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Title: Mutational analysis of the major soybean UreF paralogue involved in urease activation

Author
item Bilyeu, Kristin
item Hyten, David
item SLEPER, DAVID - University Of Missouri
item POLACCO, JOE - University Of Missouri

Submitted to: Journal of Experimental Botany
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/31/2011
Publication Date: 6/1/2011
Citation: Bilyeu, K.D., Hyten, D.L., Sleper, D., Polacco, J. 2011. Mutational analysis of the major soybean UreF paralogue involved in urease activation. Journal of Experimental Botany. 62(10):3599-3608.

Interpretive Summary: Soybean yields are in part defined by the plants’ ability to efficiently coordinate the uptake and movement of nutrients to their ultimate deposition into seeds. One enzyme important in nitrogen movement is activated urease, a nickel containing protein that has been conserved from bacteria through the plant kingdom. Unlike the situation in bacteria, it was not known which genes were responsible for the activation of the urease enzyme in soybean. The objective of this work was to investigate the molecular genetic basis for a mutation in soybean that did not allow soybean urease activation. The new genomic resources available including the soybean genome sequence and access to a new screening technology enabled the localization and identification of the mutant gene, an accessory protein similar to one that exists in bacteria. Knowledge of the urease activation process will contribute to a broader understanding of mechanisms that include plant defense, metal insertion to activate enzymes, and transport and mobilization of storage reserves that could impact yields.

Technical Abstract: In soybean, mutation at Eu2 or Eu3 eliminates the urease activities of both the embryo-specific and the tissue-ubiquitous (assimilatory) isozymes, encoded by Eu1 and Eu4, respectively. Eu3 encodes UreG, a GTP’ase necessary for proper emplacement of Ni and carbon dioxide in the urease active site. However, Eu2 has not been mapped nor its product identified. In bacteria and Arabidopsis thaliana, UreD and UreF, in addition to UreG, are essential for in vivo urease activation. The soybean genome contains a single gene encoding UreG/Eu3, and two paralogs each for UreD and UreF. We report that F2 bulked segregant analysis localizes Eu2 to soybean chromosome 2: Each of two Eu2 missense alleles co-segregates with mutation in the UreF paralog on chromosome 2 (Ch02UreF). This is the first map-based recovery of a soybean gene (Eu2) via the Universal Soy Linkage Panel (USLP1.0). Ch02UreF has 95% deduced AA identity with the Ch14UreF paralog, which was shown to complement a UreF mutant of the fungus, Schizosaccharomyces pombe. However, in soybean, Ch02UreF represents >99% of the total UreF urease-activation function - a conclusion supported by a third, null Ch02UreF Eu2 allele. Transcripts of Ch02UreF and Ch14UreF accumulate to the same extent, with highest levels in developing embryos—the stage of most active urease synthesis. Thus, the exact role of the Ch14UreF paralog remains undetermined.