Stored proteinases and the initiation of storage protein mobilization in seeds during germination and seedling growth

Müntz, Klaus; Belozersky, M. A.; Dunaevsky, Yakov E.; Schlereth, Armin; Tiedemann, Jens

doi:10.1093/jexbot/52.362.1741

Cited by 217 publications

(168 citation statements)

References 73 publications

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“…During germination and seedling growth, storage proteins are degraded and mobilized (20), and it is possible that they become unstable as a result of the change of conditions during these stages. For example, the pH in vacuoles decreases from 6.0 to 5.5 during germination (19).…”

Section: Resultsmentioning

confidence: 99%

Crystal structure of soybean 11S globulin: Glycinin A3B4 homohexamer

Adachi

Kanamori

Masuda

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

284

256

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Most plant seeds contain 11S globulins as major storage proteins for their nutrition. Soybean glycinin belongs to the 11S globulin family and consists of five kinds of subunits. We determined the crystal structure of a homohexamer of the glycinin A3B4 subunit at 2.1-Å resolution. The crystal structure shows that the hexamer has 32-point group symmetry formed by face-to-face stacking of two trimers. The interface buries the highly conserved interchain disulfide. Based on the structure, we propose that an ingenious face-to-face mechanism controls the hexamer formation of the 11S globulin by movement of a mobile disordered region to the side of the trimer after posttranslational processing. Electrostatic analysis of the faces suggests that the interchain disulfide-containing face has high positive potential at acidic pH, which induces dissociation of the hexamer into trimers that may be susceptible to proteinases after seed imbibition. This dissociation might result in the degradation and mobilization of 11S globulins as storage proteins in embryos during germination and seedling growth. P lants accumulate protein reserves in developing seeds to act as a sink of nitrogen, sulfur, and carbon. Most dicotyledonous plant seeds contain 7S and͞or 11S globulins and albumins as the major storage proteins in the embryo or cotyledons, whereas some cereals such as wheat, barley, and corn contain prolamins in the endosperm (1, 2). The 11S globulins are distributed more widely than the 7S globulins among plant seeds. Although the quaternary structures of the 7S and 11S globulins are different from each other (the former being a trimeric protein and the latter a hexameric protein), they are believed to be derived from a common ancestor because of the partial homologies in their amino acid sequences and limited proteolysis patterns (3). This assumption has been confirmed by x-ray crystallography of 7S globulins from kidney bean (4), jack bean (5), and soybean (6) and of the trimeric soybean 11S globulin precursor (7).In general, 11S globulins are composed of several kinds of subunits. For example, five major subunits have been identified from soybean: A1aB1b, A2B1a, A1bB2, A3B4, and A5A4B3. In developing seeds, the constituent subunits of 11S globulin are synthesized as a single polypeptide precursor, preproprotein, the signal sequence of which is removed cotranslationally. The resultant proproteins assemble into trimers of Ϸ8S in the endoplasmic reticulum. The proprotein trimers are transported from the endoplasmic reticulum to protein storage vacuoles (PSVs), where they then are cleaved to form acidic and basic polypeptides that are linked by a disulfide bond (8). Finally, the mature proteins assemble into hexamers. The protein reserves are stored in the dormant seed until its germination.Recently we determined the crystal structure of the soybean proglycinin A1aB1b homotrimer at 2.8-Å resolution (7). The protomer consists of N-and C-terminal modules that each include a jelly-roll ␤-barrel and a helix domain. However, the packa...

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Section: Resultsmentioning

confidence: 99%

Crystal structure of soybean 11S globulin: Glycinin A3B4 homohexamer

Adachi

Kanamori

Masuda

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

284

256

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“…Upon seed germination, aleurone cells respond to GA 3 synthesized by the embryo and trigger a signal transduction pathway that prompts the activation of a-amylases and proteases. These hydrolases are secreted and break down the stored reserves in the endosperm, releasing nutrients that support the embryo growth until the onset of photosynthesis (Muntz et al, 2001;Sreenivasulu et al, 2008). In this biological context, the reduction in the level of cystatin transcripts in the aleurone cells treated with GA, together with the GA induction of those genes encoding cathespin L-like proteases, indicate their respective functions as counterparts in the same biological process.…”

