Analysis of organelle contact sites and their metabolic rewiring upon pharmacological treatment in breast cancer lines

Organelle contact sites are defined as the physical juxtaposition between their membranes occurring at 10-80nm. They are involved in different cellular pathways such as Ca2+ signaling, lipid metabolism, apoptosis, and autophagy. Alteration of such contacts have been observed in pathological conditions like cancer and neurodegeneration. In this workflow we have investigated a putative relationship between organelle contact sites and metabolic conditions of breast cancer cells. We have employed MCF10A, MCF7, MDAMB231 breast cancer cell lines which represent different stage of tumoral progression and show different metabolic states. Contact sites between mitochondria (MT) and endoplasmic reticulum (ER), or lysosomes (LY) or nucleus (NU) were revealed by SPLICSS recombinant probes in basal condition and upon treating the cells with metabolic inhibitors (2-deoxy-D-glucose, Etomoxir, UK5099): a rewiring of contact sites have been observed upon their administration. In non-treated cells, ER-MT contacts increase proportionally to the aggressiveness of the tumor stage represented by the different cell lines, while LY-MT contacts are different (i.e., more abundant) only between MCF7 and MDAMB231 cells. The metabolic rewiring induced by metabolic inhibitors cause different effects on contact sites in the three cell lines, indicating a possible link between the tuning of metabolic pathways, organelles communication and intracellular signaling in cancer development and progression.