Virus-induced transformation without cell division

Interference with the cell cycle by vinblastine sulfate immediately after cells were infected with Rous sarcoma virus had little effect on the a development of two metabolic changes that occur in transformed cells. These results, along with an earlier demonstration of morphological changes developing in infected nondividing cells, demonstrate that the phenotypic development of the malignant state can occur without the intervention of cell divisions after infection by Rous sarcoma virus.     

周期操作对重组大肠杆菌高密度发酵表达类人胶原蛋白的影响

采用补料一分批式发酵,研究了4种不同间歇周期补氮操作对细胞生长和类人胶原蛋白表达的影响,以及发酵过程中细胞质粒稳定性、细胞菌落形成能力和乙酸生成量的变化。结果表明,在每隔10min连续补加 36mL补料氮溶液(22s完成补加操作)的周期操作下,细胞质粒稳定性明显提高,细胞菌落形成能力下降较小,乙酸生成量低,仅0．7 g／L,最终细胞密度为84 g／L,类人胶原蛋白表达量可达到14 g／L,在4种操作方式中最高。     

Restoration of gamma globulin production in agammaglobulinemic chickens

Chickens irradiated and bursectomized in the newly hatched period consistently develop agammaglobulinemia and form no circulating antibodies; if the birds are treated immediately after operations by intra-abdominal injection of unirradiated autologous bursa cells, immunoglobulin production, lymphoid germinal centers, and plasma cells are restored; however, the birds fail to produce antibody to specific antigenic challenge.     

Contribution of receptor editing to the antibody repertoire

Receptor editing, clonal deletion, and anergy are the mechanisms by which B cells maintain tolerance to self antigens. To determine the extent to which receptor editing shapes the normal antibody repertoire, we generated an immunoglobulin kappa polymorphism that facilitates the detection of editing of immunoglobulin light chains in vivo. We found that B cells are targeted for editing during a 2-hour delay in development at the pre-BII cell stage, and that about 25% of all antibody molecules are produced by gene replacement. These results suggest that receptor editing represents a major force in shaping the antibody repertoire.     

Restriction of DNA replication to the reductive phase of the metabolic cycle protects genome integrity

When prototrophic yeast cells are cultured under nutrient-limited conditions that mimic growth in the wild, rather than in the high-glucose solutions used in most laboratory studies, they exhibit a robustly periodic metabolic cycle. Over a cycle of 4 to 5 hours, yeast cells rhythmically alternate between glycolysis and respiration. The cell division cycle is tightly constrained to the reductive phase of this yeast metabolic cycle, with DNA replication taking place only during the glycolytic phase. We show that cell cycle mutants impeded in metabolic cycle-directed restriction of cell division exhibit substantial increases in spontaneous mutation rate. In addition, disruption of the gene encoding a DNA checkpoint kinase that couples the cell division cycle to the circadian cycle abolishes synchrony of the metabolic and cell cycles. Thus, circadian, metabolic, and cell division cycles may be coordinated similarly as an evolutionarily conserved means of preserving genome integrity.     

Quantitative aspects of plasma membrane-associated immunoglobulin in clones of diploid human lymphocytes

The amount of kappa and lambda chains and Fc fragment associated with the plasma membrane and in the entire cell has been measured for eight lines of human lymphocytes and 21 clones derived from two diploid lines. There was considerable variation in the nature and amount of membrane-associated immunoglobulin and total immunoglobulin among different cell lines and clones. Cells with different phenotypes for membrane-associated immunoglobulin were utilized to show that it is not simply absorbed from the culture medium onto the plasma membrane. The methods are useful for selecting variants which lack or have altered membrane-associated immunoglobulin.     

G1 events and regulation of cell proliferation

Cells prepare for S phase during the G1 phase of the cell cycle. Cell biological methods have provided knowledge of cycle kinetics and of substages of G1 that are determined by extracellular signals. Through the use of biochemical and molecular biological techniques to study effects of growth factors, oncogenes, and inhibitors, intracellular events during G1 that lead to DNA synthesis are rapidly being discovered. Many cells in vivo are in a quiescent state (G0), with unduplicated DNA. Cells can be activated to reenter the cycle during G1. Similarly, cells in culture can be shifted between G0 and G1. These switches in and out of G1 are the main determinants of post-embryonic cell proliferation rate and are defectively controlled in cancer cells.     

