Abstract
Phenylketonuria (PKU) is an autosomal recessive metabolic disorder caused by the dysfunction of the enzyme phenylalanine hydroxylase (PAH). Alterations in the level of PAH leads to the toxic accumulation of phenylalanine in the blood and brain. Protein degradation mediated by ubiquitination is a principal cellular process for maintaining protein homeostasis. Therefore, it is important to identify the E3 ligases responsible for PAH turnover and proteostasis. Here, we report that anaphase-promoting complex/cyclosome-Cdh1 (APC/C)Cdh1 is an E3 ubiquitin ligase complex that interacts and promotes the polyubiquitination of PAH through the 26S proteasomal pathway. Cdh1 destabilizes and declines the half-life of PAH. In contrast, the CRISPR/Cas9-mediated knockout of Cdh1 stabilizes PAH expression and enhances phenylalanine metabolism. Additionally, our current study demonstrates the clinical relevance of PAH and Cdh1 correlation in hepatocellular carcinoma (HCC). Overall, we show that PAH is a prognostic marker for HCC and Cdh1 could be a potential therapeutic target to regulate PAH-mediated physiological and metabolic disorders.
Highlights
Phenylalanine hydroxylase (PAH) is a highly regulated liver enzyme that plays a key role in phenylalanine catabolism
Mammalian PAH is tetrameric, and its 52 kDa subunits are composed of n-terminal regulatory domain for allosteric activation by Phenylalanine (Phe), a central catalytic domain, and c-terminal helix responsible for tetramer formation [1,2] PAH catalyzes the hydroxylation of l-phenylalanine (l-Phe) into l-tyrosine (l-Tyr) using tetrahydrobiopterin (BH4) and molecular oxygen [3,4], which is the initial step in governing the synthesis of catecholamines, thyroxin, and melatonin [5]
Understanding the importance of PAH protein turnover in maintaining a stable pool of l-Phe and l-Tyr at physiological condition, we investigated whether PAH undergoes degradation via the 26S proteasomal pathway
Summary
Phenylalanine hydroxylase (PAH) is a highly regulated liver enzyme that plays a key role in phenylalanine catabolism. Mutations or deficiency in the PAH gene leads to hyper-accumulation of l-Phe, causing phenylketonuria (PKU) in different ethnic populations and the depletion of precursors involved in neurotransmitter biosynthesis in the central nervous system that overall results in loss of cognitive ability and neurological disorders [7]. The expression profiles of most of the liver enriched genes in hepatocellular carcinoma (HCC) showed downregulation behavior in tumor tissues than that of non-tumor tissues. High expression of these liver enriched genes showed prolonged overall survival suggesting favorable prognostic effects in HCC [9]. In addition to the covalent and non-covalent regulation of kinetic properties, the amount of available enzyme in cells plays a vital role [10]
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