A novel translation initiation region from Mycoplasma genitalium that functions in Escherichia coli.

The tuf gene of Mycoplasma genitalium uses a signal other than a Shine-Dalgarno sequence to promote translation initiation. We have inserted the translation initiation region of this gene in front of the Escherichia coli lacZ gene and shown that it is recognized by the translational machinery of E. coli; the signal operates in vivo at roughly the same efficiency as a synthetic Shine-Dalgarno sequence. The M. genitalium sequence was also used to replace the native translation initiation region of the cat gene. When assayed in E. coli, the M. genitalium sequence is equivalent to a Shine-Dalgarno sequence in stimulating translation of this mRNA also. Site-directed mutagenesis enabled us to identify some of the bases that comprise the functional sequence. We propose that the sequence UUAACAACAU functions as a ribosome binding site by annealing to nucleotides 1082-1093 of the E. coli 16S rRNA. The activity of this sequence is enhanced when it is present in the loop of a stem-and-loop structure. Additional sequences both upstream and downstream of the initiation codon are also involved, but their role has not been elucidated.     

Regulation of differentiation of the BC3H1 muscle cell line through cAMP-dependent and -independent pathways

Serum mitogens, fibroblast growth factor (FGF), and type beta transforming growth factor (TGF-beta) suppress differentiation of the mouse muscle cell line BC3H1; however, the signal transduction pathways whereby these growth factors exert their effects on this system are unknown. The goal of this study was to determine whether the program for differentiation of BC3H1 cells was susceptible to negative regulation by signaling pathways involving cAMP or protein kinase C and whether these intracellular effectors participate in the mechanism by which growth factors prevent establishment of the myogenic phenotype. Exposure of BC3H1 cells to dibutyryl cAMP, 8-bromo-cAMP, or compounds that stimulate adenylate cyclase, i.e. forskolin, prostaglandin E1, and cholera toxin, prevented up-regulation of muscle-specific gene products following growth arrest in mitogen-deficient medium. Conversely, addition of cAMP to differentiated BC3H1 myocytes caused down-regulation of muscle-specific mRNAs. In contrast to the ability of cAMP to block differentiation, chronic exposure to O-tetradecanoylphorbol-13-acetate, the potent activator of protein kinase C, exhibited no apparent effects on expression of muscle-specific gene products. The proto-oncogenes c-myc and c-fos were up-regulated rapidly by cAMP in a manner similar to that observed previously by serum, FGF, and TGF-beta. However, these growth factors failed to increase intracellular cAMP levels, and they did not induce ornithine decarboxylase, which was subject to positive regulation by cAMP and O-tetradecanoyl-13-acetate. Together, these data indicate that differentiation of BC3H1 cells is subject to negative regulation through a cAMP-dependent pathway and that serum mitogens, FGF, and TGF-beta inhibit differentiation through a mechanism independent of cAMP or protein kinase C.     

Differing activities of medullary respiratory neurons in eupnea and gasping

Our purpose was to compare further eupneic ventilatory activity with that of gasping. Decerebrate, paralyzed, and ventilated cats were used; the vagi were sectioned within the thorax caudal to the laryngeal branches. Activities of the phrenic nerve and medullary respiratory neurons were recorded. Antidromic invasion was used to define bulbospinal, laryngeal, or not antidromically activated units. The ventilatory pattern was reversibly altered to gasping by exposure to 1% carbon monoxide in air. In eupnea, activities of inspiratory neurons commenced at various times during inspiration, and for most the discharge frequency gradually increased. In gasping, the peak discharge frequency of inspiratory neurons was unaltered. However, all commenced activities at the start of the phrenic burst and reached peak discharge almost immediately. The discharge frequencies of all groups of expiratory neurons fell in gasping, with many neurons ceasing activity entirely. These data are consistent with the hypothesis that brain stem mechanisms controlling eupnea and gasping differ fundamentally.     

