The effect of knockdown and mutation of the ganglioside-induced differentiation-associated protein 1 (GDAP1) on mitochondrial shape and cell metabolism
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Abstract
Mutations in the gene encoding for the ganglioside-induced differentiation-associated protein 1 (GDAP1) cause the hereditary polyneuropathy Charcot-Marie-Tooth disease (CMT). Structurally, the GDAP1 protein possesses a transmembrane domain anchored in the outer mitochondrial membrane (OMM) and two domains facing the cytosol that share similarities to glutathione-transferases (GST) – enzymes catalyzing the conjugation of the antioxidant glutathione to electrophilic residues. GDAP1 is highly involved in mitochondrial dynamics, ongoing processes of fusion and fission. Changes in GDAP1 expression levels can disturb this balance and shift the mitochondrial shape to either fragmented mitochondria when overexpressed or fused mitochondria when silenced.
This work not only confirmed that perturbed GDAP1 expression leads to more mitochondrial elongation in GDAP1 knockdown (KD) cell lines but also in neuronal cells derived from autosomal-recessive CMT4A patients caused by a GDAP1 mutation.
We observed that GDAP1 KD cells and patient-derived motoneurons exhibited a significant reduction in cytosolic fatty acids and that both in vitro models shifted their mitochondrial metabolism towards increased glutaminolysis – most likely, a compensatory effect to an impaired pyruvate conversion to acetyl-CoA, a crucial Ca2+-dependent process controlling citrate production and entry into the TCA cycle. Though GDAP1 KD cells were still less efficient for mitochondrial ATP production.
By analyzing pulled down neuronal proteins by mass spectrometry, I identified the cytosolic actin-binding factor cofilin1 as a potential GDAP1-interacting protein. Cofilin1 regulates the depolymerization of actin filaments, the abundance of the intracellular signaling hubs mitochondria-ER contact sites (MERCS) and mitochondrial fragmentation mediated by the mitochondrial fission factor dynamin-related protein 1 (DRP1). Translocation of DRP1 to the OMM was indeed reduced and MERCS had an increased width in GDAP1 KD cells resulting in an impaired Ca2+ transfer upon stimulation of the inositol-1,4,5-tris-phosphate receptor (IP3R). An altered function of cofilin1 might therefore underlie the morphological and metabolic impairments observed in this work. Hence, this work shed light on metabolic alterations in GDAP1 KD but also in CMT4A patient-derived cells and contributed to further explanation of potential GDAP1 targets.