Cysteine Dependence of Insulin-like Growth Factor Effects on Redox Status

Abstract

Cobalamin is an essential co-factor for methionine synthase (MS), which catalyzes the conversion of homocysteine to methionine, however, oxidization of cobalamin halts MS activity. In cultured neuronal cells, reactivation of MS requires the antioxidant glutathione (GSH) to produce the active cofactor methylcobalamin (MeCbl). Hydroxocobalamin is rapidly and irreversibly converted to glutathionylcobalamin (GSCbl) in the presence of glutathione, then to MeCbl in the presence of S-adenosylmethionine. Cysteine is the rate-limiting precursor for GSH synthesis, and in neurons uptake via excitatory amino acid transporter 3 (EAAT3) provides the predominant source of cysteine. Insulin growth factor 1 (IGF-1) increases cellular cysteine uptake by promoting cellular surface expression of EAAT3. We investigated the influence of IGF-1 on cellular redox status and cobalamin composition in SH-SY5Y human neuroblastoma cells. HPLC separation and electrochemical detection were used to measure six intracellular cobalamin species, as well as the levels of GSH and its oxidized GSSG form. We found that GSCbl is a prominent intracellular cobalamin species in SH-SY5Y cells. IGF-1 treatment shifted the cellular environment to a more reduced state by increasing intracellular levels of GSH and the GSH/GSSG ratio, leading to an increased level of GSCbl. This novel action of IGF-1 allows it to regulate cellular redox status.