Molecular cloning of yieldins regulating the yield threshold of cowpea cell walls: cDNA cloning and characterization of recombinant yieldin

• Published in: Plant, cell and environment cell and environment, 2000-02, Vol.23 (2), p.155-164
• Main Authors: Okamoto‐Nakazato, A., Takahashi, K., Kido, N., Owaribe, K., Katou, K.
• Format: Article
• Language: English
• Subjects: Accession: AB028025 (Vu‐YLD) in DDBJ database; Agronomy. Soil science and plant productions; Biological and medical sciences; Cowpea hypocotyl; Economic plant physiology; Fundamental and applied biological sciences. Psychology; General aspects; glycerinated hollow cylinder; Growth and development; molecular cloning of yieldin; Morphogenesis, differentiation, rhizogenesis, tuberization. Senescence; Plant physiology and development; recombinant yieldin; Vegetative apparatus, growth and morphogenesis. Senescence; Vigna unguiculata; yield threshold; yieldin
• ISSN: 0140-7791; 1365-3040
• DOI: 10.1046/j.1365-3040.2000.00526.x

ABSTRACT cDNA for yieldin of Vigna unguiculata L. was cloned with reverse transcriptase‐polymerase chain reaction using synthetic oligonucleotides as primers. The primers were designed on the basis of the N‐terminal amino acid sequence of the yieldins isolated from the wall preparation of cowpea hypocotyls. The 1·2 kbp cDNA for yieldin contained an open reading frame of 981 base pairs, encoding 327 amino acids including 23 amino acids as a putative signal sequence. An homology search of the deduced amino acid sequence revealed that the yieldin was homologous to acidic class III endochitinases (EC 3·2.1·14) and concanavalin B. A cDNA fragment containing the yieldin‐coding region was introduced to Escherichia coli cells using an expression vector to express the recombinant protein. The recombinant yieldin was obtained from the recombinant E. coli and its effect on the wall mechanical properties was examined by reconstitution experiments. The recombinant yieldin fully restored the acid‐induced change of the yield threshold tension (y) of heat denatured glycerinated hollow cylinders (GHCs) of cowpea hypocotyls. Northern hybridization analysis revealed that the yieldin mRNA was expressed mainly in the rapid and moderate elongation region of the etiolated hypocotyl of Vigna unguiculata L.