The asymmetric cleavage of beta-carotene in mammalian embryonic development

哺乳动物胚胎发育中β-胡萝卜素的不对称裂解

基本信息

批准号：
9181429
负责人：
Loredana Quadro
金额：
$ 31.57万
依托单位：
RUTGERS, THE STATE UNIV OF N.J.
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-01-01 至 2020-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9181429
关键词：
Address Affect Alcohols Aldehydes All-Trans-Retinol Beta Carotene Binding Binding Sites C10 Carotenoids Cell Line Cell Respiration Cell Survival Child Child Development Citric Acid Cycle Cleaved cell Congenital Abnormality Data Developing Countries Development Diet Dose Embryo Embryonic Development Enzymes Experimental Animal Model Experimental Models Female Female of child bearing age Fetal Death Fetus Fibroblasts Genes Genetic Transcription Geographic Locations Growth and Development function Health Homeostasis Human In Vitro Intake Intervention Knock-out Knockout Mice Ligands Link Malnutrition Metabolism Mitochondria Modeling Morphology Mothers Mouse Protein Mus Mutant Strains Mice Mutation Nutrient Nutritional status Output Oxidation-Reduction Oxidative Phosphorylation Oxidative Stress Oxides Oxygenases Pathway interactions Phenotype Phosphotransferases Physiology Play Population Pregnancy Pregnant Women Production Protein Isoforms Proteins Public Health Pyruvate Dehydrogenase Complex Reaction Reagent Regulation Respiration Retinaldehyde Retinoids Retinol Binding Proteins Role Signal Transduction Source Supplementation Testing Tissues Toxic effect Tretinoin Vitamin A Vitamin A Deficiency Vitamin B Complex Woman Work World Health Organization apocarotenal base in vivo innovation mouse model novel prevent programs public health relevance

项目摘要

DESCRIPTION (provided by applicant): Vitamin A is required for proper mammalian embryonic development, owing to defective transcriptional action of retinoic acid (RA), its active form. Deficient or excessive maternal vitamin A intake of this essential nutrient results in congenital abnormalities or fetal death in experimental animal models and humans. Notably, vitamin A deficiency (VAD) is the third most prevalent nutritional deficiency, and is an overwhelming public health issue, affecting hundreds of millions of people (predominantly women and children) in developing countries. The majority of the world population, especially in the above-mentioned geographic areas, relies on the vitamin A precursor β-carotene (BC) as a source of retinoids (vitamin A and its derivatives). In the embryo, BC obtained from the maternal diet can significantly contribute to the retinoid needs of the developing tissues upon conversion to retinaldehyde via the symmetric BC cleavage enzyme β,β-carotene-15,15'-oxygenase, CMO1. Retinaldehyde is then oxidized to retinoic acid, the master regulator of many genes that are crucial to embryogenesis. BC is also cleaved asymmetrically by β,β-carotene-9',10'-oxygenase, CMO2, generating C10 β-apocarotenal (C10 apoAL). While retinaldehyde could also be generated from C10 apoAL, the role of CMO2 and its reaction product during mammalian embryonic development is unknown. Our preliminary data indicate that the well-known detrimental effects of VAD on mouse embryogenesis are aggravated when CMO2 is inactive and BC is administered to the dams, despite expression of CMO1. We showed that the embryonic phenotype of mice lacking CMO2 on the retinol-binding protein (RBP) knockout background, an established model of VAD, was due to the low levels of C10 apoAL along with limited availability of retinoids. Supplementing CMO2-/-RBP-/- mice on a vitamin A deficient diet with C10 apoAL reduced congenital malformations. We propose that C10 apoAL serves as a ligand for PKCδ. This mitochondria-localized PKC isoform signals to the pyruvate dehydrogenase complex (PDHC), increasing its activity, with the purpose of coordinating the fuel flux to the citric acid cycle with the demands for ATP production. PKCδ is activated by redox mechanisms, with a mandatory catalytic role of vitamin A (retinol) that binds the activation domains of the kinase. CMO2 localizes to mitochondria and C10 aopAL is structurally similar to retinol. We showed that C10 apoAL interacts with the retinol-binding domain of PKCδ and modulates respiration in mouse embryonic fibroblasts, in a PKCδ-dependent manner. Overall, our data suggest that disruption of PKC signaling and mitochondrial functions when C10 apoAL and vitamin A are limiting could be the underlying cause of the exacerbated phenotypes of mice lacking CMO2 in the presence of BC. With this application we seek to further understand the function of CMO2 and the mechanisms of C10 apoAL action during mammalian embryogenesis. Specifically, we will test whether C10 apoAL is essential for embryonic survival under conditions of VAD (Aim 1) and we will define the interaction between CMO2/β- apocarotenoids and PKCδ signaling network during embryonic development (Aim 2), both in vitro and ex vivo (Aim 2A) and in vivo (Aim 2B). Unique reagents, such as pure synthetic β-apocarotenoid compounds, and new mouse models, with inactivation of the PKCδ and carotenoid metabolism pathways, will be used to address the above-mentioned questions. Understanding the role of CMO2/β-apocarotenoids in embryogenesis is relevant to human health, as it will lead to novel interventions to ameliorate VAD-associated congenital defects.

