Pathways of protein secretion in eukaryotes

Protein secretion from cells can take several forms. Secretion is constitutive if proteins are secreted as fast as they are synthesized. In regulated secretion newly synthesized proteins destined for secretion are stored at high concentration in secretory vesicles until the cell receives an appropriate stimulus. When both constitutive and regulated protein secretion can take place in the same cell a mechanism must exist for sorting the correct secretory protein into the correct secretory vesicle. The secretory vesicle must then be delivered to the appropriate region of plasma membrane. Transfection of DNA encoding foreign secretory proteins into regulated secretory cells has provided insight into the specificity of sorting into secretory vesicles.     

The molecular control of blood cell development

The establishment of a cell culture system for the clonal development of blood cells has made it possible to identify the proteins that regulate the growth and differentiation of different blood cell lineages and to discover the molecular basis of normal and abnormal cell development in blood forming tissues. A model system with myeloid blood cells has shown that (i) normal blood cells require different proteins to induce cell multiplication (growth inducers) and cell differentiation (differentiation inducers), (ii) there is a hierarchy of growth inducers as cells become more restricted in their developmental program, and (iii) a cascade of interactions between proteins determines the correct balance between immature and mature cells in normal blood cell development. Gene cloning has shown that there is a family of different genes for these proteins. Normal protein regulators of blood cell development can control the abnormal growth of certain types of leukemic cells and suppress malignancy by inducing differentiation to mature nondividing cells. Chromosome abnormalities that give rise to malignancy in these leukemic cells can be bypassed and their effects nullified by inducing differentiation, which stops cells from multiplying. These blood cell regulatory proteins are active in culture and in the body, and they can be used clinically to correct defects in blood cell development.     

紫苏腺毛的形态结构和发育的研究

 应用光学显微镜和扫描电子显微镜对紫苏腺毛的形态结构和发育进行了研究。紫苏腺毛起源于原表皮细胞,由1个基细胞、1个柄细胞和由2个或8个分泌细胞构成的头部3部份组成。根据头部分泌细胞的数目及其大小,紫苏腺毛可分成两类:大腺毛具有8个分泌细胞的头部,分布于叶脉之间;小腺毛头部有2个分泌细胞,位于叶脉之上。     

A microdomain for protein secretion in Gram-positive bacteria

Gram-positive bacteria face unique challenges in generating biologically active conformations for their exported proteins because they lack a dedicated compartment for folding secreted polypeptides. We have discovered that protein secretion by way of the general secretory (Sec) pathway in the important human pathogen Streptococcus pyogenes proceeds through a single microdomain. Unlike other mechanisms for asymmetry involving the Sec pathway, proteins destined for secretion are targeted to a single locus distal to either cell pole that has specialized to contain the Sec translocons. This subcellular organization may represent a paradigm for secretion common to other Gram-positive pathogens with profound implications for pathogenesis.     

Transition metal speciation in the cell: insights from the chemistry of metal ion receptors

The essential transition metal ions are avidly accumulated by cells, yet they have two faces: They are put to use as required cofactors, but they also can catalyze cytotoxic reactions. Several families of proteins are emerging that control the activity of intracellular metal ions and help confine them to vital roles. These include integral transmembrane transporters, metalloregulatory sensors, and diffusible cytoplasmic metallochaperone proteins that protect and guide metal ions to targets. It is becoming clear that many of these proteins use atypical coordination chemistry to accomplish their unique goals. The different coordination numbers, types of coordinating residues, and solvent accessibilities of these sites are providing insight into the inorganic chemistry of the cytoplasm.     

Unveiling the mechanisms of cell-cell fusion

Cell-cell fusion is fundamental to the development and physiology of multicellular organisms, but little is known of its mechanistic underpinnings. Recent studies have revealed that many proteins involved in cell-cell fusion are also required for seemingly unrelated cellular processes such as phagocytosis, cell migration, axon growth, and synaptogenesis. We review advances in understanding cell-cell fusion by contrasting it with virus-cell and intracellular vesicle fusion. We also consider how proteins involved in general aspects of membrane dynamics have been co-opted to control fusion of diverse cell types by coupling with specialized proteins involved in cell-cell recognition, adhesion, and signaling.     

