Expression Of Nodule-specific Genes Research Articles

Functional analysis of the Sesbania rostrata leghemoglobin glb3 gene 5'-upstream region in transgenic Lotus corniculatus and Nicotiana tabacum plants.

Expression of the Sesbania rostrata leghemoglobin glb3 gene was analyzed in transgenic Lotus corniculatus and tobacco plants harboring chimeric glb3-uidA (gus) gene fusions to identify cis-acting elements involved in nodule-specific gene expression and general transcriptional control. A 1.9-kilobase fragment of the glb3 5'-upstream region was found to direct a high level of nodule-specific beta-glucuronidase (GUS) activity in L. corniculatus, restricted to the Rhizobium-infected cells of the nodules. The same fragment directed a low level of GUS activity in tobacco, restricted primarily to the roots and to phloem cells of the stem and petiole vascular system. A deletion analysis revealed that the region between coordinates -429 and -48 relative to the ATG was sufficient for nodule-specific expression. Replacement of the -161 to -48 region, containing the glb3 CAAT and TATA boxes, with the heterologous truncated promoters delta-p35S and delta-pnos resulted in a loss of nodule specificity and reduction of GUS activity in L. corniculatus but a significant increase in tobacco, primarily in the roots. The same fragment could not direct nodule-specific expression when fused to a heterologous enhancer in cis. This region contains DNA sequences required, but not sufficient, for nodule-specific expression in L. corniculatus that function poorly or may be involved in promoter silencing in tobacco. By fusing further upstream fragments to the delta-p35S and delta-pnos promoters, two positive regulatory regions were delimited between coordinates -1601 and -670, as well as -429 and -162. The former region appears to function as a general enhancer because it significantly increased promoter activity in both orientations in L. corniculatus and tobacco. The latter region could enhance gene expression in both orientations in tobacco, but only in the correct orientation in L. corniculatus. These results show that efficient expression of the S. rostrata glb3 gene in nodules is mediated by an ATG-proximal, tissue-specific element, as well as further 5'-upstream positive elements; that the S. rostrata glb3 promoter is induced in a nodule-specific fashion in the heterologous legume L. corniculatus, suggesting a high degree of conservation of the relevant regulatory signals; and that the S. rostrata lb promoter is not silent in the nonlegume tobacco, but is expressed primarily in the roots.

Chimeric genes and transgenic plants are used to study the regulation of genes involved in symbiotic plant‐microbe interactions (nodulin genes)

Nodulin genes are plant genes specifically activated during the formation of nitrogen-fixing nodules on leguminous plants. These genes are interesting to study since they are not only induced in a specific developmental fashion by signals coming directly or indirectly from the rhizobial symbiont, but are also expressed in a tissue-specific manner. By examining the expression of chimeric nodulin-reporter genes in transgenic legume plants it has been shown that nodule specific expression is mediated by DNA sequences present in the 5 upstream region of several nodulin genes. Here we summarize the available data on these cis-acting elements and the trans-acting factors interacting with them. We also review experiments designed to identify rhizobial "signals" which may play a role in nodule specific gene expression.

Expression of nodule-specific genes in alfalfa root nodules blocked at an early stage of development.

To help dissect the molecular basis of the Rhizobium-legume symbiosis, we used in vitro translation and Northern blot analysis of nodule RNA to examine alfalfa-specific genes (nodulins) expressed in two types of developmentally defective root nodules elicited by Rhizobium meliloti. Fix- nodules were elicited by R. meliloti nif mutants; these nodules were invaded by rhizobia and contained differentiated bacteroids. 'Empty' nodules were elicited by R. meliloti exo and ndv mutants and by Agrobacterium tumefaciens strains carrying the R. meliloti nod genes; these nodules contained a nodule meristem but lacked infection threads, intracellular bacteria, and bacteroids. Fix- nodules contained a spectrum of nodulins similar to wild-type nodules. In contrast, only two nodulins, Nms-30 and a nodulin homologous to ENOD2 of soybean, were detected in empty nodules. Although R. meliloti ndv and exo mutants elicited nodules with the same defective phenotype, ndv and exo mutants (except for exoC mutants) had distinct biochemical phenotypes. R. meliloti ndvA and ndvB mutants were deficient in cyclic glucan production but not the acidic exopolysaccharide; the converse was true for exoA, exoB, and exoF mutants. exoC mutants were defective in both exopolysaccharide and cyclic glucan biosynthesis. Our results support the model that the R. meliloti nod genes produce a signal that results in nodule meristem induction. Both the exopolysaccharide and cyclic glucan, however, appear to act at the next step in the developmental process and are involved in the production of a signal (or structure) that allows infection thread formation and invasion of the nodule.