Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/35938
標題: 百合花粉管中一個附著於內質網表面的蛋白質(LLDNAJ)之特性分析 百合花粉管中一個附著於內質網表面的蛋白質
Characterization of an ER-associated protein,LLDNAJ,in germinating lily pollen tube
作者: 周享渝
Yu, Shiang
關鍵字: LLDNAJ
百合蛋白質
cochaperone
Hsp70
GST pull down
內質網
護衛蛋白
出版社: 生物科技學研究所
摘要: DnaJ為J-protein 家族以及Hsp40家族的一員,且為Hsp70的co-chaperone。在細胞中,護衛蛋白 (chaperone) 能防止新合成的蛋白質以及變性的蛋白質因不正確的折疊而產生的聚集現象。前人從鐵砲百合的花粉中得到一個全長DnaJ cDNA並將之命名為LLDNAJ。研究顯示LLDNAJ基因的表現具有花粉專一性,並且只在花粉發育後期表現。本研究針對LLDNAJ蛋白質在花粉萌發及花粉管生長期間的表現特性,及其可能的功能作進一步的探討,其中包括了該蛋白質在不同組織部位及花粉不同發育時期的表現、蛋白質在細胞中的定位,以及其分子功能的分析。鐵砲百合的花粉管中,我們利用於E. coli 表達之GST-LLDNAJ 重組蛋白質進行GST pull down的實驗証實:LLDNAJ蛋白質會和花粉管內推測的Hsp70結合。另外,免疫定位實驗發現LLDNAJ蛋白位於生長之在花粉管的細胞質及內質網中。由於花粉的萌發及花粉管的生長是一個非常快速的過程,為了因應此特性,花粉必須利用內質網和高基氏體合成大量的細胞膜和細胞壁物質,並利用分泌型囊泡將其運送到花粉管尖端快速生長區域,這其中包含了許多分泌蛋白或膜蛋白。因此我們推測LLDNAJ會與其相對應之Hsp70共同幫助這些大量的蛋白質,於粗內質網進行post-translational transportation,進而達成提供花粉管快速生長所需之結構物質。
DnaJ is a member of J-protein family that is also known as Hsp40 family molecular co-chaperones for Hsp70. The major function of this cellular chaperone machinery is to prevent the aggregation of long, unfolded nascent or denatured proteins. One lily pollen DnaJ cDNA, LLDNAJ, was obtained from previous study and RT-PCR results showed its pollen specific and late developmental expression patterns. In this study, we focused on characterization the spatial and temporal expression, localization and possible function of LLDNAJ/protein during lily pollen germination and pollen tube growth. GST pull down assay using GST-LLDNAJ recombinant protein as a bait revealed that LLDNAJ protein interacts with a putative Hsp70 partner in germinating pollen tube of lily. Immunolocalization study indicated that LLDNAJ protein mainly localized in the cytosolic side of ER in the germination pollen tube. Pollen germination and tube growth are very rapid events. To support this unusually fast growth, large amounts of membrane and cell wall materials including the integrated and secreted proteins are synthesized in ER and Golgi apparatus, and transported to plasma membrane at tip region by secreted vesicles. We hypothesize that the potential LLDNAJ-Hsp70 molecular chaperone is involved in the active protein synthesis by post-translational transportation occurring on rER in the fast germinating lily pollen tube.
URI: http://hdl.handle.net/11455/35938
Appears in Collections:生物科技學研究所

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