Interaction between hypoxia and mutations in SDH subunit genes associated with phaeochromocytoma/paragangliomas (#10)
Phaeochromocytomas (PC) and paragangliomas occur in the context of inherited syndromes in 30% of cases1. Of 11 loci that have been defined to date, 4 encode succinate dehydrogenase genes (SDHA, SDHB, SDHC, and SDHD, collectively SDHx). Mechanisms by which germline SDHx mutations stimulate tumour formation are poorly understood. Clinical data implicate an interaction with environmental hypoxia.2, 3, 4 This study investigated effects of hypoxia on cellular function in the presence or absence of SDHx mutation.
1. Wild type and mutant SDHx cDNAs tagged with GFP were expressed in HEK293 cells, and cultured in either normoxic or hypoxic conditions. Cellular localisation was visualised by confocal microscopy and confirmed by western blotting.
2. Mitochondrial function was studied in a novel model5 of SDHx dysfunction using neural stem cells obtained from patients with SDHx mutations.
3. The hypoxic-responsive miR2106 was previously measured by qPCR in PC/PGLs containing mutations in SDHB and non SDHB PC/PGLs. MiR210 was also measured in neurospheres.
Wild-type SDHx localised to mitochondriae under normoxic conditions, but less so under hypoxia. In contrast, some SDHx mutants constitutively failed to localise to mitochondriae whereas other mutants retained mitochondrial localisation in normoxic but not hypoxic conditions.
Neurosphere cells containing SDHB mutations exhibited reduced SDHB expression, reduced complex II function, and increased membrane potential.
MiR210 levels were increased in SDHB-mutated tumours compared with non-SDHB-mutated tumours or normal samples. MiR210 was increased in neurospheres containing SDHB mutations.
1. SDHx mutations impaired SDHx mitochondrial localisation, either constitutively or in conjunction with hypoxia.
2. The neurosphere model confirmed that some complex II dysfunction is evident with heterozygous SDHB mutation.
3. Elevated miR210 was evident in PC/PGLs associated with SDHB mutations, consistent with known pseudohypoxic gene signature of these tumours.
Our study suggests an interaction between SDHx mutation and hypoxia in the pathogenesis of PC/PGL.
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