Regulatory Mechanisms of Folate Transporters by Transcription Factors in the Brain

叶酸(VitB9)是哺乳动物细胞关键生物合成过程中所需的水溶性维生素。叶酸跨生物膜和组织的转运主要有三条途径：叶酸受体α(FRA)、质子偶联叶酸转运体(PCFT)和还原叶酸载体(RFC)。这些叶酸转运途径的功能表达可以受到多种转录因子的调节，如维生素D受体(VDR)。叶酸对大脑发育和功能至关重要。FRA是血-脑脊液屏障的主要叶酸运输途径，通过功能丧失突变或自身抗体的存在而失活，导致以炎症、氧化应激、线粒体功能障碍和脑髓鞘异常为特征的严重脑叶酸缺乏症。最近，作为2015年4月批准的NSERC拨款的一部分，我们证明了通过诱导特定的叶酸转运体(即RFC)来调节血脑屏障(BBB)的叶酸摄取可以提供一种在FRA缺陷的情况下增强脑叶酸输送的新策略。我们提供的证据表明，通过暴露于VDR的激活配体1，25-二羟基维生素D3(骨化三醇)来激活VDR，可以诱导血脑屏障的RFC功能，并有效地恢复FRA缺乏小鼠大脑中的叶酸水平。尽管这些结果非常令人兴奋，但我们观察到，用骨化三醇治疗的动物出现了毒性，如体重减轻和高钙血液水平。NRF-1是一种转录因子，主要因其在促进线粒体和呼吸功能方面的作用而闻名。我们最近确定，它还可以调节血脑屏障中的叶酸转运蛋白RFC。我们认为，在大脑FRA缺陷的情况下，NRF-1被其特定的配体-吡咯喹啉醌(PQQ)激活，可以增强RFC功能，增加叶酸向大脑的输送，逆转炎症、氧化应激和线粒体功能障碍。我们拟议的工作将揭示与脑叶酸缺乏相关的线粒体功能障碍、脑部炎症和氧化应激的基本机制，阐明当大脑叶酸水平恢复时这些影响如何被逆转，并为FRA功能丧失引起的神经代谢障碍提供新的治疗方法。此外，由于越来越多的证据将线粒体功能障碍与神经疾病联系起来，该项目的发现提供了更广泛的应用，并可能被用作几种已知具有非常低的中枢神经系统通透性的药物(即抗癌药物)的一种新的给药途径。随着拟议工作的开展，我们计划培养和指导高素质的人员，使其成为未来自然科学的领导者。

Folates (Vit B9) are water soluble vitamins required for key biosynthetic processes in mammalian cells. Folate transport across biological membranes and tissues is mediated by three major pathways: folate receptor alpha (FRa), proton-coupled folate transporter (PCFT), and reduced folate carrier (RFC). The functional expression of these folate transport pathways can be regulated by several transcription factors such as the Vitamin D receptor (VDR). Folates are critical for brain development and function. Inactivation of FRa, the major folate transport pathway at the blood-cerebrospinal fluid barrier, through loss-of-function mutations or the presence of FRa autoantibodies, causes severe cerebral folate deficiency characterized by inflammation, oxidative stress, mitochondrial dysfunction and abnormal brain myelination. Recently, as part of the NSERC grant approved in April 2015, we demonstrated that modulating folate uptake at the blood-brain barrier (BBB) through induction of specific folate transporters (i.e., RFC) could present a novel strategy for enhancing brain folate delivery in the context of defective FRa. We provided evidence that activation of VDR through exposure to its activating ligand, 1,25-dihydroxyvitamin D3 (calcitriol), induced RFC function at the BBB and effectively restored folate levels in the brain of mice lacking FRa. Although these results are very exciting, we observed that the animals treated with calcitriol presented toxicities such as weight loss and high calcium blood levels. NRF-1 is a transcription factor primarily known for its role in promoting mitochondrial and respiratory function. We recently determined that it can also regulate the folate transporter, RFC at the BBB. We propose that, in the context of defective FRa in the brain, activation of NRF-1 by its specific ligand, pyrroloquinoline quinone (PQQ), a natural non-toxic compound, can enhance RFC function and increase folate delivery into the brain reversing inflammation, oxidative stress, as well as mitochondrial dysfunction. Our proposed work will uncover fundamental mechanisms involved in mitochondrial dysfunction, brain inflammation and oxidative stress related to cerebral folate deficiency, elucidate how these effects can be reversed when brain folate levels are restored, and provide novel treatment approaches for neurometabolic disorders caused by loss of FRa function. Further, the findings of this project present broader applications due to the growing body of evidence associating mitochondrial dysfunction to neurological disorders and may also be exploited as a novel delivery route for several pharmacological agents known to have very poor CNS permeability (i.e., anticancer drugs). With the undertaking of the proposed work, we plan to train and mentor highly qualified personnel to become future leaders in the natural sciences.