Journal Article

The Rice Nucellin Gene Ortholog <i>OsAsp1</i> Encodes an Active Aspartic Protease Without a Plant-specific Insert and is Strongly Expressed in Early Embryo

Xuezhi Bi, Gurdev S. Khush and John Bennett

in Plant and Cell Physiology

Published on behalf of Japanese Society of Plant Physiologists

Volume 46, issue 1, pages 87-98
Published in print January 2005 | ISSN: 0032-0781
Published online January 2005 | e-ISSN: 1471-9053 | DOI: http://dx.doi.org/10.1093/pcp/pci002
The Rice Nucellin Gene Ortholog OsAsp1 Encodes an Active Aspartic Protease Without a Plant-specific Insert and is Strongly Expressed in Early Embryo

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  • Biochemistry
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The barley nucellin gene was reported to be nucellus specific in its expression and was hypothesized to play a role in the programmed cell death of the nucellus as an aspartic protease. Here we provide direct evidence that the rice ortholog encodes an active aspartic protease, but we prefer the name aspartic protease1 (OsAsp1) to nucellin after a detailed analysis of its expression pattern in rice and barley. Northern blots, RT–PCR and RNA in situ hybridization showed that OsAsp1 is expressed most abundantly in zygotic embryos 1–2 d after fertilization. It is also expressed in pollen, nucellus, ovary wall, shoot and root meristem, coleoptiles of immature seeds, and somatic embryos. A parallel study in barley showed that the barley nucellin gene was expressed not only in the nucellus but also strongly in embryos. Recombinant protein proOsAsp1 expressed in the bacterium Escherichia coli refolded and autolysed at acidic pH 3.5 in vitro, and the mature peptide displayed protease activity. Nucellin has three close homologs in rice on chromosomes 11 and 12 and in Arabidopsis on chromosomes 1 and 4. They lack the plant-specific insert that distinguishes the typical plant aspartic protease from aspartic proteases of other organisms. They constitute a new class of aspartic protease that is present in both monocots and dicots but whose function remains to be explored further.

Keywords: Keywords: Aspartic protease — Embryo — Gene expression — Recombinant protein — Rice (Oryza sativa L.).; Abbreviations: DAF, days after flowering; DIG, digoxygenin; DH, double haploid; ER, endoplasmic reticulum; EST, expressed sequence tag; FTC, fluorescein thiocarbamoyl; IPTG, isopropyl 1-thio-β-d-galactopyranoside; MALDI-TOF, matrix-assisted laser desorption/ionization time of flight; PCD, programmed cell death; PSI, plant-specific insert; PTS, peroxisome targeting sequence; RT–PCR, reverse transcriptase–polymerase chain reaction; SDS-PAGE, UTR, untranslated region.

Journal Article.  8342 words.  Illustrated.

Subjects: Biochemistry ; Molecular and Cell Biology ; Plant Sciences and Forestry

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