Incrimination of Heterogeneous Nuclear Ribonucleoprotein E1 (hnRNP-E1) as a Candidate Sensor of Physiological Folate Deficiency

Abstract

The mechanism underlying the sensing of varying degrees of physiological folate deficiency, prior to adaptive optimization of cellular folate uptake through the translational up-regulation of folate receptors (FR) is unclear. Because homocysteine, which accumulates intracellularly during folate deficiency, stimulated interactions between heterogeneous nuclear ribonucleoprotein E1 (hnRNP-E1) and an 18-base FR-α mRNA cis-element that led to increased FR biosynthesis and net up-regulation of FR at cell surfaces, hnRNP-E1 was a plausible candidate sensor of folate deficiency. Accordingly, using purified components, we evaluated the physiological basis whereby L-homocysteine triggered these RNA-protein interactions to stimulate FR biosynthesis. L-homocysteine induced a concentration-dependent increase in RNA-protein binding affinity throughout the range of physiological folate deficiency, which correlated with a proportionate increase in translation of FR in vitro and in cultured human cells. Targeted reduction of newly synthesized hnRNP-E1 proteins by siRNA to hnRNP-E1 mRNA reduced both constitutive and L-homocysteine-induced rates of FR biosynthesis. Furthermore, L-homocysteine covalently bound hnRNP-E1 via multiple protein-cysteine-S-S-homocysteine mixed disulfide bonds within K-homology domains known to interact with mRNA. These data suggest that a concentration-dependent, sequential disruption of critical cysteine-S-S-cysteine bonds by covalently bound L-homocysteine progressively unmasks an underlying RNA-binding pocket in hnRNP-E1 to optimize interaction with FR-α mRNA cis-element preparatory to FR up-regulation. Collectively, such data incriminate hnRNP-E1 as a physiologically relevant, sensitive, cellular sensor of folate deficiency. Because diverse mammalian and viral mRNAs also interact with this RNA-binding domain with functional consequences to their protein expression, homocysteinylated hnRNP-E1 also appears well positioned to orchestrate a novel, nutrition-sensitive (homocysteine-responsive), posttranscriptional RNA operon in folate-deficient cells.