Biochemical and molecular biological studies on infection (Ascochyta rabiei)-induced thaumatin-like proteins from chickpea plants (Cicer arietinum L.).

A pathogenesis-related protein induced by infection with Ascochyta rabiei was purified from intercellular washing fluid of chickpea (Cicer arietinum L.) leaves. Amino-terminal sequencing identified the protein, named PR-5a, as a thaumatin-like protein. The isoelectric point was determined with 6.5 and the molecular mass is 16 kDa. Therefore, chickpea PR-5a is the first dicot member of a TLP subgroup containing small TLPs with a molecular weight between 15 and 18 kDa. PR-5a shows no antifungal activity towards A. rabiei. Screening of a chickpea cDNA library led to the isolation of a cDNA clone (p5a-241) for this protein. A second cDNA clone (ELR112) encoding a TLP was isolated using differential hybridisation of cDNA libraries obtained from elicited and water treated cell suspension cultures of chickpea. The deduced protein (PR-5b) has a molecular mass of 22 kDa. PR-5b is postulated to be located in the vacuole due to the presence of a respective N-terminal signal peptide and a carboxy-terminal extension. Southern blot analyses showed that ELR112 and p5a-241 represent single copy genes. During fungal infection of chickpea plants expression of both genes proceeds much faster in an A. rabiei resistant cultivar than in a susceptible one.

Genes expressed in Ascochyta rabiei-inoculated chickpea plants and elicited cell cultures as detected by differential cDNA-hybridization.

In response to the exogenous application of elicitors and attempted invasion by pathogens, plants exhibit a wide range of defense reactions. To understand the defense mechanisms at the level of gene activation and deactivation, differential screenings were performed to isolate cDNA clones which are differentially expressed in pathogen-inoculated resistant chickpea plants and elicitor-treated cell cultures. A plenty of genes were isolated and arranged in 5 groups, namely defense-related pathways, signal transduction pathways, regulation of gene expression, catabolic pathways and primary metabolism. Most of these genes were activated although several genes were also found to be suppressed. We discuss the plausible functions of cDNA products in plant defense responses. The cDNAs provide a variety of tools to investigate molecular mechanisms of defense responses and clearly reflect the massive genomic and metabolic changes which occur during manifestation of antimicrobial defense.