Primary cells derived from Tuberous Sclerosis Complex patients show autophagy alteration in the haploinsufficiency state.

Clévia Rosset,Larissa Brussa Reis,Eduardo Filippi-Chiela,Mariane Da Cunha Jaeger,Cristina Brinckmann Oliveira Netto,Ivaine Taís Sauthier Sartor,Caroline Brunetto De Farias,Patricia Ashton-Prolla,Rafael Roesler

doi:10.1590/1678-4685-gmb-2020-0475

Abstract

Tuberous sclerosis complex (TSC) is an autosomal dominant cancer predisposition disorder caused by heterozygous mutations in TSC1 or TSC2 genes and characterized by mTORC1 hyperactivation. TSC-associated tumors develop after loss of heterozygosity mutations and their treatment involves the use of mTORC1 inhibitors. We aimed to evaluate cellular processes regulated by mTORC1 in TSC cells with different mutations before tumor development. Flow cytometry analyses were performed to evaluate cell viability, cell cycle and autophagy in non-tumor primary TSC cells with different heterozygous mutations and in control cells without TSC mutations, before and after treatment with rapamycin (mTORC1 inhibitor). We did not observe differences in cell viability and cell cycle between the cell groups. However, autophagy was reduced in mutated cells. After rapamycin treatment, mutated cells showed a significant increase in the autophagy process (p=0.039). We did not observe differences between cells with distinct TSC mutations. Our main finding is the alteration of autophagy in non-tumor TSC cells. Previous studies in literature found autophagy alterations in tumor TSC cells or knock-out animal models. We showed that autophagy could be an important mechanism that leads to TSC tumor formation in the haploinsufficiency state. This result could guide future studies in this field.

Highlights

Tuberous sclerosis complex (TSC) is an autosomal dominant cancer predisposition disorder with a birth incidence estimated to be 1 in 6000, with an equal male/ female distribution (Osborne et al, 1991)
Cell cycle distribution and autophagy in heterozygous TSC1 or TSC2 mutated and wildtype fibroblasts, before and after treatment with rapamycin
We found for the first time that autophagy is already altered in heterozygous mutant TSC non-tumor cells and this alteration could be an initial mechanism to lead TSC tumor formation

Summary

Introduction

Tuberous sclerosis complex (TSC) is an autosomal dominant cancer predisposition disorder with a birth incidence estimated to be 1 in 6000, with an equal male/ female distribution (Osborne et al, 1991). It is caused by mutations in either of two genes, TSC1 or TSC2, which code for hamartin and tuberin, respectively (Povey et al, 1994). TSC is a complex disorder with many features, ranging from mild to severe symptoms It is characterized by neuropsychiatric disorders and multiple hamartomas, mainly in the brain, kidneys, heart, and lungs (Crino et al, 2006). In parallel to autophagy induction, the inactivation of mTORC1 inhibits cell growth (Balgi et al, 2009; Hung et al, 2012)

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