Induction of apoplastic caspase- 3 like activity attributable to a subtilisin- like protease in potato- Phytophthora infestans interaction

DALEO; FERNANDEZ; LAMENZA; DALEO; GUEVARA

Austin

Congreso; Plant Biology 2012 ASPB; 2012

American Society of Plant Biologist

Caspases, a group of cysteine aspartate specific proteases, are universal mediators of animal apoptosis, a type of programmed cell death (PCD). Despite in plants there are not animal caspase orthologues, several caspase- like activities have been characterized in different plant PCD models, including during defense responses. Here we report the involvement of caspase- 3 like activity (DEVDase) during disease development induced by P. infestans infection in potato leaves. This DEVDase activity was mainly localized in the apoplast after 48h of infection, and was sensitive to serine (PMSF) and serine and cysteine protease (chymostatin) inhibitors. DEVDase activity from total protein extracts accounts for a 60% of serine proteases; however, this percentage increases to 100% in the apoplast. In order to purify apoplastic proteases with DEVDase activity, intercellular washing fluids (IWF) from 24h and 48h infected potato leaves were incubated with biotinylated chymostatin. Precipitated protein complexes were analyzed by 12% SDS- PAGE and mass spectrometric analysis of two bands (28.5 and 16.5 kDa.) showed sequence similarity to Solanaceae subtilisin- like proteases. Other extracellular Ser- dependent proteases from the subtilase family, phytaspases and saspases, do not exert DEVDase activity. Thus, caspase- 3 like activity found during potato- P. infestans interaction, represents a new extracellular serine protease with caspase- like activity. The importance of this activity to restrict pathogen spread remains elusive. Further assays would be necessary in order to elucidate the role of caspase-3- like activity in potato resistance to P. infestans. *Equal contribution.