| Budget Amount *help |
¥15,500,000 (Direct Cost: ¥15,500,000)
Fiscal Year 2005: ¥7,600,000 (Direct Cost: ¥7,600,000)
Fiscal Year 2004: ¥7,900,000 (Direct Cost: ¥7,900,000)
|
|---|
| Research Abstract |
The aim of the present study was to evaluate the functional role of MLPK, a membrane-anchored cytosolic protein kinase found as a causal factor of a self-compatible Brassica mutant. The major findings are summarized as follows.
1.Expression and localization of MLPK.
We identified two MLPK transcripts, termed MLPKf1 and MLPKf2, in stigma. They are generated by using alternative transcriptional start sites. While MLPKf2 is specifically expressed in stigma, MLPKf1 is expressed in almost all tissues. Although both MLPK isoforms localized to the plasma membrane, the membrane localization of MLPKf1 is myristoylation dependent manner, while not in the case of MLPKf2. Transient expression of MLPKf2 complemented the mlpk mutation as in the case of MLPKf1, suggesting that both forms function in the self-incompatibility signaling. The mutated form of MLPKf1, lacking the myristoylation site, failed to complement the mlpk mutation, suggesting that the membrane localization of MLPK is essential for its biological function.
2.Regulatory mechanism of MLPK
Pull down experiment suggested that MLPK does not form a receptor complex with SRK. MLPK was presumed to function in the self-incompatibility signaling downstream of SRK. However, we could not detect the direct interaction nor the reciprocal phosphorylation between SRK and MLPK.
3.Target molecule of MLPK
We searched for MLPK-interacting stigmatic protein by a yeast two-hybrid screening. We selected four candidates and confirmed their MLPK-binding activities in vitro. The four candidates were also shown to be phosphorylated by the recombinant MLPK in vitro.
|
