| Summary: | 碩士 === 國立陽明大學 === 生物化學研究所 === 92 === The Arabidopsis TUBBY-like protein (AtTLP) gene family has been recently characterized as a distinct class of F-box proteins in Arabidopsis. AtTLP9, one member in this family, has been shown to be involved in phytohormone abscisic acid (ABA) signaling (Lai et al., 2004). In the present work, AtTLP2, a representative member of AtTLPs, was shown to specifically interact with Arabidopsis SKP1-like 1 (ASK1) using the yeast two-hybrid system. Serial deletion analyses demonstrated that both F-box domain and tubby domain were required for AtTLP2-ASK1 interaction. Three AtTLPs (AtTLP5, AtTLP7 and AtTLP8) were shown to be unable to interact with ASK1. Domain swapping indicated the replacement of F-box domain of AtTLP2 by that of AtTLP7 resulted in the loss of interacting activity with ASK1. Bioinformatics analysis suggested that residue P52 in AtTLP2 corresponding to P113 in human SKP2 might be involved in AtTLPs-ASK1 interaction. Site directed mutagenesis of P52L in AtTLP2 was confirmed to greatly reduce the interacting activity with ASK1. Transient expression of AtTLP2 in Arabidopsis mesophyll protoplasts indicated that AtTLP2 is localized in the nucleus. These results strongly suggest that AtTLP2 probably acts through targeting some regulatory protein(s) in nucleus for ubiquitin-mediated proteolysis.
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