Promoter of soybean early nodulin gene<Emphasis Type="Italic">enod2B</Emphasis> is induced by rhizobial Nod factors in transgenic rice

Nod factors, which are signaling molecules produced by Rhizobia, are the principal determinants of host specificity in Rhizobium-legume symbiosis. Nod factors can elicit a number of characteristic developmental responses in the roots of legumes, such as depolarization of the membrane potential in epidermal cells, specific expression of early nodulin genes and changes in the flux of calcium in root hairs, deformation of root hairs, cell division in the root cortex and formation of the nodule primordinm. Whether the rice plant can respond to signaling molecules (i.e. Nod factors) is an important question, as it could establish the potential for symbiotic nitrogen fixation in rice. The promoter of the soybean (Glycine max) early nodulin gene Gmenod2B fused to the β-glucuronidase (GUS) reporter gene was used as a molecular marker to explore whether Nod factors can be recognized by rice cells as signaling molecules. Transgenic rice plants harboring the chimeric gene Gmenod2BP-GUS were obtained via an Agrobacterium tumefaciens-mediated system. NodNGR factors produced by a broad-host-range Rhizobium strain NGR234(pA28) were used as probes to investigate the activity of the Gmenod2B promoter in rice. Our results showed that the early nodulin gene Gmenod2B promoter was induced by NodNGR factors in transgenic rice, and that it was specifically expressed in rice plant roots. Moreover, GUS gene expression driven by the Gmenod2B promoter in transgenic rice was regulated by nitrogen status. These findings indicated that rice possessed the ability to respond to Nod factor signals, and that this signal transduction system resulted in activation of the Gmenod2B promoter. Thus, we predict that the Nod-factor inducible nodulin expression system, which is similar to Rhizobium-legume symbiosis, may also exist in rice.     

Expression of the E. coli uvrA gene is inducible

UvrA+-dependent excision repair is one of the most important systems in Escherichia coli for repairing UV-induced pyrimidine dimers and a variety of other forms of DNA damage. The uvrA protein acts in conjunction with the uvrB and uvrC gene products to introduce a nick at the of a DNA lesion and thus initiate the repair process. We have recently used the Mud(Ap, lac) operon fusion vector to identify a set of genes whose expression is induced by DNA damage. One Mud(Ap, lac) insertion mapped at the uvrA locus and made the cells sensitive to UV light. In this fusion strain, beta-galactosidase expression was induced by DNA-damaging agents in a recA+lexA+-dependent fashion. We were surprised by this result because uvrA+-dependent excision repair is observed both in cells in which protein synthesis has been inhibited and in recA- and lexA- cells, findings which have led to the conclusion that the uvrA gene product is constitutively expressed and not under the control of the complex recA+lexA+ regulatory circuitry (see below). We have investigated this possibility further and describe here the generation and characterization of a set of fusions of the lac genes to the promoter of the uvrA gene. We confirm that the uvrA gene product is induced by DNA damage in a recA+lexA+-dependent fashion.     

Demonstration by genetic suppression of interaction of GroE products with many proteins

The way in which proteins attain and maintain their final form is of fundamental importance. Recent work has focused on the role of a set of ubiquitous proteins, termed chaperonins, in the assembly of phage and multisubunit proteins. The range of chaperonin action is unknown; they could interact with most cellular polypeptides or have a limited subset of protein partners. Included in the chaperonin family is the essential heat-shock regulated Escherichia coli groEL gene product. Over-expression of the groE operon in E. coli causes enhanced assembly of heterologously expressed ribulose bisphosphate carboxylase subunits and suppresses the heat-sensitive mutant phenotype of several dnaA alleles. It has been inferred that suppression of heat-sensitive mutations is confined to dnaA alleles and that this confinement could reflect an interaction between the groE operon products and a dnaA protein aggregate at the replication origin. We now report that multiple copies of the groE operon suppress mutations in genes encoding several diverse proteins. Our data indicate a general role for the groE operon products, the GroEL and GroES proteins, in the folding-assembly pathways of many proteins.     

