Acquired disease resistance can be induced in rose by acibenzolar-S-methyl (BTH), a novel synthetic chemical which has been shown to induce a broad-spectrum disease resistance in many plant species. BTH was applied by dipping whole in vitro rose shoots into the chemical at different concentrations for a few seconds before returning them to shoot growth medium. Four days later the shoots were challenge inoculated in vitro with Diplocarpon rosae or Agrobacterium tumefaciens under otherwise aseptic conditions. Pretreatment of rose shoots with BTH led to resistance to D. Rosae and A. tumefaciens by significantly reducing the disease severity of blackspot and frequency of crown gall formation and gall sizes, respectively. The induced disease resistance by BTH may provide a novel approach to protect rose plants from pathogen infection in the field. The biochemical mechanism behind the induced resistance was investigated based on analyses of protein changes in the intercellular spaces of rose leaves by SDSPAGE, 2D-PAGE and western blotting of 1D- or 2D-gels against antisera of the four major classes of tobacco PR proteins. The results indicate that BTH-mediated enhanced resistance in rose was accompanied by the induction and accumulation of a set of extracellular proteins, including PR-1, PR-2, PR-3 and PR-5 proteins. Most of the extracellular proteins activated by BTH were also induced and found to accumulate in leaves upon infection with D. rosae. However, their accumulation was much more pronounced in BTH-pretreated leaves than in water-pretreated leaves upon a challenge inoculation with D. rosae. The induction and accumulation of PR-2 and PR-3 proteins in the intercellular spaces of BTH-treated leaves or BTH-pretreated leaves followed by infection with the pathogen was further confirmed by assays of β-1,3-glucanase and chitinase activities. All these results demonstrate that extracellular PR proteins were involved in BTH-induced disease resistance in rose. However, not all isoforms of the PR proteins seemed to be associated with rose disease resistance based on western blots and native PAGE with specific staining of β-1,3-glucanase and chitinase activities. Some isoforms of the PR proteins such as 15 kDa PR-1, 36 and 37 kDa PR-2 proteins may be more important in the expression of resistance, whereas some isoforms may be involved in the growth and development of rose shoots. In conclusion, here it has been demonstrated that BTH is an effective inducer of PR proteins and acquired disease resistance in rose.