TFE3 fusions escape from controlling of mTOR signaling pathway and accumulate in the nucleus promoting genes expression in Xp11.2 translocation renal cell carcinomas

Abstract

Background Xp11.2 translocation renal cell carcinoma (tRCC) is mainly caused by translocation of the TFE3 gene located on chromosome Xp11.2 and is characterized by overexpression of the TFE3 fusion gene. Patients are diagnosed with tRCC usually before 45 years of age with poor prognosis. We investigated this disease using two tRCC cell lines, UOK109 and UOK120, in this study. Methods The purpose of this study was to investigate the pathogenic mechanism of TFE3 fusions in tRCC based on its subcellular localization, nuclear translocation and transcriptional activity. The expression of TFE3 fusions and other related genes were analyzed by quantitative reverse transcription PCR (qRT-PCR) and Western blot. The subcellular localization of TFE3 was determined using immunofluorescence. The transcriptional activity of TFE3 fusions was measured using a luciferase reporter assay and ChIP analysis. In some experiments, TFE3 fusions were depleted by RNAi or gene knockdown. The TFE3 fusion segments were cloned into a plasmid expression system for expression in cells. Results Our results demonstrated that TFE3 fusions were overexpressed in tRCC with a strong nuclear retention irrespective of treatment with an mTORC1 inhibitor or not. TFE3 fusions lost its co-localization with lysosomal proteins and decreased its interaction with the chaperone 14–3-3 proteins in UOK109 and UOK120 cells. However, the fusion segments of TFE3 could not translocate to the nucleus and inhibition of Gsk3β could increase the cytoplasmic retention of TFE3 fusions. Both the luciferase reporter assay and ChIP analysis demonstrated that TFE3 fusions could bind to the promoters of the target genes as a wild-type TFE3 protein. Knockdown of TFE3 results in decreased expression of those genes responsible for lysosomal biogenesis and other target genes. The ChIP-seq data further verified that, in addition to lysosomal genes, TFE3 fusions could regulate genes involved in cellular responses to hypoxic stress and transcription. Conclusions Our results indicated that the overexpressed TFE3 fusions were capable of escaping from the control by the mTOR signaling pathway and were accumulated in the nucleus in UOK109 and UOK120 cells. The nuclear retention of TFE3 fusions promoted the expression of lysosomal genes and other target genes, facilitating cancer cell resistance against an extreme environment. Electronic supplementary material The online version of this article (10.1186/s13046-019-1101-7) contains supplementary material, which is available to authorized users.

A renal cell carcinoma with EWSR1‐TFE3 fusion gene

“…Consistently, all TFE3 fusion partner genes including EWSR1 are ubiquitously expressed with constitutive active promoters. In addition, wild type TFE3 is mostly cytoplasmic by its interaction with 14‐3‐3; however, as seen in this case, rearranged TFE3 is translocated to the nucleus (Figure C) . The 14‐3‐3 binding site of TFE3 includes Ser321, which can be phosphorylated by mTORC1.…”

“…In addition, wild type TFE3 is mostly cytoplasmic by its interaction with 14-3-3; however, as seen in this case, rearranged TFE3 is translocated to the nucleus ( Figure 1C). 21 The 14-3-3 binding site of TFE3 includes Ser321, which can be phosphorylated by mTORC1. Both RCC cases of EWSR1-TFE3 fusion retain the TFE3 Ser321.…”

TFE3 activation in a TSC1‐altered malignant PEComa: challenging the dichotomy of the underlying pathogenic mechanisms

“…Transcriptome analysis detected an increase in levels of TFE3 mRNA in tumor cells compared to dermal fibroblast controls after normalization for expression of housekeeping genes (see supplementary material, Figure S1). Therefore, similar to Xp11 translocation cancers, TFE3 overexpression is one likely oncogenic driver in this tumor [15].…”

Sunitinib treatment promotes metastasis of drug-resistant renal cell carcinoma via TFE3 signaling pathway

“…It has been demonstrated that 3-kinase (PI3K) is induced by VEGF/VEGFR2 can phosphorylate phosphoinositide 3-kinase (PI3K) and AKT, both of which augment mammalian target of rapamycin (mTOR) activation. mTOR complex 1 (mTORC1) can respond to the nutrients level change by regulating the intracellular distribution of TFE3 20 , 21 , 36 . In fed condition, TFE3 at serine 321 (Ser321) residue of TFE3 is phosphorylated by phosphorylated mTOR as a result of recruiting cytosolic chaperone 14-3-3 to bind to TFE3.…”