The induction of Sinorhizobium meliloti C4-dicarboxylate transport system （Dct） is regulated by oxygen concentration

The Sinorhizobium meliloti C4-dicarboxylate transport （Dct） system is essential for symbiotic nitrogen fixation. The dctA gene, encoding the C4-dicarboxylate permease, is expressed in both free living and symbiotic cells. But in free living cells expression of dctD and dctB is absolutely required for the expression of dctA. In this study, in order to investigate the effect of oxygen concentration on the induction of Dct system, E. coli DH5a strain which carries the plasmid-encoded dctABD operon was used in tube assays. It was found that the specific induction of Dct system oc- curred only at a certain depth under the surface of M63-0.6% agar media, suggesting that Dct system could respond to oxygen concentration during succinate-induced expression. Furthermore, when measured at different oxygen concentrations, the highest expression level was observed at oxygen concentration of 2%. Thus, we predict that in addition to dicarboxylates, the induction of Dct system may also regulated by oxygen concentration.     

Modification of the rib operon derived from Bacillus subtilis and its expression in Escherichia coli

A riboflavin operon （rib operon） derived from Bacillus subtilis 368 was modified on structure and the resulting operons were expressed in various strains of Escherichia coli. The results showed that the optimization of the rib operon and the host strain used for expression are two main factors affecting the riboflavin production. Replacing the promoterl and rfn box of the rib operon with a strong constructive promoter spol drastically increased the expression of the rib genes. When E. coli JM109 was used as the host strain, the highest riboflavin production reached 95.3μg/mL （about eight times higher than that of the unmodified r/b operon）. In addition, when tetracycline （20 μg/mL） was used as the selective pressure, compared with the ampicillin resistant transformants, a higher riboflavin yield was obtained in tetracycline resistant host strain.     

Transfer of a cloned immunoglobulin light-chain gene to mutant hybridoma cells restores specific antibody production

The expression of immunoglobulin (Ig) genes is regulated at several levels. For example, although kappa-chain production requires a DNA rearrangement that juxtaposes variable and joining segments, this rearrangement is not sufficient for kappa-chain gene expression; that is, some cell types do not permit immunoglobulin production. The mechanisms responsible for the regulation of the expression of rearranged immunoglobulin genes are poorly understood. The technique of modifying cloned genes in vitro and transferring the modified genes to cells in culture provides a tool for identifying the structural features required for gene expression. To analyse immunoglobulin genes in this manner, however, it is first necessary to use, as recipients, cells that normally permit immunoglobulin production. We report here that a cloned kappa-chain gene is expressed in immunoglobulin-producing hybridoma cells. Furthermore, the product of the transferred kappa-chain gene is capable of restoring specific antibody production to the transformed cells.     

Immunodeficiency virus rev trans-activator modulates the expression of the viral regulatory genes

The pathogenic human retrovirus human immunodeficiency virus type 1 (HIV-1) encodes two trans-acting nuclear proteins, tat and rev, whose functional expression is essential for viral replication in vitro. The tat protein greatly enhances the expression of both structural and regulatory genes of HIV-1 (linked to the viral long-terminal-repeat promoter element), whereas the rev gene product (previously termed art or trs) has only been shown to be required for the synthesis of structural proteins. Here, we demonstrate that rev also moderates the expression of regulatory genes of HIV-1. It decreases the expression of messenger RNAs that encode the full-length form of the viral tat gene product or the rev protein itself, and induces the synthesis of a previously unreported, truncated tat protein. These actions of rev are mediated by a dramatic shift in the ratio of spliced to unspliced cytoplasmic HIV-1 mRNA. Therefore rev not only activates the synthesis of the viral structural proteins, but also modulates the level and quality of HIV-1 regulatory gene expression.     

The locus of sequence-directed and protein-induced DNA bending

The bending locus of trypanosome kinetoplast DNA, identified by gel electrophoresis, has tracts of a simple repeat sequence (CA5-6 T) symmetrically distributed about it, with a repeat interval of 10 base pairs. The analogous bending induced when catabolite gene activating protein binds to its recognition sequence near the promoter of the Escherichia coli lac operon is centred on a site about 5-7 base pairs away from the centre of the protein binding site.