Opposing effects of HIF1α and HIF2α on chromaffin cell phenotypic features and tumor cell proliferation: Insights from MYC‐associated factor X

Abstract

Pheochromocytomas and paragangliomas (PPGLs) are catecholamine-producing chromaffin cell tumors with diverse phenotypic features reflecting mutations in numerous genes, including MYC-associated factor X (MAX). To explore whether phenotypic differences among PPGLs reflect a MAX-mediated mechanism and opposing influences of hypoxia-inducible factor (HIF)s HIF2a and HIF1a, we combined observational investigations in PPGLs and gene-manipulation studies in two pheochromocytoma cell lines. Among PPGLs from 140 patients, tumors due to MAX mutations were characterized by gene expression profiles and intermediate phenotypic features that distinguished these tumors from other PPGLs, all of which fell into two expression clusters: one cluster with low expression of HIF2a and mature phenotypic features and the other with high expression of HIF2a and immature phenotypic features due to mutations stabilizing HIFs. Max-mutated tumors distributed to a distinct subcluster of the former group. In cell lines lacking Max, re-expression of the gene resulted in maturation of phenotypic features and decreased cell cycle progression. In cell lines lacking Hif2a, overexpression of the gene led to immature phenotypic features, failure of dexamethasone to induce differentiation and increased proliferation. HIF1a had opposing actions to HIF2a in both cell lines, supporting evolving evidence of their differential actions on tumorigenic processes via a MYC/MAX-related pathway. Requirement of a fully functional MYC/MAX complex to facilitate differentiation explains the intermediate phenotypic features in tumors due to MAX mutations. Overexpression of HIF2a in chromaffin cell tumors due to mutations affecting HIF stabilization explains their proliferative features and why the tumors fail to differentiate even when exposed locally to adrenal steroids.

DNA Methylation Profiling in Pheochromocytoma and Paraganglioma Reveals Diagnostic and Prognostic Markers

“…Our results verified PNMT hypermethylation previously reported in SDHBrelated PPGLs (2). PNMT hypomethylation in RET-and NF1-and hemimethylation in VHL-related PPGLs is highly consistent their associated neurochemical phenotypes (2,20,29). However, this model does not explain why other "cluster 1" tumors do not display an adrenergic phenotype, even when PNMT is hemimethylated.…”

“…Also, this does not explain why MAX-associated tumors, showing PNMT hypomethylation comparable to RET and NF1 tumors, do not display a predominantly adrenergic phenotype. In this context, Qin and colleagues (2014) provided evidence that addressed these paradoxes in other "cluster 1" tumors characterized by both EPAS1 expression and stabilization of HIF protein (20). In that study, the authors showed that EPAS1 completely blocked effects of steroid-induced PNMT expression in PC12 cells (29).…”

“…As previously described, downloaded gene expression profiles were quantile normalized and antigenomic probes were used as background probes (3,19,20).…”

A Defined, Controlled Culture System for Primary Bovine Chromaffin Progenitors Reveals Novel Biomarkers and Modulators

“…Specifically, it reduced the ratio of epinephrine to norepinephrine and increased the percentage of dopamine relative to total catecholamines. High dopamine levels in adrenomedullary tumors associate with a more aggressive phenotype, suggesting the presence of immature cancer cells [23][24][25]. On the other hand, the increase in relative dopamine content can be interpreted as a result of enhanced differentiation/ maturation of dopamine-producing TH+ cells.…”

Endocrine targets of hypoxia-inducible factors

“…Overexpression of HIF-2α is relatively common in VHL deficiency-related pheochromocytoma, whereas HIF-1α is more frequent in succinate dehydrogenase-deficient pheochromocytoma (Pollard et al 2006). Interestingly, it was reported that HIF-1α and HIF-2α exert opposite effect in pheochromocytoma, suggesting the signaling pathways may be similar but the downstream effects are divergent (Qin et al 2014). The distinct effects of HIF-1α and HIF-2α were also observed in adrenal neuroblastoma.…”