PI-55

Preparation Method

PI-55 (6-(2-hydroxy-3-methylbenzylamino)purine) was first dissolved in DMSO to obtain a 10mM stock solution and was kept at low temperature in the dark. Immediately before use, the stock was diluted with kinase assay buffer so that the final DMSO content in the reaction did not exceed 1%. Recombinant Arabidopsis thaliana cytokinin receptors (CRE1/AHK4 or AHK3) were incubated in a defined reaction mixture containing ATP and an appropriate peptide or protein substrate. PI-55 was added at the indicated concentrations to evaluate its ability to interfere with cytokinin-dependent receptor activity, typically in the presence of a natural cytokinin such as trans-zeatin.

Reaction Conditions

1–50µM, 30min.

Applications

PI-55 acted as a competitive antagonist of cytokinin receptors. At low micromolar concentrations it suppressed trans-zeatin–induced signaling, with CRE1/AHK4 exhibiting the highest sensitivity.

Cultured tissue lines

Arabidopsis thaliana tissues such as seedlings, root explants, and ARR5:GUS reporter lines, together with medicinal plant seedlings including Bulbine natalensis and Rumex crispus maintained on MS-based media, are commonly used systems for evaluating the biological activity of PI-55 in plant research.

Preparation Method

PI-55 was prepared as a 10 mM DMSO stock and diluted into sterile culture medium immediately before use. Explants or seedlings were transferred onto solid or liquid medium supplemented with PI-55 at the required concentrations, while control cultures received the same amount of DMSO without PI-55. The compound was present throughout the cultivation period to continuously modulate cytokinin perception during organogenesis and stress treatments. In experiments combining PI-55 with heavy metal stress, cadmium was added to the medium together with PI-55, and seedlings were grown under controlled light and temperature conditions for several days to weeks.

Reaction Conditions

0.1–5µM, 7–21 days.

Applications

PI-55 suppressed cytokinin signaling, shifting organogenesis toward root formation. At low micromolar concentrations (0.1–10µM, 7–21 days), it reduced cadmium-induced growth inhibition, supported biomass accumulation, maintained balanced hormone activity, and enhanced stress tolerance.

References:
[1]. Spíchal, Lukás et al. “The purine derivative PI-55 blocks cytokinin action via receptor inhibition.” The FEBS journal vol. 276,1 (2009): 244-53.
[2]. Aremu, Adeyemi O et al. “Dissecting the role of two cytokinin analogues (INCYDE and PI-55) on in vitro organogenesis, phytohormone accumulation, phytochemical content and antioxidant activity.” Plant science : an international journal of experimental plant biology vol. 238 (2015): 81-94.