四季鹅甲状旁腺在产蛋期与停产期的显微和超微结构的比较研究

鹅甲状旁腺实质仅主细胞构成,没有嗜酸性细胞,Ｈ－Ｅ染色。产蛋鹅主细胞较大,胞质深染,胸核大而圆,核仁明显,停产 细胞质淡染,较清亮。电镜下产蛋期主细胞电子密度中等,胞人含有丰富的细胞器;线粒体多,常膨大成圆形;高尔基复合体发达;南网稍扩张;脂质滴与分泌前颗粒均较多,成熟的分泌颗粒大而少,细胞这样期主细胞的细胞较和,核常染色质少,其相对稳定。     

Cationic protein-bearing granules of polymorphonuclear leukocytes: separation from enzyme-rich granules

The cationic, antibacterial proteins of polymorphonuclear leukocytes are associated with a unique subcellular particle that is separable through zonal density gradient centrifugation from acid phosphatase-containing particles as well as from particles that contain alkaline phosphatase and lysozyme. Normal macrophages, macrophages stimulated by bacillus Calmette-Guérin, and liver cells lack this particle and the associated group of cationic proteins. Particles physically and biochemically similar to slower sedimenting enzyme-rich particles of polymorphonuclear leukocytes are shared by all the tlhree cell types.     

长双歧杆菌NCC2705分泌蛋白的全基因组预测和功能分析

以长双歧杆菌NCC2705基因组序列为研究对象,使用Signal P 4.0、Lipo P、TMHMM 2.0软件分析该基因组中的分泌蛋白及其信号肽的类型,同时采用COG(Cluster of Orthologous Groups of proteins)功能数据库对预测的分泌蛋白进行功能注释和聚类分析。结果表明,长双岐杆菌NCC2705中共有37个Sec途径分泌蛋白,其中Sec途径分泌蛋白包括27个被I型(SPaseⅠ)信号肽酶和10个Ⅱ型信号肽酶(Spase II)识别的蛋白,Sec途径分泌蛋白信号肽长度最多的有52个氨基酸,最少的有10个氨基酸。分泌蛋白的功能分析表明,该菌株分泌蛋白中含有大部分的假定蛋白,主要参与氨基酸代谢与转运、碳水化合物代谢转运,无机盐离子代谢转运,细胞壁和细胞膜生物合成的功能有关。     

Molecular aspects of lens cell differentiation

I have presented a series of observations on macromolecular interactions which occur during the terminal stages of lens cell differentiation. These are summarized in Fig. 2. Other cell types that undergo similar changes are the erythrocyte and skin cells (epidermis) during the process of keratinization. These other cells are also involved in the synthesis of highly specific proteins, and there are indications that molecular alterations similar to those described for the lens may also occur in these cells (26). Thus, elucidation of a specific series of macromolecular initeractions such as those described may provide a basis for the biochemical definition of the terminal stages of cellular differentiation. Differentiation of the reticulocyte, for example, involves inactivation of the nucleus, stabilization of mRNA, and possibly a ribosomal breakdown such as I have described here (26). Furthermore, elucidation of the mechanisms of reactions involving the initiation of tissue-specific protein synthesis and suLbsequent nuclear inactivation, stabilization of mRNA, and breakdown of the ribosomes may provide a basis for defining the mechanisms of terminal cellular differentiation. The lens cell has reached its highest form of cellular differentiation when it has formed the fiber cell. With respect to the mechanism of lens fiber cell formation, we would like to know whether specific biochemical changes such as gamma-crystallin synthesis are intiniately linked to fiber cell formation-that is, whether gamma-crystallins are required to bring about the formation of a fiber cell. The potential for synthesizing gamma-crystallins is inherent in the genome of the cell. This part of the genome is nonfunctional in the epithelial cell. Can these genes be activated without bringing about a simultaneous cellular elongation, nuclear inactivation and loss of cellular replication, stabilization of mRNA, and breakdown of the ribosomes? The degree of coupling or uncoupling of tissue-specific-protein synthesis to morphogenesis is an important part of the mechanism of cellular differentiation. We feel that we have now reached the stage where we can begin to answer these questions.     

Diversity of G proteins in signal transduction

The heterotrimeric guanine nucleotide-binding proteins (G proteins) act as switches that regulate information processing circuits connecting cell surface receptors to a variety of effectors. The G proteins are present in all eukaryotic cells, and they control metabolic, humoral, neural, and developmental functions. More than a hundred different kinds of receptors and many different effectors have been described. The G proteins that coordinate receptor-effector activity are derived from a large gene family. At present, the family is known to contain at least sixteen different genes that encode the alpha subunit of the heterotrimer, four that encode beta subunits, and multiple genes encoding gamma subunits. Specific transient interactions between these components generate the pathways that modulate cellular responses to complex chemical signals.     

