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item Horvath, David

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 3/16/2005
Publication Date: 3/22/2005
Citation: Sonju, R., Horvath, D.P. 2005. Cloning and expression of KRP genes from adventitious buds of the perennial weed leafy spurge. [Abstract]. Midwestern Society of the American Society of Plant Biology.

Interpretive Summary: We have cloned several slightly different copies (alleles) a key gene involved in regulating growth from the perennial noxious weed leafy spurge. This gene is a member of a small family of cell cycle regulatory known as KIP-related proteins (KRP’s). The gene we have cloned and characterized from leafy spurge is most similar to the KRP4 genes from Arabidopsis and several other model plants. KRP4 from leafy spurge appears to be very rapidly “turned off” in underground buds when they are induced to grow when the above-ground portion of the plant is cut off. Also, KRP4 appears to be turned off in field-grown buds in August when such buds are beginning to enter a dormant state in preparation for fall. We have also identified two other members of this KRP gene family (KRP3 and KRP7) from our collection of 35,000 sequenced genes from leafy spurge. Additionally, we are attempting to clone a copy of KRP1 or KRP2 from leafy spurge since these genes have been shown to play a critical role in growth inhibition in response to drought and cold stress in model plants. Towards this end, we have prepared short sequences (primers) needed to amplify (make lots of copies of) these genes and are in the process of determining if our amplified DNA is from KRP1 or KRP2.

Technical Abstract: We are interested in studying bud dormancy in the perennial weed leafy spurge. Adventitious buds of leafy spurge are capable of exhibiting all three types of dormancy (para-dormancy or apical dominance, endo-dormancy or innate dormancy, and eco-dormancy). Dormancy is defined as the temporary cessation of growth, and thus implies regulation of the cell division process. KIP-related proteins (KRP’s), also known as inhibitor of cyclin-dependent kinases (ICKs) are inhibitors of cell division. Their function is known to be through the binding of cyclins and cyclin-dependent kinases (CDK’s) thereby negatively regulating their activity of these cell cycle promoting complexes. We have hypothesized that KRP’s potentially play a role in dormancy through inhibition of cell cycle in dormant buds. Thus, our goal has been to clone and characterize the expression of KRP genes in buds of leafy spurge. KRP3, 4, and 7, were identified and cloned by several different methods, including primer design from related species, and blast searches of our leafy spurge EST database. Attempts to identify KRP1, 5 and 6 are ongoing. Studies performed to analyze the temporal expression of these genes following dormancy release, and in various tissues and after a gauntlet of treatments have provided information needed for understanding the role of KRPs in dormancy. Semi-quantitative RT-PCR on total RNA derived from the root buds of leafy spurge in a time course experiment revealed that expression of KRP4 was relatively higher during dormancy and the initial hours after growth induction and decreased to a basal level during growth. Similar experiments using RNA from buds collected at various times during the growing season demonstrated that KRP4 was slightly up-regulation in August and September, a time when the buds are considered to be para-dormant.