Author
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Chao, Wun |
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SERPE, MARCELO - BOISE STATE UNIVERSITY |
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Submitted to: Meeting Abstract
Publication Type: Abstract Only Publication Acceptance Date: 6/10/2004 Publication Date: 7/23/2004 Citation: Chao, W.S., Serpe, M.D. 2004. Does dormancy break in root bud of leafy spurge (Euphorbia esula L.) require phosphorylation of CDK-activating kinase (CAK)?. [Abstract]. July 24-28, 2004. Lake Buena Vista, FL. Interpretive Summary: Leafy spurge is a perennial weed that propagates vegetatively from an abundance of underground adventitious buds (UABs) located on the roots and crown and is the primary characteristic leading to its invasive nature. These buds develop during the normal growing season but are maintained in a quiescent state through correlative inhibition. Existing evidence indicates that growth-arrest may be a result of interactions between signaling pathways controlling dormancy and those controlling the cell cycle. To enhance our understanding of growth and development during vegetative propagation, we have cloned a cell cycle gene from leafy spurge encoding CDK-activating kinase (CAK). This enzyme is involved in a phosphorylation cascade linked to early stages of cell cycle progression. An antibody for leafy spurge CAK has been generated. We have examined the expression and biochemical character of this gene and protein using RT-PCR, RNA and protein blot analysis, immunoprecipitation, and phosphorylation in dormant and non-dormant UABs of leafy spurge. Current data shows that CAK is expressed constitutively at both transcriptional and translational levels, indicating a preference of post-transcriptional regulation. Site-directed mutagenesis of CAK identified that two threonine residues are mutually responsible for autophohphorylation and for phosphorylating its substrate protein, cyclin-dependent kinase (CDK). In addition, autophosphorylation of CAK is an intra-molecular event, and ATP concentration directly affects CAK's activity on CDK phosphorylation. We are in the process of generating a phosphoprotein-specific antibody. The regulation of CAK phosphorylation in relation to dormancy break will be reported. Key words: Leafy Spurge, Dormancy, CDK-Activating Kinase. Technical Abstract: Leafy spurge is a perennial weed that propagates vegetatively from an abundance of underground adventitious buds (UABs) located on the roots and crown and is the primary characteristic leading to its invasive nature. These buds develop during the normal growing season but are maintained in a quiescent state through correlative inhibition. Existing evidence indicates that growth-arrest may be a result of interactions between signaling pathways controlling dormancy and those controlling the cell cycle. To enhance our understanding of growth and development during vegetative propagation, we have cloned a cell cycle gene from leafy spurge encoding CDK-activating kinase (CAK). This enzyme is involved in a phosphorylation cascade linked to early stages of cell cycle progression. An antibody for leafy spurge CAK has been generated. We have examined the expression and biochemical character of this gene and protein using RT-PCR, RNA and protein blot analysis, immunoprecipitation, and phosphorylation in dormant and non-dormant UABs of leafy spurge. Current data shows that CAK is expressed constitutively at both transcriptional and translational levels, indicating a preference of post-transcriptional regulation. Site-directed mutagenesis of CAK identified that two threonine residues are mutually responsible for autophohphorylation and for phosphorylating its substrate protein, cyclin-dependent kinase (CDK). In addition, autophosphorylation of CAK is an intra-molecular event, and ATP concentration directly affects CAK's activity on CDK phosphorylation. We are in the process of generating a phosphoprotein-specific antibody. The regulation of CAK phosphorylation in relation to dormancy break will be reported. Key words: Leafy Spurge, Dormancy, CDK-Activating Kinase. |
