Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress

Verona, Italy - 24/27 September, 2003

ISBN 88-900622-4-X

Poster Abstract - 5.24

A. THALIANA NONSYMBIOTIC HAEMOGLOBIN 2 (AHB2): EXPRESSION PROFILE ANALYSIS AND CHARACTERIZATION OF TRANSGENIC PLANTS ALTERED IN THE LEVEL OF THE PROTEIN

C. ANZI, D. EDERLE, E. ZAGO, M. DELLEDONNE

Università degli Studi di Verona, Dipartimento Scientifico e Tecnologico, Strada Le Grazie 15, 37134 Verona

nonsymbiotic plant haemoglobin, gene expression

Two different groups of haemoglobins have been found in plants: symbiotic and non-symbiotic. The first group has been well characterized in legumes and other symbiotic plants, and its function during the symbiotic process is now clear. On the other hand, nonsymbiotic plant haemoglobins (nsHb) represent a challenge for researchers as they are widespread in plant kingdom, but we are still far from finding their proper (if any) physiological function. Phylogenetic analysis showed the existence of two class of nsHb in dicots. Arabidopsis thaliana possesses two non-symbiotic haemoglobins, AHb1 (class 1) and AHb2 (class 2). AHb1 is induced in root and rosette leaves by low oxygen levels. AHb1 has a very high oxygen affinity. Analysis of plants grown on MS0 medium revealed a low constitutive level of AHb2 transcript in rosette leaves of A. thaliana and the induction of AHb2 transcription was observed during cold stress. AHb2 shows a lower affinity for oxygen than AHb1. Here we show that the level of protein is extremely stable. Recombinant AHb2 protein, expressed in E. coli, was purified and used to obtain polyclonal rabbit antisera for western blot analysis. We never observed changes in protein amount, no matter what the level of the corresponding transcript, suggesting a post transcriptional control of AHb2 level. Moreover, to get a deeper insight, we monitored mRNA and protein level in plants grown in soil. This analysis showed a different AHb2 transcription profile between plants grown on MS0 medium or in soil, but once again, in both growth condition protein level didn’t change. To test whether an increased level of GLB2 has effects on plant physiology, we generated transgenic plants containing 35S::AHb2 construct. We selected three lines: one showing a lower protein level than wild type, not detectable with western blot analysis; the other two lines having higher protein content. Their phenotypic and molecular characterization is presented.