Research Accomplishment Reports 2007

26S Proteasome-Dependent Control of Cytokinin Sensitivity

J.A. Smalle, J. Kurepa
Department of Plant and Soil Sciences

 

Project Description

Activities:

Analysis of the degradation rates of cytokinin response regulators in wild type and 26S proteasome mutant backgrounds.

Determining the Ubiquitin-dependent and -independent proteolysis rates in 26S proteasome mutants.

Results of this project were presented at the 19th IPGSA meeting in Puerta Vallarta (Mexico, July 2007) and at the CSREES Genes to Products Awardee Workshop in Washington (March, 2007)

Products:

Novel insights into 26S and 20S Proteasome function in plant cells. Identification of unstable proteins in the cytokinin response pathway.

New method to determine Ubiquitin-dependent degradation rates in plant cell extracts.

Transgenic Arabidopsis lines expressing epitope-tagged ARR1 and ARR5 in wild type and proteasome mutant backgrounds.

Impact

During the previous year, the cytokinin response regulators ARR5 and ARR1 were shown to be targets for 26S proteasome-dependent proteolysis. While using proteasome mutants for this project it was discovered that these mutant lines, as expected, have decreased Ub-dependent proteolysis, but surprisingly have increased Ub-independent proteolysis rates due to an increase in the abundance of the 20S proteasome. This finding is important for the correct interpretation of the cytokinin response phenotypes of these mutants. While it is expected that the cytokinin response regulators are degraded by the proteasome in a Ub-dependent manner, it remains possible that the degradation of these targets does not require prior ubiquitination. In the latter case, the original hypothesis driving this research needs to be adjusted. Instead of a stabilization of a cytokinin response inhibitor explaining the decreased cytokinin sensitivity of these mutants, a more likely explanation would be that the degradation of a response activator is enhanced. To distinguish between these two possibilities, transgenic lines have been generated that express MYC-tagged ARR1 and ARR5. These lines have been crossed into the proteasome mutants rpn10-1 and rpn12a-1. Currently, the degradation rates of these MYC-tagged ARRs in wt and proteasome mutant backgrounds are being compared as well as in response to cytokinin treatments.

Publications

Kurepa J., A. Toh-e, and J. Smalle. 2007. 26S proteasome regulatory particle mutants have increased oxidative stress tolerance. The Plant Journal (in press).

Kurepa J., and J. Smalle. 2007. Structure, function and regulation of plant proteasomes. Biochimie. (in press).

Kurepa J., Y. Li, and J. Smalle. 2007. 26S Proteasome-dependent control of cytokinin signaling. CSREES Genes to Products Awardee Workshop Abstract Book. Washington, March, 2007

Kurepa J., Y. Li, and J. Smalle. 2007. 26S Proteasome-dependent control of cytokinin signaling. 19th IPGSA meeting Abstract Book. Puerta Vallarta, Mexico. July, 2007