Nuove frontiere della ricerca: definire il ruolo di Tau/MAPT nella progressione tumorale ​

Tau protein, also known as MAPT, is a major member of microtubules associated protein (MAP) family. The immature primary transcript coded by MAPT gene can undergo alternative splicing leading to the generation of multiple Tau isoforms. The C-terminal region of the protein named “microtubule assembly domain” is essential for binding microtubules and promoting their assembly. Therefore, post-translational modifications (PTMs) occurring at this level may be responsible for MAPT binding dysfunction. Among all the potential PTMs involved in Tau proteostasis, phosphorylation has been extensively studied. In fact, MAPT hyperphosphorylation has been suggested responsible for neurofibrils generation at axonal level in accumulation diseases called tauopathies, including Alzheimer’s disease. Although tauopathies affect post-mitotic cells, Tau protein can also modulate cell cycle progression due to its role as a ligand of the mitotic spindle. Therefore, Tau dysfunction should be re-interpreted also in the light of the effects it could exert to the modulation of cell cycle in tumor cells. Indeed, as reported by “The Human Protein Atlas”, different tumors can express MAPT at high levels, including neuroblastoma (NB), a pediatric extra-cranial solid tumor. This preliminary study aimed to evaluate whether modulation of Tau protein phosphorylation and localization can influence tumor cell differentiation towards a neuronal-like phenotype, clarifying its potential role as a therapeutic target in combination with a differentiation treatment. NB CHP-126 and SH-SY5Y cell lines were used as reference tumor models and retinoic acid (RA) as differentiation treatment. A 9-day RA-based differentiation protocol was established to promote tumor cell proliferation arrest and differentiation. Enzymatic inhibition acting on Tau phosphorylation/dephosphorylation status was evaluated to understand the potential association with RA-induced antiproliferative effect. In addition, RA washout was considered as a model mimicking an acquired resistance to the therapy. Cell cultures, Western Blot, flow cytometry and immunofluorescence were the main experimental methodologies performed. Tau expression in NB cells occurs as two main monomeric isoforms. MAPT hyperphosphorylation was confirmed to hugely increase during G2/M phase suggesting an association between microtubules and Tau phosphorylation in potentially modulating mitotic spindle formation. Following the differentiation protocol, total Tau was detected to polarize along neo-forming neurites induced by RA-based differentiation, whereas phosphorylated Tau (p-Tau231) persisted mainly at cytoplasmatic level. Phosphorylation inhibition due to the treatment with 1-azakenpaullone seems to further enhance the differentiation process, however more analyses are required to strengthen this evidence. RA washout at the end of differentiation protocol revealed the occurrence of peculiarly new NB cell morphologies, associated with the recovery of tumor growth and, interestingly, associated also with an evident downmodulation in phosphorylation of one of the two main Tau isoforms. This study has allowed to highlight a preliminary evidence about the peculiar behavior of Tau proteostasis during RA differentiation of NB cells. Further experimental investigation will permit to understand if modulation of Tau phosphorylation could represent a useful adjuvant strategy in NB differentiation therapy.