【摘要】：海帶是一種重要的大型海洋經(jīng)濟藻類,具有富集碘元素的生物學(xué)特性,是生物界中重要的碘存儲器之一,其吸收、儲存與釋放過程是碘生物地球化學(xué)循環(huán)的重要組成部分。海帶中碘積累的主要形態(tài)是碘化物,它參與藻體的免疫反應(yīng),可以有效清除體內(nèi)的活性氧(ROS),所以,海帶中的碘是一種非常有效的無機抗氧化劑。在逆境條件的刺激下,藻體中的碘會釋放,在海帶葉片表面以及質(zhì)外體,碘化物可以和體內(nèi)產(chǎn)生的氧化劑或者空氣中的臭氧反應(yīng),從而導(dǎo)致大量的分子碘釋放到體表。分子碘隨后會被臭氧O3氧化成高吸濕性的碘氧化物,逐步形成云凝集核,從而影響O3的含量。由此可見,海帶對碘元素的生物地球化學(xué)循環(huán)起到不可或缺的作用。另外,在海帶的碘代謝過程中有一種關(guān)鍵酶—釩依賴型鹵代過氧化物酶(vanadium-dependent haloperoxidase,vHPOs),在催化鹵素離子與過氧化氫反應(yīng)生成鹵素的氧化形態(tài)的過程中發(fā)揮著重要的催化作用。此外,vHPOs還可能參與調(diào)控海帶游孢子體和配子體附著發(fā)育及細胞壁增厚機制。因此,研究海帶中的碘代謝機制具有一定的可行性和較高的研究價值。受人類活動與全球氣候變化的雙重影響,大氣中二氧化碳(CO2)排放量持續(xù)增加,造成溫室效應(yīng),引起了全球變暖。海洋作為一個天然的CO2儲存庫,吸收了人類排放CO2總量的約30%,又引起海洋酸化現(xiàn)象。海洋系統(tǒng)在碘元素的生物地球化學(xué)循環(huán)中是重要的組成部分,而海帶作為海洋中重要的碘儲存器,全球氣候變化所引起的海洋酸化及溫度升高可能會影響海帶中碘元素的含量,從而影響人類食用碘的含量。另外,低潮狀態(tài)下的海帶會將碘釋放到空氣中,而我國又是世界上海帶栽培面積和養(yǎng)殖產(chǎn)量最大的國家,主要養(yǎng)殖品種為日本真海帶(Saccharina japonica),由海洋酸化引起的海帶中碘含量的改變會影響碘的生物地球化學(xué)循環(huán)。由于海帶中碘含量隨氣候變化會對人類健康及碘的生物地球化學(xué)循環(huán)兩方面均有重要影響,本研究從實驗生態(tài)學(xué)和分子生物學(xué)兩方面探究海帶(S.japonica)中碘代謝對氣候變化的應(yīng)答與調(diào)控機制。首先,我們通過設(shè)置不同的溫度及酸化梯度,探究海帶(S.japonica)對海水溫度升高和酸化的應(yīng)答;其次,我們對海帶中碘的抗氧化機制進行了研究;最后由于vHPOs在海帶碘代謝中重要的催化作用,我們在海帶(S.japonica)全基因組測序的基礎(chǔ)上,對其中的vHPOs基因家族進行鑒定和結(jié)構(gòu)功能分析。本文的主要研究結(jié)果如下:1)采用單因子實驗,分別研究溫度和酸化誘導(dǎo)對海帶碘代謝的影響。實驗結(jié)果表明,溫度變化和酸化誘導(dǎo)均對海帶中碘含量有顯著影響,在實驗溫度5℃,10℃,15℃,20℃,23℃范圍內(nèi),海帶中碘含量呈現(xiàn)出先升高后下降的趨勢,在15℃取得最大值;酸化實驗結(jié)果表明,在海水pCO2(二氧化碳分壓)(400μatm,700μatm,1000μatm,1500μatm,2000μatm)范圍內(nèi),海帶中碘含量也呈現(xiàn)先升高后下降的趨勢,在1000μatm取得最大值,且酸化培養(yǎng)對海帶碘吸收具有促進作用。此外,15℃和20℃與酸化耦合在一定程度上對海帶中的碘含量具有促進作用。在海區(qū)中開展的中試酸化實驗也驗證了海洋酸化條件下海帶的個體大小以及總碘含量明顯高于正常條件下的海帶,但總體上隨著海帶的生長其碘含量逐漸下降。另外,本研究通過對海帶中無機形態(tài)碘的測定發(fā)現(xiàn),碘離子的變化趨勢與總碘的變化趨勢一致,而碘酸根離子不在檢測限度內(nèi)。最后,通過在海帶的培養(yǎng)體系中添加100μg/m L的低聚古羅糖醛酸,發(fā)現(xiàn)在3h時海帶中碘釋放速率達到最大。結(jié)果顯示當海帶在氧化應(yīng)激狀態(tài)時,體內(nèi)碘離子大量釋放與活性氧反應(yīng)來消除活性氧,起到了抗氧化作用。2)通過生物信息學(xué)方法在S.japonica基因組中檢索鑒定出75個vHPO基因,其中58個釩依賴型碘過氧化物酶(vanadium-dependent iodoperoxidase,vIPO),17個釩依賴型溴過氧化物酶(vanadium-dependent bromoperoxidase,vBPO)。通過與其他真核和原核生物的vHPOs全長蛋白質(zhì)序列構(gòu)建系統(tǒng)進化樹,發(fā)現(xiàn)所有的vHPOs蛋白質(zhì)被分為三個不同的亞家族:釩依賴型氯過氧化物酶(vanadium-dependent chloroperoxidase,vCPO)、vBPOs和vIPOs。通過進一步分析進化樹拓撲結(jié)構(gòu)發(fā)現(xiàn)海洋藻類中的vHPO起源于真菌的vCPO,其中vBPO在紅藻和褐藻中獨立進化,海帶中vIPO由褐藻中的vBPO祖先進化而來。3)通過對海帶(S.japonica)雌雄配子體、小孢子體及成熟孢子體不同部位的轉(zhuǎn)錄組數(shù)據(jù)分析發(fā)現(xiàn),vHPOs在海帶不同世代及組織中具有明顯的表達特異性。在海帶不同世代中,雌、雄配子體中表達的vHPO的數(shù)目最多,其次是小孢子體世代,最后是成熟孢子體。在成熟孢子體的不同組織中,梢部和根莖表達的vHPO數(shù)目最多。另外,為了從分子生物學(xué)水平上揭示脅迫條件下vHPOs基因表達量的變化,我們對高溫及CO2充氣培養(yǎng)3h下S.japonica的熒光定量PCR結(jié)果分析發(fā)現(xiàn)高溫及CO2充氣培養(yǎng)條件下各有8個vHPO基因表達量明顯增加,推測這些基因在高溫及酸化脅迫條件下的海帶碘的吸收中發(fā)揮重要作用。
[Abstract]:Zosterae marinae is an important large-scale marine economic algae, which has the biological characteristics of enriching the iodine element, is one of the important iodine storage in the biological community, and the absorption, storage and release process of the kelp is an important part of the biogeochemical cycle of the iodine. The main form of iodine accumulation in the kelp is iodide, which participates in the immune response of the algae body and can effectively remove the active oxygen (ROS) in the body, so the iodine in the kelp is a very effective inorganic antioxidant. Under the stimulation of the stress condition, the iodine in the algae body is released, and