Role of propeptide glycan in post-translational processing and transport of barley lectin to vacuoles in transgenic tobacco.

Wilkins, Thea A.; Bednarek, Sebastian Y.; Raikhel, Natasha V.

doi:10.1105/tpc.2.4.301

Cited by 82 publications

(65 citation statements)

References 45 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…In support of this hypothesis is the fact that retention of the vacuolar pea vicilin protein in the ER by engineering a ER retention signal onto the protein stabilized the modified protein 100-fold over unmodified vicilin protein when synthesized in leaves of transgenic plants (Wandelt et al, 1992). However, many vacuolar proteins such as the pea lectin (Edwards et al, 1991), barley lectin (Wilkins et al, 1990), and cowpea trypsin inhibitor (Hilder et al, 1987) have been shown to be stable in leaves of transgenic plants. Thus, it follows that the stability of a protein is not determined solely by its subcellular location but may be controlled by the intrinsic properties of the protein itself.…”

mentioning

confidence: 84%

Accumulation of 15-Kilodalton Zein in Novel Protein Bodies in Transgenic Tobacco

et al. 1995

View full text Add to dashboard Cite

Zeins, the seed storage proteins of maize, are a group of alcoholsoluble polypeptides of different molecular masses that share a similar amino acid composition but vary i n their sulfur amino acid composition. They are synthesized on the rough endoplasmic reticulum (ER) in the endosperm and are stored in ER-derived protein bodies. Our goal i s to balance the amino acid composition of the methionine-deficient forage legumes by expressing the sulfur amino acid-rich 15-kD zeins in their leaves. However, it is crucial to know whelher this protein would be stable i n nonseed tissues of transgenic plants. The major focus of this paper is to compare the accumulation pattern of the 15-kD zein protein with a vacuolar targeted seed protein, p-phaseolin, in nonseed tissues and to determine the basis for i t s stability/instability. We have introduced the 15-kD zein and bean p-phaseolin-coding sequences behind the 35s cauliflower mosaic virus promoter into tobacco (Nicofiana fabacum) and analyzed the protein's accumulation pattern in different tissues. Our results demonstrate that the 15-kD seed protein i s stable not only i n seeds but in all nonseed tissues tested, whereas the P-phaseolin protein accumulated only i n mid-and postmaturation seeds. Interestingly, zein accumulates in novel protein bodies both in the seeds and in nonseed tissues. We attribute the instability of the p-phaseolin protein i n nonseed tissues to the fact that it is targeted to protease-rich vacuoles. The stability of the 15-kD zein could be attributed to its retention i n the ER or to the proteaseresistant nature of the protein.Seed storage proteins constitute a potentially useful class of proteins for the improvement of forage crops if they can be made to accumulate in leaves. It is a fairly simple genetic engineering feat to introduce a seed protein-coding sequence behind a strong constitutive promoter into transgenic plants and ensure high rates of synthesis of the corresponding protein. However, stability of the protein in an alien environment is still not clearly defined and has to be treated on a case by case basis.Seed proteins are synthesized during seed development and accumulate in protein bodies. These proteins are then utilized by the emerging seedling during germination. The major seed protein in dicotyledonous plants are the saltsoluble globulins, which are stored in vacuole-derived protein bodies. Most monocot seed storage proteins are '

show abstract

mentioning

confidence: 84%

Accumulation of 15-Kilodalton Zein in Novel Protein Bodies in Transgenic Tobacco

et al. 1995

View full text Add to dashboard Cite

show abstract

“…Although other plant proteins have been expressed and shown to be correctly processed in transgenic tobacco (Hoffman et al, 1987;Sturm et al, 1988;Sonnewald et al, 1989;Wilkins et al, 1990), expression of heterologous proteins in this system sometimes results in accumulation of molecular forms not normally observed in the native tissue (Higgins et al, 1988;Post-Beitenmiller et al, 1989;Altabella and Chrispeels, 1990). It is, therefore, significant that polygalacturonase, whose complex maturation process includes core glycosylation, oligosaccharide modification, and at least three proteolytic processing steps, appears to undergo identical processing events in the transgenic host, including the heterogeneous reactions associated with maturation of the PG2A and PG2B isozymes.…”

