     

Dexamethasone induces an increase in intracellular and membrane-associated lipocortin-1 (annexin-1) in rat astrocyte primary cultures

Summary 1. The antiinflammatory actions of glucocorticoid steroids are thought to occur through induction of the protein lipocortin-1 (LC-1; annexin-1). The purpose of the current study was to investigate whether astrocytic LC-1 content was increased in the presence of a synthetic glucocorticoid, dexamethasone.2. Steroid-induced changes in cellular levels of LC-1 in astrocytes were determined by electrotransfer and immunoblotting techniques. Separate cell fractions were investigated to study the influence of dexamethasone on astroglial LC-1 content. The effect of culture state on LC-1 expression was also examined.3. Intracellular LC-1 content was found to decrease after initiation of culture, with a substantial rise in both cell proliferation and LC-1 expression occurring after the replenishment of medium containing steroid-free serum. A further increase in intracellular LC-1 occurred upon incubation with dexamethasone. The glucocorticoid-induced change in intracellular LC-1 was a time-dependent event and coincided with an increase in membrane-associated LC-1.4. The findings in this study indicate that astrocytic LC-1 content is influenced by cell culture conditions and, in the presence of glucocorticoid steroids, the cellular localization of LC-1 is altered. This may indicate that LC-1 has functions at more than one cellular locality.     

植物ASR基因研究进展

ASR（abscisic acid,stress,ripening-induced）基因是近年来从植物中发现的一类受ABA、胁迫和成熟诱导表达的基因,具有保守的ABA/WDS结构域。ASR基因不仅参与植物对干旱、高盐、低温以及脱落酸的胁迫应答,而且参与植物生命活动的许多过程,如果实发育、成熟和糖代谢等。本文综述了近年来国内外ASR基因的研究进展,主要包括ASR基因和蛋白结构特点、ASR基因家族的进化、ASR基因的表达及可能具有的功能,为植物ASR基因研究提供参考。     

PLAG基因家族研究进展

多形性腺瘤基因(pleomorphic adenoma gene, PLAG)家族包括PLAG1、PLAGL1和PLAGL2共3个成员,为锌指蛋白的新亚科。多项研究表明,PLAG家族蛋白作为核蛋白转录调节因子,主要对机体多种重要基因表达发挥调控作用。本研究旨在介绍PLAG家族蛋白的结构和生物学功能,并详细介绍PLAG家族在肿瘤治疗和动物育种中的作用和研究进展,以为今后的研究方向提供参考。     

GSTT1 、GSTM1 基因多态性与重度慢性牙周炎易感性关系的研究

目的：研究GSTTI＋／0和GSTM1＋／0基因型及其联合基因型与重度慢性牙周（chronic pefiodontitis,cp）易感性的关系。方法：用聚合酶链反应检测50例重度慢性牙周炎患者和51例正常对照者的GSIT1＋／0、GSTM1＋／0的基因型。结果：GSTM1（0／0）和GSTT1（0／0）基因型及GSTMI（0／0）与GSTT1（0／0）联合基因型对重度慢性牙周炎相对危险度（OR）分别为9．56（95％CI．3．88—23．59）,8．68（95％CI,3．50—21．51）,36．83（95％CI,10．42—130．13）。结论：在内蒙古汉族人群中,基因型GSTT1（0／0）和GSTM1（0／0）增加了个体对重度慢性牙周炎易感性,且上述两种基因型间存在协同作用。     

C型产气荚膜梭菌α、β_1毒素基因的融合

利用PCR技术,从C型产气荚膜梭菌染色体DNA中扩增出α和β1毒素基因,通过分离、纯化、内切酶酶切、连接和转化,构建了含αβ1融合基因表达质粒重组菌株BL21(DE3)(pETXAB1)。经酶切鉴定和核苷酸序列测定证实,构建的重组质粒pETXAB1含有αβ1融合基因,且基因序列和阅读框架均正确。经ELISA检测,重组菌株表达的αβ1融合蛋白能够被α、β1毒素抗体识别。免疫实验结果表明,αβ1融合蛋白免疫的小鼠可以抵抗1MLD的C型产气荚膜梭菌C5944毒素攻击,表明构建的重组菌株可以作为预防仔猪红痢基因工程亚单位苗的候选菌株。