MAPK (mitogen-activated protein kinase)訊息傳遞途徑可以傳遞外界的訊號，以引發植物生理及形態上的適應，進而調節植物的生長與發育。本研究主要在分析紅杉（Sequoia sempervirens ）新找到之SsMAPKA及SsMEKA之特性及功能，由我們的結果顯示，SsMAPKA基因表現與紅杉生長相的轉變有關聯性表現；另外，SsMEKA基因會隨著植物生長相的轉變而發生剪接選擇性（alternative splicing）。
　　根據親緣演化分析之結果，SsMAPKA屬於A2族群之MAPK，其氨基酸序列與芫荽PsD5具有82%之相同度。以南方墨點分析證實SsMAPKA基因在紅杉基因組中為單一基因。為進一步了解SsMAPKA基因之特性與功能，以北方墨點分析顯示，SsMAPKA基因在癒傷組織、根、莖及葉均有表現；在生長相的轉變上，SsMAPKA基因在紅杉老樹、1x至3x嫁接植株的頂端及基部的表現較多於4x至5x嫁接植株及年輕的植株。而在酵素活性的偵測上，於E.coli中大量表現GST-SsMAPKA融合蛋白，再以anti-phosphotyrosine抗體偵測發現，僅有GST-SsMAPKA以IPTG誘導後，可見一約64kDa分子量的蛋白質有被磷酸化的特性。更進一步選殖SsMAPKA基因啟動子，取得全長為1838bp，結果顯示，SsMAPKA之啟動子存在著11個ABREs的序列（ACGT）在cis-acting element的部位，另外尚有2個ABREs-like的序列（ACGTG）及6個與花粉發育（AGAAA）相關的序列。我們推測SsMAPKA在紅杉生長相的轉變可能扮演著調控的角色。
　　實驗選殖SsMAPKA上游調控因子―SsMEKA。根據序列分析結果顯示，在紅杉植株中共找到4種不同型式的MEKs，由親緣演化分析結果顯示，SsMEKA屬於A2族群之MEK，與菸草的NtMEK1有65％以上的相似度，參與植物細胞分裂的過程。此4種不同型式的MEKs，其中一種型式為在序列上多出一小段42bp的片段，含有intron的特性，將其命名為SsMEKA ⅡA。以南方墨點分析證實SsMEKA基因在紅杉基因組中為單一基因。在組織特異性表現分析顯示，SsMEKA基礎轉錄量均相同。分析SsMEKA ⅡA以RT-PCR產物，經由定序及南方墨點分析顯示，此基因在紅杉老樹、1x至3x嫁接植株表現較多於4x至5x嫁接植株及年輕的植株。此為MAPK cascades調節植物生長與發育中，首次提出中游的MEK經由選擇性剪接參與植物的發育。
　　我們的實驗亦首度發現，樹木成熟過程中，genomic DNA重組參與在紅杉生長相的轉變。

　　Phase change or maturation can be observed in all higher plants and affects reproductive competence, the morphology and growth rate of the vegetative body, as well as the regenerative potential of tissue explants. Efforts to elucidate the underlying mechanism of phase change of plants, or maturation and rejuvenation, have disclosed biochemical correlations, some of which could aid in furthering the investigations. Hence, the aims of this project are to characterize these phase-regulated signal transduction components.
　　Plants have developed elaborate mechanisms to perceive external signals and to manifest adaptive responses with proper physiological and morphological changes. The mitogen-activated protein (MAP) kinase cascade is one of the well-characterized intracellular signaling modules, and that cross-talk between various signal-transduction pathways might be concentrated at MEK level in plant MAPK cascades. In this study, two novel genes of Sequoia sempervirens, SsMAPKA, and SsMEKA have been identified and characterized.
　　According to the phylogenetic analysis based on the amino acid sequences, SsMAPKA belongs to the Group A2 MAPK subfamily, and shares high homology with PsD5 (82% identity) from Pisum sativum. Southern blot analysis showed that SsMAPKA is a single copy gene in S. sempervirens. Northern blot analysis showed that SsMAPKA was expressed in all plant organs. The mRNA level of SsMAPKA was accumulated in adult and 1x- to 3x- grafted shoots than in juvenile and 4x- to 5x- grafted shoots. SsMAPKA was fused to glutathione-S-transferase (GST) and the fusion protein was expressed in Escherichia coli. Results showed that the recombinant SsMAPKA was recognized by monoclonal anti-phosphotyrosine antibody. This result indicated that SsMAPKA is a functional protein kinase that has features characteristics of MAP kinases. The promoter of the SsMAPKA gene was isolated from S. sempervirens. The cloned promoter regions upstream of the transcription start site of SsMAPKA gene is 1,838bp. The most prominent matches identified include those to the consensus binding size for ABA-responsive promoters（ACGT or ACGTG）and for pollen-specific regulatory elements（AGAAA）.
　　Then we made an attempt to isolate the SsMAPKA upstream factor, SsMEKA in signal transduction components. Four types of S. sempervirens PCR product have been isolated that encode putative protein kinases, designated SsMEKA ⅠA, SsMEKA ⅠAs, SsMEKA ⅡA and SsMEKA ⅡAs. According to the phylogenetic analysis based on the amino acid sequences, SsMEKA belongs to the Group A2 MEK subfamily. These kinases exhibit a high degree of homology to NtMEK1（65％ identity）, a tobacco protein that is a membrane of the family of mitogen-activated protein kinase kinase （MAPKKs）, which appears to function in a cell-cycle-dependent manner. One of these isoforms SsMEKA IIA which contained the region that flanked the 42bp intron-like sequence was found in S. sempervirens. Southern blot analysis showed that SsMEKA is a single copy gene in S. sempervirens. Tissue-specific analysis indicated that the basal transcription level of SsMEKA was expressed in all organs in plants. Our results revealed that transcripts of the SsMEKA ⅡA gene accumulated predominantly in adult and 1x- to 3x- grafted shoots, and a small amount of the transcript was found in juvenile and 4x- to 5x- grafted shoots of S. sempervirens. Furthermore, we provide the first report that concentrated MEK by alternative splicing regulating phase change of plants in the MAPK cascades.
　　In this study, we propose that DNA rearrangement involved in woody plant maturation.

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