Section: Discussionmentioning

confidence: 99%

Analysis of tomato polygalacturonase expression in transgenic tobacco.

et al. 1990

View full text Add to dashboard Cite

Tomato polygalacturonase is a cell wall enzyme secreted in large amounts during tomato fruit ripening. Polygalacturonase is synthesized as a glycoprotein precursor that undergoes numerous cotranslational and post-translational processing steps during its maturation, yielding three isozymes in tomato fruit, PG1, PGPA, and PGPB. To investigate the physiological roles of the three isozymes and the functional significance of the polygalacturonase processing domains in its intracellular transport and activity, we have examined polygalacturonase expression in transgenic tobacco plants. A full-length polygalacturonase cDNA was placed under control of the cauliflower mosaic virus 35s promoter and introduced into tobacco by way of Agrobacterium-mediated transformation. Analysis of transgenic tobacco plants indicated that (1) immunologically detectable polygalacturonase can be extracted from leaves, roots, and stems of transgenic tobacco plants; ( 2 ) only PGSA and PGPB were detectable in transgenic tobacco; (3) the polygalacturonase isozymes present in transgenic tobacco were electrophoretically indistinguishable from the tomato isozymes; (4) the N-terminal sequence, degree of N-linked glycosylation, and extent of oligosaccharide processing were similar in polygalacturonase from transgenic tobacco and tomato; (5) polygalacturonase was properly localized in cell walls of transgenic tissue; (6) the protein was enzymically active in vitro; however, (7) accumulation of PGPA and PGSB in cell walls of transgenic tobacco did not result in pectin degradation in vivo. These results indicated that tomato polygalacturonase was properly processed and transported to the cell wall of tobacco. However, accumulation of the two polygalacturonase isozymes expressed in this heterologous host was insufficient to promote polyuronide degradation in tobacco leaf tissue.

show abstract

“…Peroxisomal targeting was amino termini (Subramani, 1992;van den Bosch et ai., 1992). In adeletional analysis of the propeptide that contains ayeast vacuolar targeting signal, QRPL, Johnson et al (1987) found All barley lectin (BL) mutant carboxyl-terminal propeptide (CTPP) constructs were prepared by site-specific mutagenesis as described by ther by modification of the CTPP coding region of the wild-type clone described by Wilkins et al (1990) or by the addition of specific nucleotide sequences between the final codon for the mature Drotein and ako mediated by different signals 'Ocated at the carboxy and et ai. (1990).…”

Section: Preparation Of Bl-mutant Ctpp Constructsmentioning

confidence: 99%

Determination of the functional elements within the vacuolar targeting signal of barley lectin.

et al. 1993

Self Cite

View full text Add to dashboard Cite

We have previously demonstrated that the carboxyl-terminal propeptide of barley lectin is both necessary and sufficient for protein sorting to the plant vacuole. Specific mutations were constructed to determine which amino acid residues or secondary structural determinants of the carboxyl-terminal propeptide affect proper protein sorting. We have found that no consensus sequence or common structural determinants are required for proper sorting of barley lectin to the vacuole. However, our analysis demonstrated the importance of hydrophobic residues in vacuolar targeting. In addition, at least three exposed amino acid residues are necessary for efficient sorting. Sorting was disrupted by the addition of two glycine residues at the carboxyl-terminal end of the targeting signal or by the translocation of the glycan to the carboxy terminus of the propeptide. These results suggest that some components of the sorting apparatus interact with the carboxy terminus of the propeptide.

show abstract

Role of propeptide glycan in post-translational processing and transport of barley lectin to vacuoles in transgenic tobacco.

Cited by 82 publications

References 45 publications

Accumulation of 15-Kilodalton Zein in Novel Protein Bodies in Transgenic Tobacco

Accumulation of 15-Kilodalton Zein in Novel Protein Bodies in Transgenic Tobacco

Analysis of tomato polygalacturonase expression in transgenic tobacco.

Determination of the functional elements within the vacuolar targeting signal of barley lectin.

Contact Info

Product

Resources

About