A DUF4281 domain-containing protein (homologue of ABA4) of Phaeodactylum tricornutum regulates the biosynthesis of fucoxanthin
A DUF4281 domain-containing protein (homologue of ABA4) of Phaeodactylum tricornutum regulates the biosynthesis of fucoxanthin
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三角褐指藻含有 DUF4281 结构域的蛋白（ABA4 同源物）调节岩藻黄质的生物合成
DOI：
10.1016/j.algal.2022.102728
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发表时间：
2022-06
期刊：
影响因子：
--
通讯作者：
Jichang Han
中科院分区：
文献类型：
--
作者：
Xiaotong Shen;Kehou Pan;Lin Zhang;Baohua Zhu;Yun Li;Jichang Han
岩藻黄质是最有价值的类胡萝卜素之一，但其生物合成途径尚不清楚。新藻黄素近年来被认为是岩藻黄素的中间体，但仍有许多细节有待探索。先前在拟南芥中进行的研究表明，含有duf4281结构域的蛋白（ABA4）是新黄质生物合成所必需的，ABA4的突变导致色素组成的巨大改变。在三角褐指藻（Phaeodactylum tricornutum）中，一种功能未知蛋白（本研究命名为PtABA4）与ABA4具有高度同源性，并且含有DUF4281结构域。为了研究其对新黄素合成和岩藻黄素积累的作用，我们分别对PtABA4进行了内源性过表达和敲低（RNA干扰），并利用重组蛋白进行了体外活性测定。PtABA4的过表达和敲低分别导致岩藻黄素的急剧增加和急剧减少，表明PtABA4在岩藻黄素积累中起着关键作用。出乎意料的是，PtABA4基因的下调导致了新黄质上游类胡萝卜素的减少，而不是代偿性的增加。在体外活性测定方面，大肠杆菌产生的重组蛋白和过表达的转化蛋白都没有表现出新黄质合成酶的活性。综合以上结果，我们推测PtABA4可能与其他膜相关酶构成一种多酶复合物，参与多种类胡萝卜素的形成，而不仅仅是新黄质。DUF4281结构域在所有分类群中的普遍存在以及细菌中叶黄素的缺失为我们的假设提供了间接证据。总之，本研究为通过基因工程方法提高岩藻黄质含量提供了一个靶基因，加深了对DUF4281结构域功能的认识。•PtABA4对岩藻黄质积累起重要作用。•本研究提供了提高岩藻黄质含量的靶基因。•本研究加深了对DUF4281结构域功能的认识。
Although fucoxanthin is one of the most valuable carotenoids , its biosynthesis pathway is still poorly understood. Neoxanthin has been regarded as the intermediate of fucoxanthin in recent years, however, multiple details still need to be explored. Previous study performed in Arabidopsis thaliana demonstrated a DUF4281-domain containing protein (ABA4) was necessary for the biosynthesis of neoxanthin, and the mutation of ABA4 led to a huge alteration of pigment composition. In Phaeodactylum tricornutum , a function unknown protein (designated as PtABA4 in this study) displayed high homology to ABA4, and also contained DUF4281 domain. To investigate its function on neoxanthin synthesis and fucoxanthin accumulation, we conducted endogenous overexpression and knockdown (RNA interference) of PtABA4 individually, as well as in vitro activity assay using recombinant protein . The overexpression and knockdown of PtABA4 lead to sharp increases and decrease of fucoxanthin, respectively, demonstrating a key role of PtABA4 on fucoxanthin accumulation. Unexpectedly, the knockdown of PtABA4 results in the decreases of upstream carotenoids of neoxanthin, rather than compensatory increases. In terms of in vitro activity assay, neither the recombinant protein produced by Escherichia coli nor the overexpressed transformant displays neoxanthin synthase activity. Taking all results into consideration, we speculate that the PtABA4 may constitute one multienzyme complexes with other membrane associated enzymes and participate in the formation of several carotenoids, rather than neoxanthin only. The ubiquity of DUF4281 domain across all taxa and the absence of xanthophylls in bacteria provide indirect evidence for our assumption. Above all, this study here provides a target gene for enhancing the fucoxanthin content by genetic engineering methods, and deeps the understanding about the function of DUF4281 domain. • PtABA4 plays an important role on fucoxanthin accumulation. • This study provides a target gene for enhancing the fucoxanthin content. • This study deeps the understanding about the function of DUF4281 domain.
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