Subtilase activity and gene expression during germination and seedling growth in barley

Proteases play a main role in the mobilization of storage proteins during seed germination. Until today, there is little information about the involvement of serine proteases, particularly subtilases, in the germination of barley grains. The aims of the present work were to study the contribution of serine proteases to the total proteolytic activity induced during germination of barley grains and evaluate the specific involvement of subtilases in this process. Proteolytic activity assayed against azocasein in the presence of specific inhibitors, showed that serine proteases contributed between 10 and 20% of total activity along germination. Subtilase activity increased from day 1 after imbibition with a peak between days 4-5. Moreover, in vivo determination of subtilase activity in germinating grains revealed increasing activity along germination mainly localized in the seed endosperm and developing rootlets. Finally, the expression of 19 barley genes encoding subtilases was measured by real time PCR during germination. Three of the analyzed genes increased their expression along germination, five showed a transient induction, one was down-regulated, nine remained unchanged and one was not expressed. The present work demonstrates the involvement of subtilases in germination of barley grains and describes the positive association of eight subtilase genes to this process.