Protein-DNA interactions in vivo upstream of a cell cycle-regulated human H4 histone gene

Cell cycle-dependent histone genes are transcribed at a basal level throughout the cell cycle, with a three- to fivefold increase during early S phase. Protein-DNA interactions in the 5' promoter region of a cell cycle-regulated human H4 histone gene have been analyzed at single-nucleotide resolution in vivo. This region contains two sites, with four potential protein-binding domains, at which the DNA is protected from reaction with dimethyl sulfate in cells and from digestion with deoxyribonuclease I in nuclei. These protein-DNA interactions persist during all phases of the cell cycle and dissociate with 0.16 to 0.2M sodium chloride.     

AID-driven deletion causes immunoglobulin heavy chain locus suicide recombination in B cells

Remodeling of immunoglobulin genes by activation-induced deaminase (AID) is required for affinity maturation and class-switch recombination in mature B lymphocytes. In the immunoglobulin heavy chain locus, these processes are predominantly controlled by the 3' cis-regulatory region. We now show that this region is transcribed and undergoes AID-mediated mutation and recombination around phylogenetically conserved switchlike DNA repeats. Such recombination, which we term locus suicide recombination, deletes the whole constant region gene cluster and thus stops expression of the immunoglobulin of the B cell surface, which is critical for B cell survival. The frequency of this event is approaching that of class switching and makes it a potential regulator of B cell homeostasis.     

Control of the reversibility of cellular quiescence by the transcriptional repressor HES1

The mechanisms by which quiescent cells, including adult stem cells, preserve their ability to resume proliferation after weeks or even years of cell cycle arrest are not known. We report that reversibility is not a passive property of nondividing cells, because enforced cell cycle arrest for a period as brief as 4 days initiates spontaneous, premature, and irreversible senescence. Increased expression of the gene encoding the basic helix-loop-helix protein HES1 was required for quiescence to be reversible, because HES1 prevented both premature senescence and inappropriate differentiation in quiescent fibroblasts. In some human tumors, the HES1 pathway was activated, which allowed these cells to evade differentiation and irreversible cell cycle arrest. We conclude that HES1 safeguards against irreversible cell cycle exit both during normal cellular quiescence and pathologically in the setting of tumorigenesis.     

Diurnal expression of transducin mRNA and translocation of transducin in rods of rat retina

The messenger RNA (mRNA) that encodes alpha subunit of the guanosine triphosphate-binding protein transducin (T alpha) and T alpha immunoreactivity were localized and measured in the rat retina during the light-dark cycle with in situ hybridization and immunohistochemistry. Both T alpha mRNA and T alpha immunoreactivity were observed only in photoreceptors. Within the photoreceptor T alpha mRNA was present primarily in the inner segments and to a lesser extent in the outer nuclear layer at all times during the day and night. However, the distribution of T alpha immunoreactivity varied profoundly with the light-dark cycle; during the day, T alpha immunoreactivity was highest in the inner segments, and at night the outer segments were more immunoreactive. The amounts of T alpha mRNA and T alpha immunoreactivity also depended on the light-dark cycle. Levels of T alpha mRNA were high immediately before and after lights on; levels were low for the rest of the light-dark cycle. During the day, T alpha immunoreactivity increased in the inner segments following the increase in T alpha mRNA. After the lights were turned off, T alpha immunoreactivity decreased in the inner segments and increased in the outer segments. Thus, it appears that T alpha is synthesized in the inner segments after a morning increase in T alpha mRNA. Newly synthesized T alpha remains in the inner segments until it is transported to the outer segments at night, where it may be involved in the increase in the sensitivity of photoreceptor rods at night.     

Cell cycle dependence of chloride permeability in normal and cystic fibrosis lymphocytes

Cystic fibrosis (CF) is a genetic disease characterized by abnormal regulation of epithelial cell chloride channels. Nonepithelial cells, including lymphocytes and fibroblasts, may exhibit a similar defect. Two independent techniques were used to assess the macroscopic chloride permeability (PCl) of freshly isolated B lymphocytes and of B and T lymphocyte cell lines. Values for PCl increased specifically during the G1 phase of the cell cycle and could be further enhanced by increasing intracellular adenosine 3',5'-monophosphate (cAMP) or calcium. In lymphocytes from CF patients, regulation of PCl during the cell cycle and by second messengers was absent. Characterization of the cell cycle-dependent expression of the chloride permeability defect in lymphocytes from CF patients increases the utility of these cells in the analysis of the functional consequences of mutations in the CF gene.     

