| Abstract: | 生理多樣性是基因多樣性的結果之一，更是各物種適應其生態棲位的基礎，但也是現今討論生物多樣性時，最常被忽視的一環。螃蟹廣泛分佈在各種棲地(如：深海熱泉、各類海域、潮間帶、河口區、溪流、陸地；是在地球上分佈最廣、生存棲地變異最大的類群之一。這個廣佈的特性，使本類群非常合適用來進行生理多樣性的研究。在適應這些環境時，螃蟹的調控機制又視物種以及其適陸性，有很大的變異與多樣性。主要參與調節機制的鰓是一個多功能的器官，至少負責氣體交換、離子調節、含氮廢物的排除及酸鹼平衡等重要功能。這些功能是由多種酵素的共同作用完成，其中最主要的酵素是Na+, K+-ATPase、 carbonic anhydrase 以及H+-ATPase。目前主要的論點是：在海水環境時，離子調節主要是藉由細胞基底膜上Na+, K+-ATPase 的主動運輸，產生離子濃度差來作為離子調節的原動力﹔而在淡水的調節機制中，卻是藉由細胞頂部(apical)膜上的H+-ATPase 來做為原動力 (Morris 2001)。這樣的觀點，有許多值得商榷的地方。因此本計劃的核心問題是，由較為原始的海生型螃蟹到陸生型或淡水型螃蟹間，三種酵素序列變動的速度，是否相同? 有沒有在哪一個類群的螃蟹中，發現三種酵素的分佈不相同的? 這些變異具有什麼適應上的意義或價值? 本計劃將利用(1) 對三種酵素進行Molecular cloning、比對及分析各自的相對變異程度，(2) 以quantitative RT-PCR 對三種酵素同步測量其相對量，並且(3) 以穿透式電子顯微鏡的方式，觀察金粒子在離子調節細胞的位置。
Physiological biodiversity is one of the end products of genetic biodiversity, and it is also the basis of how a species possesses its pariticular ecological niche. However, it is overlooked by most of the biodiversity issues. Crabs are one of the few taxa that can survive in almost any kind of habitats, including hydrothermal vent, seawater, freshwater, intertidal, estuarine and terrestrial environments. This feature makes the crustaceans ideal for studying physiological biodiversity. In adapting to these environments, there is a great variation and diversity in the physiological mechanisms, depending on the species and its terrestriality. Crab gill is a multi-functional organ responsible for respiration, ion regulation, nitrogen excretion and acid-base balance. These physiological functions are controled by several enzymes, including Na+, K+-ATPase、carbonic anhydrase and H+-ATPase. Currently, it is widely accepted that the basolateral Na+, K+-ATPase is the driving force for osmoregulation in seawater, whereas the apical H+-ATPase being the one for that in fresh water. However, some of the points deserve further investigation. The major questions asked in this proposal are: 「From the presumably more primitive marine crabs to terristrial or freshwater species, are the rates for amino acid substitutions among the three key enzymes the same? Do these crabs have a similar sub-cellular distribution for these three enzymes? What is the adaptive significance for any variation? For all the three enzymes, I plan to include (1) molecular cloning for estimating the rates of the amino acid substitutions and phylogenetic analyses, (2) quantitative RT-PCR for monitoring their relative abundance, and (3) TEM examination for the gold particle localization. |
