Characterization of cDNA for nodulin-75 of soybean: A gene product involved in early stages of root nodule development

Abstract

Establishment of a nitrogen-fixing root nodule is accompanied by a developmentally regulated expression of nodulin genes, only some of which, the so-called early nodulin genes, are expressed in stages preceding actual nitrogen fixation. We have isolated soybean cDNA clones representing early nodulin genes and have studied clone pENOD2 in detail. The cDNA insert of this clone hybridizes to nodulespecific RNA of 1200 nucleotides in length. The RNA that was hybrid-selected by the cloned ENOD2 DNA was in vitro translated to produce two nodulins with an apparent Mr of 75,000, the N-75 nodulins. These two nodulins differ slightly in charge and one does not contain methionine. The amino acid sequence deduced from the DNA sequence shows that proline accounts for 45% of the 240 residues in these nodulins and the sequence contains at least 20 repeating heptapeptide units. The amino acid composition of none of the (hydroxy)proline-rich (glyco)proteins described in plants resembles the composition of the N-75 nodulins, especially with respect to the high glutamic acid and the low serine content. This suggests that the N-75 nodulins belong to a hitherto unidentified class of presumably structural proteins. The genes encoding the N-75 nodulins were found to be expressed in nodule-like structures devoid of intracellular bacteria and infection threads, indicating that these nodulins do not function in the infection process but more likely function in nodule morphogenesis.The formation of nitrogen-fixing nodules on the roots of leguminous plants induced by bacteria of the genera Rhizobium and Bradyrhizobium involves the specific expression of a number of plant genes called nodulin genes (1-3). In a description of nodule development, Vincent (4) distinguishes between three stages in nodule development denoted as "preinfection", "infection and nodule formation", and "nodule function". In the preinfection stage, the Rhizobium bacteria recognize their host plants and attach to the root hairs, an event that is followed by root hair curling. At the moment, nothing is known about specific plant genes that are involved in this stage. In the next stage, the bacteria enter the roots by infection threads while concomitantly the dedifferentiation of some cortical cells results in the formation of meristems. The infection threads grow toward the meristematic cells; bacteria are released into the cytoplasm of about half of these cells and develop into bacteroids. In the final stage, further differentiation of nodule cells occurs leading up to a nitrogen-fixing nodule. Most studies on the expression of nodulin genes so far have been confined to the final stage of root nodule development. But the steps involved in root nodule formation show that major decisions determining the development of a root nodule are made in the stages preceding the establishment of a nitrogen-fixing nodule. We have shown (5) that nodulin genes are differentially expressed during development and that in pea at least two nodulin genes are transcribed in the seco...

Pipeline to Identify Hydroxyproline-Rich Glycoproteins

“…4), and SbPRP3 (Glyma11g21616.1; Hong et al, 1987Hong et al, , 1990) are found in HRGP class 18 (PRP bias, high Y) because they have a higher number of EXT motifs than PRP motifs. A sequence (Glyma10g33050.2) similar to a fourth soybean PRP (ENOD2; Franssen et al, 1987) has only 9.8% HRGP motifs and, therefore, is found in MAAB class 24. These findings highlight the need to increase the range of PRP motifs used by MAAB.…”

Conserved regulation of the soybean early nodulin ENOD2 gene promoter in determinate and indeterminate transgenic root nodules

“…Interzone 11-111 constitutes a few cell layers of amyloplast-rich cells between the non-fixing apical zones and nitrogen-fixing zone 111. At A number of temporally and spatially regulated plant genes expressed specifically during nodule development have been characterized (Govers et a/., 1987;van Kammen, 1984;Nap and Bisseling, 1990;Sandal eta/., 1987). Early nodulin genes are expressed shortly after infection with Rhizobia, and are most likely involved in infection and nodule morphogenesis rather than nodule functioning (Nap and Bisseling, 1990 (Dehio and de Bruijn, 1992 …”

Expression of DcPRP1 Is Linked to Carrot Storage Root Formation and Is Induced by Wounding and Auxin Treatment

“…For example, they are expressed during the early stages of pea and soybean root nodule formation (Govers et al, 1986;Franssen et al, 1987;Van de Wiel et al, 1990) and during seedling, leaf, and seed development in soybean (Tierney et al, 1988;Hong et al, 1989;Kleis-San Francisco and Tierney, 1990;Lindstrom and Vodkin, 1991;Wyatt et al, 1992;Suzuki et al, 1993a). In many cases, expression is limited to specific cell types or organs.…”

“…PRPs were first identified as wound-inducible transcripts encoding a cell wall protein (p33) in carrot storage roots (Chen and Varner, 1985;Tierney et al, 1988) and are basic proteins composed mainly of Pro, Hyp, Val, Tyr, Lys, Thr, His, and Glu (Averyhart-Fullard et al, 1988;Kleis-San Francisco and Tierney, 1990;Lindstrom and Vodkin, 1991). Amino acid and DNA sequence analysis indicates that PRPs consist of a signal sequence that is cleaved during secretion of the protein into the cell wall, followed by tandem repetitions of the sequence Pro-Pro-Val-X-Lys (ProHyp-Val-X-Lys in the native protein sequence), where X is often Tyr, His, or Glu (Chen and Varner, 1985;Franssen et al, 1987;Hong et al, 1987;Hong et al, 1990;Datta and Marcus, 1990;Kleis-San Francisco and Tierney, 1990;Lindstrom and Vodkin, 1991;Sheng et al, 1991). Some PRPs (SbPRPl, 2, and 3) are composed only of tandem repeats of ' Salaries and research support were provided in part by a grant from the National Science Foundation (DCB-8911987) and by state and federal funds appropriated to the Ohio Agricultura1 Research and Development Center, The Ohio State University, manuscript NO, * Corresponding author; fax 1-614-292-5379.…”