鱼类免疫球蛋白分子生物学研究进展

综述了近年来鱼类免疫球蛋白分子生物学的最新研究进展,主要涉及鱼类Ig基因序列分析、组成形式、重排机制以及转录调控4个方面.鱼类Ig重链和轻链基因克隆分析证实有多种不同的重链、轻链类型存在;重链和轻链基因由不同染色体上的多基因座编码,在不同的鱼类中具有不同的基因组织形式;重组信号序列在重链、轻链基因组中均有发现,其重排过程主要由重组活化酶进行调控;增强子和启动子在鱼类Ig转录调控中发挥重要作用,其中增强子序列在系统发育中具有保守性.Ig作为鱼体特异性体液免疫应答的主要因子,其分子生物学的深入研究对于阐明脊椎动物免疫系统进化具有重要意义.     

An origin unwinding activity regulates initiation of DNA replication during mammalian cell cycle

An in vitro assay was developed to study the positive factors that regulate the onset of DNA replication during the mammalian cell cycle. Extracts prepared from cells at defined positions in the cell cycle were used to examine the replication of SV40 DNA in a cell free system. Extracts prepared from S phase cells were ten times more efficient at initiating replication at the SV40 origin than were extracts from G1 cells, whereas elongation rates were similar in G1 and S reactions. At a discrete point in the cell cycle, just before the cell's entry into S, an activity appeared that was required, in conjunction with SV40 T antigen, for site specific initiation at the SV40 origin. This factor had a role in unwinding DNA at the replication origin.     

Methylation of an immediate-early inducible gene as a mechanism for B cell tolerance induction

Stage-specific gene regulation is important in determining cell function during development. Immature B cells expressing membrane-bound immunoglobulin M (mIgM) are sensitive to antigen-induced tolerance, whereas mature B cells are activated by antigen. Previous studies have established an association between Egr-1 gene induction and antigen receptor (mIgM)-mediated activation of mature B cells. Here it is shown that the immature B cell line WEHI-231 and tolerance-sensitive bone marrow-derived B cells do not express Egr-1. It is further shown that lack of inducible expression in these cells is due to specific methylation of the Egr-1 gene. Thus, covalent inactivation of an activation-associated gene may explain tolerance sensitivity at specific stages of B cell development.     

酵母基因表达调控关系的构建及其统计特性分析

[目的]从复杂网络的角度研究酵母细胞周期调控网络的统计特性。[方法]首先对酵母的细胞周期基因表达数据采用皮尔逊相关性度量,计算基因间的关联矩阵,即建立简单的基因调控网络;同时为了能够得到在宏观角度对大规模的基因调控有一个全貌的了解,进一步分析所建立调控网络的统计特性,通过计算所建立网络的统计特性：即计算网络的聚类系数、平均路径长度和网络的度分布。[结论]发现所建立的网络具有无标度和小世界特性,即所建立的酵母基因调控网络属于复杂网络,这为进一步深入研究基因调控网络的统计学和动力学特性打下了基础。     

Herpes simplex virus glycoprotein D: human monoclonal antibody produced by bone marrow cell line

Normal bone marrow cells from a donor positive for herpes simplex virus were transformed with Epstein-Barr virus. The resulting lymphoblastoid cell line has secreted immunoglobulin G1 of the kappa type continuously for 2 years. This immunoglobulin, detected both on the cell surface and in the cytoplasm, reacts with cells infected with herpes simplex virus. It defines an antigen that comigrates with the 55-kilodalton glycoprotein D of herpes simplex virus type 1 and neutralizes the infectivity of herpes simplex viruses 1 and 2.     

Effects of mosquito genes on Plasmodium development

Malaria parasites must complete a complex developmental cycle in an Anopheles mosquito vector before transmission to a vertebrate host. Sexual development of the parasite in the midgut is initiated in the lumen immediately after the mosquito ingests infected blood, and the resulting ookinetes must traverse the surrounding epithelial layer before transforming into oocysts. The innate immune system of the mosquito is activated during midgut invasion, but to date, no evidence has been published identifying mosquito immune genes that affect parasite development. Here, we show by gene silencing that an Anopheles gambiae leucine rich-repeat protein acts as an antagonist and two C-type lectines act as protective agonists on the development of Plasmodium ookinetes to oocysts.