Characterisation of cDNAs homologous to Rab5-GTP binding protein expressed during early somatic embryogenesis in chicory

• Published in: Plant science (Limerick) Vol. 162; no. 3; pp. 413 - 422
• Main Authors: Randoux, Béatrice, Quillet, Marie-Christine, Rambaud, Caroline, Vasseur, Jacques, Hilbert, Jean-Louis
• Format: Journal Article
• Language: English
• Published: Shannon Elsevier Ireland Ltd 01.03.2002; Elsevier Science
• Subjects: Agronomy. Soil science and plant productions; Biological and medical sciences; Cichorium; Differential display; Economic plant physiology; Fundamental and applied biological sciences. Psychology; In vitro culture; Plant physiology and development; Rab5, GTP-protein; Somatic embryogenesis; Tissue cultures, protoplasts; Differential display; Cichorium; S.E., somatic embryogenesis; Rab5, GTP-protein; Somatic embryogenesis; Embryonic development; Homology; Compositae; Somatic embryo; Characterization; Binding protein; Vegetable crop; Complementary DNA; Gene; Dicotyledones; Angiospermae; Spermatophyta; Aminoacid sequence
• ISSN: 0168-9452; 1873-2259
• DOI: 10.1016/S0168-9452(01)00582-9

Transcripts from the Cichorium hybrid ‘474’ ( Cichorium intybus L., var. sativum X C. endivia L., var. latifolia) leaf tissues were compared by differential display to identify cDNA corresponding to genes that are expressed early during somatic embryogenesis (S.E.). B1.5, one of the isolated cDNAs had a strong sequence similarity with small Ypt/Rab5 GTP binding proteins. Northern blot analyses showed that transcripts hybridising with this cDNA were present in leaf tissues of the embryogenic responsive genotype ‘474’ submitted to S.E.-inducing conditions, but were absent under these conditions when using a non-embryogenic responsive chicory cultivar. Performing 5′RACE PCR, we were able to identify CHI-GTP1 and CHI-GTP2, 2 cDNAs that represent two highly homologous genes or two alleles of one gene encoding Rab5-like GTP-binding proteins. Amplification of genomic DNA revealed the presence of 2–4 genes in the chicory ‘474’ hybrid genome. The 2 proteins deduced from CHI-GTP1 and CHI-GTP2 are 200 amino-acids long with sizes of about 22 kDa. They differed only in 3 AA. Comparing the different sequences obtained in chicory with GTP-binding proteins from others plants B1.5 most likely represent a product obtained from CHI-GTP1. Expressed in Escherichia coli, the CHI-GTP1 protein was able to bind GTP confirming its identity. In view of the proposed roles of Rab5 GTP-binding protein, the possible function of the protein encoded by CHI-GTP1 in S.E. process is discussed.