Section: Discussionmentioning

confidence: 99%

Characterization of the Entire Cystatin Gene Family in Barley and Their Target Cathepsin L-Like Cysteine-Proteases, Partners in the Hordein Mobilization during Seed Germination

et al. 2009

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Plant cystatins are inhibitors of cysteine-proteases of the papain C1A and legumain C13 families. Cystatin data from multiple plant species have suggested that these inhibitors act as defense proteins against pests and pathogens and as regulators of protein turnover. In this study, we characterize the entire cystatin gene family from barley (Hordeum vulgare), which contain 13 nonredundant genes, and identify and characterize their target enzymes, the barley cathepsin L-like proteases. Cystatins and proteases were expressed and purified from Escherichia coli cultures. Each cystatin was found to have different inhibitory capability against barley cysteine-proteases in in vitro inhibitory assays using specific substrates. Real-time reverse transcriptionpolymerase chain reaction revealed that inhibitors and enzymes present a wide variation in their messenger RNA expression patterns. Their transcripts were mainly detected in developing and germinating seeds, and some of them were also expressed in leaves and roots. Subcellular localization of cystatins and cathepsin L-like proteases fused to green fluorescent protein demonstrated the presence of both protein families throughout the endoplasmic reticulum and the Golgi complex. Proteases and cystatins not only colocalized but also interacted in vivo in the plant cell, as revealed by bimolecular fluorescence complementation. The functional relationship between cystatins and cathepsin L-like proteases was inferred from their common implication as counterparts of mobilization of storage proteins upon barley seed germination. The opposite pattern of transcription expression in gibberellin-treated aleurones presented by inhibitors and enzymes allowed proteases to specifically degrade B, C, and D hordeins stored in the endosperm of barley seeds.

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“…As has been suggested for glycinin, limiting the degree of trimer interaction through subunit heterogeneity or electrostatic repulsion on the IE face may promote disassembly and mobilization of seed storage proteins during germination [7]. Such interactions will also be affected by changes in pH and ionic strength that occur within the PSV after imbibition [42,43].…”

Section: Implications Of Cruciferin Phosphorylation For Processing Amentioning

confidence: 99%

Phosphorylation of the 12 S globulin cruciferin in wild-type and abi1-1 mutant Arabidopsis thaliana (thale cress) seeds

Wan

Ross

Yang

et al. 2007

Biochemical Journal

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Cruciferin (a 12 S globulin) is the most abundant storage protein in the seeds of Arabidopsis thaliana (thale cress) and other crucifers, sharing structural similarity with the cupin superfamily of proteins. Cruciferin is synthesized as a precursor in the rough endoplasmic reticulum. Subunit assembly is accompanied by structural rearrangements involving proteolysis and disulfidebond formation prior to deposition in protein storage vacuoles. The A. thaliana cv. Columbia genome contains four cruciferin loci, two of which, on the basis of cDNA analysis, give rise to three alternatively spliced variants. Using MS, we confirmed the presence of four variants encoded by genes At4g28520.1, At5g44120.3, At1g03880.1 and At1g3890.1 in A. thaliana seeds. Two-dimensional gel electrophoresis, along with immunological detection using anti-cruciferin antiserum and antibodies against phosphorylated amino acid residues, revealed that cruciferin was the major phosphorylated protein in Arabidopsis seeds and that polymorphism far exceeded that predicted on the basis of known isoforms. The latter may be attributed, at least in part, to phosphorylation site heterogeneity. A total of 20 phosphorylation sites, comprising nine serine, eight threonine and three tyrosine residues, were identified by MS. Most of these are located on the IE (interchain disulfide-containing) face of the globulin trimer, which is involved in hexamer formation. The implications of these findings for cruciferin processing, assembly and mobilization are discussed. In addition, the protein phosphatase 2C-impaired mutant, abi1-1, was found to exhibit increased levels of cruciferin phosphorylation, suggesting either that cruciferin may be an in vivo target for this enzyme or that abi1-1 regulates the protein kinase/phosphatase system required for cruciferin phosphorylation.

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Stored proteinases and the initiation of storage protein mobilization in seeds during germination and seedling growth

Cited by 217 publications

References 73 publications

Crystal structure of soybean 11S globulin: Glycinin A3B4 homohexamer

Crystal structure of soybean 11S globulin: Glycinin A3B4 homohexamer

Characterization of the Entire Cystatin Gene Family in Barley and Their Target Cathepsin L-Like Cysteine-Proteases, Partners in the Hordein Mobilization during Seed Germination

Phosphorylation of the 12 S globulin cruciferin in wild-type and abi1-1 mutant Arabidopsis thaliana (thale cress) seeds

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