植物表达重组蛋白的N-糖基化研究进展

分子农业中,植物作为生物反应器生产药用重组蛋白是近十几年来基因工程研究的一大热点。植物具有完整的真核表达系统、重组蛋白通常可以正确组装、折叠和修饰,同时,由于植物表达体系的方便、廉价等优点,利用转基因植物生产药用蛋白的研究迅猛发展,已有100余种蛋白中得到表达,有些已进入商业化生产。但是植物与动物对蛋白的转译后修饰并不是完全保守的,植物是否能够真正“忠实”地表达出实用、可靠的目的蛋白产品已成为备受瞩目的话题,由于大多数药用蛋白都是分泌型蛋白,因此植物表达重组蛋白的N-糖基化成为新的研究热点。笔者对植物表达体系对药用蛋白的N-糖基化修饰途径和人源化改造植物表达重组蛋白的研究进展两个方面加以综述,并对生物反应器的未来发展方向进行了探讨和展望。     

Thymic requirement for clonal deletion during T cell development

During T cell differentiation, self tolerance is established in part by the deletion of self-reactive T cells within the thymus (negative selection). The presence of T cell receptor (TCR)-alpha beta + T cells in older athymic (nu/nu) mice indicates that some T cells can also mature without thymic influence. Therefore, to determine whether the thymus is required for negative selection, TCR V beta expression was compared in athymic nu/nu mice and their congenic normal littermates. T cells expressing V beta 3 proteins are specific for minor lymphocyte stimulatory (Mlsc) determinants and are deleted intrathymically due to self tolerance in Mlsc+ mouse strains. Here it is shown that V beta 3+ T cells are deleted in Mlsc+ BALB/c nu/+ mice, but not in their BALB/c nu/nu littermates. Thus, the thymus is required for clonal deletion during T cell development.     

Molecular sorting in the secretory pathway

Proteins can be secreted from animal cells by either a constitutive or a regulated pathway; those destined for regulated secretion are actively sorted into dense-core secretory granules. Although sorting is generally assumed to be accomplished by specific carriers, the nature of these carriers remains elusive. In this study, peptide hormones were used as affinity ligands to purify a set of 25-kilodalton proteins from canine pancreatic tissue. Their ligand specificities and patterns of expression have the characteristics of sorting carriers.     

Regulatory role for GTP-binding proteins in endocytosis

Guanosine 5'-triphosphate (GTP)-binding proteins have been implicated in the transport of newly synthesized proteins along the secretory pathway of yeast and mammalian cells. Early vesicle fusion events that follow receptor-mediated endocytosis as measured by three in vitro assays were blocked by guanosine 5'-O-(3-thiotriphosphate) and aluminum fluoride. The effect was specific for guanosine nucleotides and depended on the presence of cytosolic factors. Thus, GTP-binding proteins may also have a role in the transport of molecules along the endocytic pathway.     

Bioinformatic analysis and functional characterization of the cfem proteins in maize anthracnose fungus Colletotrichum graminicola

Fungal secreted proteins that contain the Common in Fungal Extracellular Membrane(CFEM) domain are important for pathogenicity. The hemibiotrophic fungus Colletotrichum graminicola causes the serious anthracnose disease of maize. In this study, we identified 24 CgCFEM proteins in the genome of C. graminicola. Phylogenic analysis revealed that these 24 proteins(CgCFEM1–24) can be divided into 2 clades based on the presence of the trans-membrane domain. Sequence alignment analysis indicated that the amino acids of the CFEM domain are highly conserved and contain 8 spaced cysteines, with the exception that CgCFEM1 and CgCFEM24 lack 1 and 2 cysteines, respectively. Ten CgCFEM proteins with a signal peptide and without the trans-membrane domain were considered as candidate effectors and, thus were selected for structural prediction and functional analyses. The CFEM domain in the candidate effectors can form a helical-basket structure homologous to the Csa2 protein in Candida albicans, which is responsible for haem acquisition and pathogenicity. Subcellular localization analysis revealed that these effectors accumulate in the cell membrane, nucleus, and cytosolic bodies. Additionally, 5 effectors, CgCFEM6, 7, 8, 9 and 15, can suppress the BAX-induced programmed cell death in Nicotiana benthamiana with or without the signal peptide. These results demonstrate that these 10 CgCFEM candidate effectors with different structures and subcellular localizations in host cells may play important roles during the pathogenic processes on maize plants.     

Structural basis for tail-anchored membrane protein biogenesis by the Get3-receptor complex

Tail-anchored (TA) proteins are involved in cellular processes including trafficking, degradation, and apoptosis. They contain a C-terminal membrane anchor and are posttranslationally delivered to the endoplasmic reticulum (ER) membrane by the Get3 adenosine triphosphatase interacting with the hetero-oligomeric Get1/2 receptor. We have determined crystal structures of Get3 in complex with the cytosolic domains of Get1 and Get2 in different functional states at 3.0, 3.2, and 4.6 angstrom resolution. The structural data, together with biochemical experiments, show that Get1 and Get2 use adjacent, partially overlapping binding sites and that both can bind simultaneously to Get3. Docking to the Get1/2 complex allows for conformational changes in Get3 that are required for TA protein insertion. These data suggest a molecular mechanism for nucleotide-regulated delivery of TA proteins.