Table_1_Regulatory and functional divergence among members of Ibβfruct2, a sweet potato vacuolar invertase gene controlling starch and glucose content.docx

Sweet potato [Ipomoea batatas (L.) Lam.] is an important food and industrial crop. Its storage root is rich in starch, which is present in the form of granules and represents the principal storage carbohydrate in plants. Starch content is an important trait of sweet potato controlling the quality and yield of industrial products. Vacuolar invertase encoding gene Ibβfruct2 was supposed to be a key regulator of starch content in sweet potato, but its function and regulation were unclear. In this study, three Ibβfruct2 gene members were detected. Their promoters displayed differences in sequence, activity, and cis-regulatory elements and might interact with different transcription factors, indicating that the three Ibβfruct2 family members are governed by different regulatory mechanisms at the transcription level. Among them, we found that only Ibβfruct2-1 show a high expression level and promoter activity, and encodes a protein with invertase activity, and the conserved domains and three conserved motifs NDPNG, RDP, and WEC are critical to this activity. Only two and six amino acid residue variations were detected in sequences of proteins encoded by Ibβfruct2-2 and Ibβfruct2-3, respectively, compared with Ibβfruct2-1; although not within key motifs, these variations affected protein structure and affinities for the catalytic substrate, resulting in functional deficiency and low activity. Heterologous expression of Ibβfruct2-1 in Arabidopsis decreased starch content but increased glucose content in leaves, indicating Ibβfruct2-1 was a negative regulator of starch content. These findings represent an important advance in understanding the regulatory and functional divergence among duplicated genes in sweet potato, and provide critical information for functional studies and utilization of these genes in genetic improvement.

甘薯（Ipomoea batatas (L.) Lam.）作为一种重要的粮食和工业作物，其储存根富含淀粉，淀粉以颗粒形式存在，并构成了植物中的主要储存碳水化合物。淀粉含量是控制甘薯工业产品质量与产量的重要性状。编码液泡性蔗糖异构酶的基因Ibβfruct2曾被认为是调控甘薯淀粉含量的关键调节因子，但其功能与调控机制尚不明确。本研究检测到了三个Ibβfruct2基因成员，其启动子表现出序列、活性和顺式调控元件的差异，并可能与其他转录因子相互作用，表明这三个Ibβfruct2家族成员在转录水平上受不同的调控机制所控制。其中，我们发现仅Ibβfruct2-1展现出高表达水平和启动子活性，并编码具有蔗糖异构酶活性的蛋白质，其中保守结构域和三个保守基序NDPNG、RDP和WEC对该活性至关重要。与Ibβfruct2-1相比，Ibβfruct2-2和Ibβfruct2-3编码的蛋白质序列分别检测到两个和六个氨基酸残基的变化；尽管这些变化不在关键基序内，但它们影响了蛋白质结构和对催化底物的亲和力，导致功能缺陷和活性降低。在拟南芥中异源表达Ibβfruct2-1降低了叶片中的淀粉含量，但增加了葡萄糖含量，表明Ibβfruct2-1是淀粉含量的负调节因子。这些发现对于理解甘薯中重复基因的调控和功能分化具有重要意义，并为这些基因的功能研究及其在遗传改良中的应用提供了关键信息。