in the surface of the laminaria leaf and the mass outer body, the iodide can react with an oxidizing agent or an ozone generated in the body to cause a large amount of molecular iodine to be released to the body surface. The molecular iodine is then oxidized to the high-hygroscopic iodine oxide by the ozone O3, and the cloud aggregation core is gradually formed, so that the content of the O3 is affected. It can be seen that kelp plays an indispensable role in the biogeochemical cycle of iodine. In addition, in the process of iodine metabolism of the kelp, a key enzyme, such as vanadium-dependent haloperoxidase (vHPOs), plays an important catalytic role in the process of generating the oxidation state of the halogen by the reaction of the catalytic halogen ion and the hydrogen peroxide. In addition, vHPOs may also be involved in the regulation of the attachment and development of the sporophytes and gametophytes of Laminaria japonica and the mechanism of cell wall thickening. Therefore, the study of the iodine metabolism mechanism in the kelp has a certain feasibility and higher research value. The double effects of human activities and global climate change, the continued increase in carbon dioxide (CO2) emissions in the atmosphere, contributing to global warming. The oceans, as a natural CO2 repository, absorb about 30% of the total amount of CO2 emitted by humans and cause ocean acidification. The marine system is an important part of the biogeochemical cycle of the iodine element, while the sea tangle is an important iodine storage in the ocean, and the ocean acidification and the temperature rise caused by the global climate change can affect the content of the iodine element in the kelp, thereby affecting the content of the human edible iodine. in addition, the kelp in the low-water state can release the iodine into the air, and the country is the country with the largest culture area and the breeding yield in the world, and the main breeding variety is the Saccharina japonica, The change of iodine content in the kelp caused by ocean acidification can affect the biogeochemical cycle of iodine. Since the content of iodine in the sea tangle has an important influence on the biological and geochemical cycle of human health and iodine, the response and regulation mechanism of iodine metabolism in the sea tangle (S. japonica) to climate change is explored from the aspects of experimental ecology and molecular biology. First, we study the response of the sea-tangle (S. japonica) to the temperature rise and the acidification of the sea water by setting different temperature and acidification gradients; secondly, we study the anti-oxidation mechanism of iodine in the kelp; and finally, because of the important catalytic effect of vHPOs in the iodine metabolism of the kelp, On the basis of the whole genome sequencing of the S. japonica, we identified and structure the vHPOs gene family. The main results of this paper are as follows:1) The effect of temperature and acidification on the metabolism of kelp iodine was studied by single factor experiment. The experimental results show that the change of temperature and the induction of acidification have a significant effect on the content of iodine in the kelp, and the content of iodine in the kelp is at the temperature of 5 鈩，

本文編號：2483590