Tissue and Cell-Type Localization and Partial Characterization of a Xylem Papain-Type Cysteine Protease From Arabidopsis

Cysteine proteases are associated with xylem tracheary element differentiation. XCP1 was recently identified as a xylem-specific cysteine protease in Arabidopsis (Zhao, et al., 2000). For this study a recombinant polyhistidine-tagged XCP1 (XCP1H6) was expressed and purified from an E. coli expression system. A polyclonal anti-XCP1 antibody was produced using purified XCP1H6. Immunoblot analysis of a developmental time course of xylem and bark protein extracted from root-hypocotyl segments demonstrated that XCP1 was expressed in xylem only. Further analysis under optimized immunoblot conditions, however, revealed that anti-XCP1 antibody reacted with protein present in both xylem and bark. The vast majority of immunoreactivity, however, was restricted to xylem. Cell-type localization of GUS expression under the control of a putative XCP1 promoter indicated that the XCP1 promoter specifies expression of XCP1 in tracheary elements in leaves, stems, roots and flowers. XCP1 promoter-driven GUS activity was not associated with senescing tissues.