Twenty novel mutations in nicotinamide nucleotide transhydrogenase (NNT) causing FGD

Lou Metherell; Eirini Meimaridou; Julia Kowalczyk; Leo Guasti; Claire Hughes; Nicholas Mann; Ritwik Banerjee; Peter King; Adrian Clark

OC1.6

Prev Next

Section Contents Cite

Endocrine Abstracts (2011) 27 OC1.6

BSPED2011 Oral Communications Oral Communications 1 (8 abstracts)

Twenty novel mutations in nicotinamide nucleotide transhydrogenase (NNT) causing FGD

Lou Metherell ¹ , Eirini Meimaridou ¹ , Julia Kowalczyk ¹ , Leo Guasti ¹ , Claire Hughes ¹ , Nicholas Mann ² , Ritwik Banerjee ³ , Peter King ¹ & Adrian Clark ¹

Err

Author affiliations

¹William Harvey Research Institute, QMUL, London, UK; ²Royal Berkshire Hospital, Reading, UK; ³Luton and Dunstable Hospital, Luton, UK.

Familial glucocorticoid deficiency (FGD;OMIM 202200) results from the inability of the adrenal cortex to produce cortisol in response to ACTH stimulation. Half of all cases are caused by mutations in MC2R, MRAP or STAR. SNP array genotyping of FGD patients of unknown aetiology mapped a disease locus to chromosome 5p13-q12. Targeted exome sequencing of 5p13-q12 in one patient identified a homozygous mutation, p.Ala533Val, in nicotinamide nucleotide transhydrogenase (NNT), a protein involved in antioxidant defence. Conventional sequencing of 100 FGD patients identified 20 further homozygous or compound heterozygous mutations in 14 other families.

C57BL6/J mice (a natural nnt mutant) had slightly hyperplastic, disorganized zonae fasciculata with higher levels of apoptosis than wild-type mice. These mutant mice also had lower basal and stimulated levels of corticosterone than their wild-type counterparts. Knockdown of NNT by shRNA in H295R cells increased reactive oxygen species (ROS) and reduced cortisol production. RT-PCR of a tissue panel revealed NNT is highly expressed in the human adrenal.

NNT encodes an integral protein of the inner mitochondrial membrane. Under most physiological conditions, this enzyme uses energy from the mitochondrial proton gradient to produce high concentrations of NADPH. The resulting NADPH is used for biosynthesis and in ROS detoxification by enzymes such as glutathione peroxidases. Previous studies have shown that ROS-mediated disruption of Leydig cell mitochondria inhibits STAR function and Leydig cell steroidogenesis.

Cellular defences against ROS such as O₂⁻ and H₂O₂ are highly developed and species have evolved several overlapping pathways to deal with it. Taken together our findings that NNT is highly expressed in the adrenal, that its knockdown/out leads to increased ROS and that mutations in the gene result in FGD suggest that NNT is a critical enzyme for ROS detoxification in adrenocortical cells, with its loss leading to defective oxidative stress responses, an impairment of steroidogenesis and hence adrenal insensitivity to ACTH.