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ARS Home » Research » Publications at this Location » Publication #166097


item Hardin, Shane
item Huber, Steven

Submitted to: Plant Physiology and Biochemistry
Publication Type: Abstract Only
Publication Acceptance Date: 5/25/2004
Publication Date: 5/25/2004
Citation: Hardin, S.C., Huber, S.C. 2004. Proteasome activity and the post-translational control of sucrose synthase stability in maize leaves [abstract]. Plant Physiology and Biochemistry. 42:197-208.

Interpretive Summary:

Technical Abstract: The serine-170 (S170) calcium-dependent protein kinase phosphorylation site of maize (Zea mays L.) sucrose synthase (SUS) (EC has been implicated in the post-translational regulation of SUS protein stability. To clarify the proteolytic process and the role of phosphorylation, SUS degradation and proteasome activities were studied in the maize leaf elongation zone. Size-exclusion chromatography resolved two peaks of proteasome-like proteolytic activity. The large molecular mass (1350 kDa) peak required Mg2+ and ATP for maximal activity and was inhibited by the proteasome inhibitors MG132 and NLVS. Anion-exchange chromatography resolved a similar proteolytic activity that was activated by ATP, characteristics that are consistent with those of a 26S-proteasome. S170 phosphorylated SUS (pS170-SUS) was detected as both high molecular mass (HMM) forms and proteolytic fragments that co-eluted with 26S-proteasome activities on both size-exclusion and anion-exchange columns. Conditions that maintained maximal 26S-proteasome activity reduced the amounts of pS170-SUS recovered. In vitro, the 26S-proteasome degraded SUS and proteasome-specific inhibitors reduced SUS proteolysis. HMM-SUS conjugates were produced in vitro and immunoprecipitations suggested that some SUS might be ubiquitinated in vivo. The results suggest that S170 phosphorylation promotes the formation of HMM, ubiquitin-SUS conjugates that are targeted for 26S-proteasome-dependent degradation. Current studies are using directed mutants of SUS1 to establish a role for site-specific phosphorylation in protein degradation in vitro using isolated proteasomes. We are also exploring the possibility that phosphorylation of proteasome subunits may regulate protein degradation activity of the complex.