Transient and specific expression of a cysteine endopeptidase associated with autolysis during differentiation of Zinnia mesophyll cells into tracheary elements.

Endopeptidase activities during the differentiation of Zinnia cells into tracheary elements (TEs) were examined with several peptidyl 4-methyl-7-coumarylamido (MCA) as substrates. The activity that hydrolysed carbobenzoxy-Phe-Arg-MCA (Z-Phe-Arg-MCA) at pH 5 increased in a differentiation-related manner: this activity, which was not observed in freshly isolated mesophyll cells was induced by the combination of alpha-naphthaleneacetic acid (NAA) and 6-benzyladenine (BA) that is necessary for differentiation of TEs, but not by NAA or BA alone. The activity in cells cultured in TE-inductive medium that contained both NAA and BA increased very rapidly between the 48th and 60th hours of culture, when the number of TE increased rapidly. A protease responsible for this activity with a molecular mass of 30 kDa was partially purified from cells which had been cultured in the TE-inductive medium and included many immature TEs. Strong inhibition by [L-3-trans-carboxyoxiran-2-carbonyl]-L-Leu- -agmatin (E-64), and activation by dithiothreitol (DTT) indicated that this protease belongs to a family of cysteine endopeptidases (EC 3.4.22).