Seim, G. | Search bioRxiv

bioRxiv2023

Reactive nitrogen species inhibit branched chain alpha-ketoacid dehydrogenasecomplex and impact muscle cell metabolism

Branched chain -ketoacid dehydrogenase complex (BCKDC) is the rate limiting enzyme in branched chain amino acid (BCAA) catabolism, a metabolic pathway with great importance for human health. BCKDC belongs to the mitochondrial -ketoacid dehydrogenase complex family, which also includes pyruvate dehydrogenase complex (PDHC) and oxoglutarate dehydrogenase complex (OGDC). Here we revealed that BCKDC can be substantially inhibited by reactive nitrogen species (RNS) via a mechanism similar to what we recently discovered with PDHC and OGDC -- modifying the lipoic arm on its E2 subunit. In addition, we showed that such reaction between RNS and the lipoic arm of the E2 subunit can further promote inhibition of the E3 subunits of -ketoacid dehydrogenase complexes. We examined the impacts of this RNS-mediated BCKDC inhibition in muscle cells, an important site of BCAA metabolism, and demonstrated that the nitric oxide production induced by cytokine stimulation leads to a strong inhibition of BCKDC activity and BCAA oxidation in myotubes and myoblasts. More broadly, nitric oxide production reduced the level of functional lipoic arms across the multiple -ketoacid dehydrogenases and led to intracellular accumulation of their substrates (-ketoacids), reduction of their products (acyl-CoAs), and a lower cellular energy charge. This work revealed a new mechanism for BCKDC regulation, demonstrated its biological significance, and elucidated the mechanistic connection between RNS-driven inhibitory modifications on the E2 and E3 subunits of -ketoacid dehydrogenases. Together with previous work, we revealed a general mechanism for RNS to inhibit all -ketoacid dehydrogenases, which has numerous physiological implications across multiple cell types.

biochemistry↗

bioRxiv2022

Asparagine starvation suppresses histone demethylation through iron depletion

Nutrient availability can impact epigenome to modify gene expression and dictate cell fate decision (Etchegaray and Mostoslavsky, 2016; Kinnaird et al., 2016). -ketoglutarate is an indispensable substrate for the Jumonji family of histone demethylases (JHDMs) mediating most of the cellular demethylation reactions on histone subunits (Schvartzman et al., 2018). Since -ketoglutarate is an intermediate of the tricarboxylic acid (TCA) cycle and a product of transamination, its intracellular levels are regulated by the metabolism of several amino acids (Baksh et al., 2020; Carey et al., 2015; Raffel et al., 2017; Vardhana et al., 2019). Here we show that asparagine starvation suppresses global histone demethylation. This process is neither due to the change of expression of histone modifying enzymes, nor due to the change of intracellular level of -ketoglutarate. Rather, asparagine starvation reduces intracellular pool of labile iron (Fe2+), which is a key cofactor for the JHDMs to function. Mechanistically, asparagine starvation post-transcriptionally suppresses the expression of iron responsive element binding protein 2 (IREB2), an iron sensing protein, which then reduces the mRNA expression of the transferrin receptor (TFRC), a major carrier for iron uptake (Hentze et al., 2010). Furthermore, iron supplementation to the culture medium restores histone demethylation and alters global gene expression to accelerate cell death under conditions of asparagine starvation. Collectively, our results uncover that suppression of iron-dependent histone demethylation is part of the cellular adaptive response to asparagine starvation.

cell biology↗