INDUCTION OF EARLY MEVALONATE PATHWAY ENZYMES AND BIOSYNTHESIS OF END-PRODUCTS IN POTATO (SOLANUM-TUBEROSUM) TUBERS BY WOUNDING AND ELICITATION

In plants, several important classes of terpenoid compounds are synthe sized via the mevalonate pathway. In addition to essential constitutiv e metabolites, potato (Solanum tuberosum L.) tubers synthesize antifun gal sesquiterpenoid phytoalexins in response to fungal infection or ar achidonic acid elicitation, and toxic steroid glycoalkaloids in respon se to wounding. The activity of the early pathway enzyme 3-hydroxy-3-m ethylglutaryl coenzyme A reductase (HMGR) has previously been shown to increase rapidly and then decrease in response to these stimuli. Duri ng an investigation of the possible post-translational control of this enzyme, it was found that the inclusion of the cysteine protease inhi bitors leupeptin and E-64 -3-trans-carboxyran-2-carbonyl)-L-leucyl]agm atine} in the enzyme extraction buffer increased nine-fold the total H MGR activity recovered in the microsomal fraction and greatly increase d the ratio of microsomal to soluble activity. Incubation of microsoma l HMGR preparations with soluble protein extracts, Mg2+ and ATP caused an apparent inhibition of HMGR, consistent with published reports of post-translational inactivation of HMGR by phosphorylation. The appare nt inhibition was completely reversed, however, by 5 mM mevalonate and was found to be an artefact caused by the presence of mevalonate kina se, the next enzyme in the pathway, in the soluble fraction. HPLC assa ys for mevalonate kinase and mevalonate phosphate kinase were develope d and used to measure the activities of these enzymes following woundi ng and elicitation. While HMGR levels increased 30-fold following arac hidonic acid treatment and 15-fold following wounding, mevalonate kina se and mevalonate phosphate kinase only increased two- to four-fold fo llowing these treatments, and the levels in arachidonic acid treated t ubers were only 20-40% higher than in wounded tubers. While HMGR level s are extremely low in untreated tissues, the activities of the two ki nases are relatively high, suggesting that they do not serve as contro l points for the synthesis of terpenoids.