描述（由申请人提供）：由于视黄酸（RA）及其活性形式的转录作用有缺陷，因此维生素 A 是哺乳动物胚胎正常发育所必需的，母体维生素 A 摄入不足或过多会导致先天性畸形或胎儿。值得注意的是，维生素 A 缺乏症 (VAD) 是第三大最普遍的营养缺乏症，也是一个压倒性的公共卫生问题，影响着数亿处于发育阶段的人（主要是妇女和儿童）。世界上大多数人口，尤其是上述地理区域的人口，依赖维生素 A 前体 β-胡萝卜素 (BC) 作为类维生素A（维生素 A 及其衍生物）的来源。在胚胎中，BC 是从胚胎中获得的。母体饮食可通过对称 BC 裂解酶 β,β-胡萝卜素-15,15'-加氧酶 (CMO1) 转化为视黄醛，从而显着满足发育组织对类视黄醇的需求。然后视黄醛被氧化为视黄酸，视黄酸是许多对胚胎发生至关重要的基因的主要调节因子，BC 也会被 β,β-胡萝卜素-9',10'-加氧酶、CMO2、C10 β-脱胡萝卜素 (C10 apoAL) 不对称地裂解。虽然视黄醛也可以由 C10 apoAL 生成，但 CMO2 及其反应产物在哺乳动物过程中的作用。我们的初步数据表明，尽管 CMO1 表达，但当 CMO2 失活且给母鼠施用 BC 时，VAD 对小鼠胚胎发生的众所周知的不良影响会加剧。视黄醇结合蛋白 (RBP) 敲除背景是一种已建立的 VAD 模型，其原因是 C10 apoAL 水平较低以及类视黄醇补充剂的可用性有限。缺乏维生素 A 且含有 C10 apoAL 的 CMO2-/-RBP-/- 小鼠可减少先天性畸形，我们认为 C10 apoAL 作为 PKCδ 的配体，向丙酮酸脱氢酶复合体 (PDHC) 发出信号。增加其活性，目的是协调柠檬酸循环的燃料通量与 ATP 生成的需求。 PKCδ 被氧化还原激活。机制，维生素 A（视黄醇）结合位于线粒体的 CMO2 激活域，并且 C10 aopAL 在结构上与视黄醇相似。我们发现 C10 apoAL 与 PKCδ 的视黄醇结合域相互作用。以 PKCδ 依赖性方式调节小鼠胚胎成纤维细胞的呼吸，我们的数据表明，当 PKC 信号传导和线粒体功能受到破坏时。 C10 apoAL 和维生素 A 的限制可能是 BC 存在时缺乏 CMO2 的小鼠表型恶化的根本原因。通过该应用，我们试图进一步了解 CMO2 的功能和 C10 apoAL 在哺乳动物胚胎发生过程中的作用机制。我们将测试 C10 apoAL 是否对 VAD 条件下的胚胎存活至关重要（目标 1），并且我们将定义 CMO2/β-类胡萝卜素和 PKCδ 信号传导之间的相互作用胚胎发育过程中的网络（目标 2），体外和离体（目标 2A）和体内（目标 2B）独特的试剂，例如纯合成 β-类胡萝卜素化合物，以及新的小鼠模型，具有 PKCδ 和 PKCδ 失活功能。类胡萝卜素代谢途径，将用于解决上述问题，了解 CMO2/β-类胡萝卜素在胚胎发生中的作用与人类健康相关。导致改善 VAD 相关先天性缺陷的新干预措施。