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 redoxmechanisms, 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.
|Effective start/end date||1/1/16 → 12/31/20|
- National Institutes of Health (NIH)