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Additional file 1.

Charcot-Marie-Tooth (CMT) disease 4A is an autosomal-recessive polyneuropathy caused by mutations of ganglioside-induced differentiation-associated protein 1 (GDAP1), a putative glutathione transferase, which affects mitochondrial shape and alters cellular Ca2+ homeostasis. Here, we identify the underlying mechanism. We found that patient-derived motoneurons and GDAP1 knockdown SH-SY5Y cells display two phenotypes: more tubular mitochondria and a metabolism characterized by glutamine dependence and fewer cytosolic lipid droplets. GDAP1 interacts with the actin-depolymerizing protein Cofilin-1 in a redoxdependent manner, suggesting a role for actin signaling. Consistently, GDAP1 loss causes less F-actin close to mitochondria, which restricts mitochondrial localization of the fission factor dynamin-related protein 1, instigating tubularity. Changes in the actin cytoskeleton also disrupt mitochondria-ER contact sites. This results in lower mitochondrial Ca2+ levels and inhibition of the pyruvate dehydrogenase complex, explaining the metabolic changes upon GDAP1 loss of function. Together, these findings reconcile GDAP1-associated phenotypes and implicate disrupted actin signaling in CMT4A pathophysiology.

Cisplatin is one of the most potent anticancer agents used in the treatment of various solid tumors. Unfortunately the onset of resistance is the main limit of this therapy and severely compromises the treatment effectiveness. Although several studies regarding cisplatin resistance have been performed, the molecular mechanisms are not completely understood. Classically, cisplatin is studied as a DNA-damaging chemotherapy agent, but more recent investigations showed that only 5-10% of intracellular platinum is bound to nuclear DNA, while the great majority of the intracellular drug can interact with a variety of cellular component including proteins, RNA and mitochondrial DNA. MtDNA, unlike nDNA, does not possess efficient repair systems; therefore it is more susceptible to the onset of mutations often associated to cancer development, loss of tumor suppressor, activation of oncogenes and mitochondrial dysfunctions related with an increase of glycolytic activity. The Warburg effect indicates the alteration of energetic metabolism used by tumor cells as a strategy to adapt and grow independently from the substrate availability. This evidence suggested us to verify the hypothesis that a similar metabolic strategy might be of relevance in cisplatin resistance. Therefore, our aim was to investigate the energetic metabolism and the mitochondrial dynamic of cisplatin-resistant and sensitive cancer cells with different experimental approaches, in order to reveal targets useful to overcome the resistance. In our laboratory we have already revealed that cisplatin resistant ovarian cancer cell line C13, as compared to sensitive line 2008, exhibits metabolic changes. Indeed, resistant clone showed a different mitochondrial and metabolic profile characterized by an increase of glucose and glutamine uptake, a decrease of the mitochondrial membrane potential and mitochondrial mass. In this scenario, we proceeded to phenotype other cancer cells that present acquired or intrinsic resistance in order to identify new targets to sensitize to cisplatin treatment. Our results pointed out alterations in mitochondrial fusion and fission in chemoresistant cancer cells. Moreover, data obtained showed that resistant clones, with an imbalance toward fission process, present a faster mitochondrial turn-over using mitophagy as a mitochondrial quality control mechanism. Furthermore, the data showed a mitochondrial network differently organized in resistant variants underlining a probable implication of dynamic process in resistance mechanisms. Having regard to the data about metabolic reprogramming, breast cancer cells that have an innate resistance to cisplatin were evaluated. The expression of c-Myc nuclear transcription factor, involved in the metabolic reprogramming of tumor cells, has been evaluated highlighting a different expression of some of its target genes involved in the glycolylisis and glutaminolysis, besides an increased dependency of glucose in cisplatin resistant cells.

Cisplatin (CDDP) is commonly used to treat a multitude of tumors including sarcomas, ovarian and cervical cancers. Despite recent investigations allowed to improve chemotherapy effectiveness, the molecular mechanisms underlying the development of CDDP resistance remain a major goal in cancer research. Here, we show that mitochondrial morphology and autophagy are altered in different CDDP resistant cancer cell lines. In CDDP resistant osteosarcoma and ovarian carcinoma, mitochondria are fragmented and closely juxtaposed to the endoplasmic reticulum; rates of mitophagy are also increased. Specifically, levels of the mitophagy receptor BNIP3 are higher both in resistant cells and in ovarian cancer patient samples resistant to platinum-based treatments. Genetic BNIP3 silencing or pharmacological inhibition of autophagosome formation re-sensitizes these cells to CDDP. Our study identifies inhibition of BNIP3-driven mitophagy as a potential therapeutic strategy to counteract CDDP resistance in ovarian carcinoma and osteosarcoma.