Insulin suppresses hepatitis B surface antigen expression in human hepatoma cells

The human hepatoma Hep3B cells contain integrated hepatitis B viral genome and continually secret hepatitis B surface antigen (HBsAg). The production of HBsAg (but not alpha-fetoprotein) was suppressed by addition of low concentrations (0.1-1 nM) of insulin into serum-free medium. In addition, the suppression of HBsAg production by insulin was paralleled with the decrease in HBsAg mRNA abundance. Insulin also cause a rapid rate of disappearance of HBsAg mRNA (t 1/2, 2 h) in Hep3B cells. The Hep3B cells carry specific receptor with high affinity for insulin (Kd = 1.8 nM). The receptor showed an insulin-dependent protein tyrosine kinase activity. The half-maximal insulin concentration for the activation of the receptor kinase was about 5 nM. Only very high concentrations of insulin-like growth factor I and human proinsulin can compete for the insulin receptor binding and suppress HBsAg production, this suggests that insulin may act through its receptor binding to suppress HBsAg expression in human hepatoma Hep3B cells.     

Loss of Rb activates both p53-dependent and independent cell death pathways in the developing mouse nervous system.

Extensive apoptosis occurs in the nervous system of mouse embryos homozygous mutant for a targeted disruption of the retinoblastoma (Rb) gene. This cell death is present in both the central (CNS) and peripheral nervous systems (PNS) and is associated with abnormal S phase entry of normally post-mitotic neurons. Aberrant proliferation in the CNS correlates with increased free E2F DNA binding activity and increased expression of cyclin E, an E2F target gene and critical cell cycle regulator. Cell death in the CNS is accompanied by increased levels of the p53 tumor suppressor gene product and increased expression of the p53 target gene, p21Waf-1/Cip-1. However, induction of p53 is not observed in the PNS of Rb-mutant embryos, nor does loss of p53 function inhibit cell death in the PNS. Surprisingly, p21Waf-1/Cip-1 is induced in the sensory ganglia of Rb-mutant embryos in a p53-independent manner. Although loss of p53 gene function prevents cell death in the CNS of Rb-mutant embryos, it does not restore normal proliferative control.     

马桑菌株的生物学特性

对从湖北、四川、云南等地采集的尼泊尔马桑(Coriaria nepalensis Wall.)根瘤中分离的20株内生菌纯培养物所进行的研究表明,其形态均具有弗兰克氏菌属的特征。在粗细不一的分枝菌丝体上有泡囊和孢囊,有的还有串珠状结构的菌丝体。不同菌株在液体培养基上生长速度很不一致;菌丝体多呈絮状或颗粒状沉淀,个别还呈薄膜状沉淀;绝大多数菌株的菌丝体具有不同色调的色素;在S培养基中大多数都能产生可溶性色素。上述这些培养特征常因培养基成分和培养方式的不同而有差异。大多数菌株的胞壁组分属Ⅱ型,少数菌株为Ⅲ型;全细胞糖型变化很大,20个菌株可划分为6个糖型,而且绝大多数糖型与已知弗兰克氏菌有所不同。此外,有65％的菌株还具有明显的抗菌活性。     

Overproduction of Three Genes Leads to Camphor Resistance and Chromosome Condensation in Escherichia Coli

We isolated and characterized three genes, crcA, cspE and crcB, which when present in high copy confer camphor resistance on a cell and suppress mutations in the chromosomal partition gene mukB. Both phenotypes require the same genes. Unlike chromosomal camphor resistant mutants, high copy number crcA, cspE and crcB do not result in an increase in the ploidy of the cells. The cspE gene has been previously identified as a cold shock-like protein with homologues in all organisms tested. We also demonstrate that camphor causes the nucleoids to decondense in vivo and when the three genes are present in high copy, the chromosomes do not decondense. Our results implicate camphor and mukB mutations as interfering with chromosome condensation and high copy crcA, cspE and crcB as promoting or protecting chromosome folding.