Endocrine Abstracts

Mitochondrial steroidogenic cytochrome P450 (CYP) enzymes, such as P450 side-chain cleavage, rely on electron transfer from the redox partner ferredoxin (FDX1) for catalytic activity. Previous in vitro data suggest these cofactors are key regulators of CYP enzyme activity. This study aims to establish the role of redox regulation on steroidogenesis using zebrafish as a vertebrate in vivo model.

In contrast to humans, zebrafish have two FDX1 genes, zFdx1 and zFdx1b. Our RT-PCR data shows that zFdx1 is maternally expressed and maintained throughout zygotic development. Its paralog, zFdx1b begins expression between 24 and 36 h post-fertilisation (hpf), which mirrors the expression of the main zebrafish P450 side-chain cleavage enzyme, zCyp11a2. Importantly, this time point coincides with the development of the zebrafish interrenal – the counterpart of the mammalian adrenal gland. While zFdx1 is ubiquitously expressed in adult zebrafish, zFdx1b expression is restricted to the main steroidogenic tissues: interrenal, gonads and brain. This suggests zFdx1b is the redox partner of steroidogenic mitochondrial CYP enzymes in the adult zebrafish. The in vivo function of the two zFdx paralogs was characterised using a transient morpholinos knockdown approach. Consistent with the expression data, knockdown of zFdx1 shows early morphological abnormalities during zebrafish development. Alternatively, zFdx1b morphants develop late onset metabolic abnormalities, consisting of an enlarged yolk and a delay in the inflation of their swim bladder at 120 hpf. Cortisol was extracted from zFdx1b deficient larvae and measured by liquid chromatography/tandem mass spectrometry. zFdx1b morphants failed to synthesise cortisol, compared to the injected controls.

This study gives in vivo insights into the molecular mechanism of mitochondrial redox regulation of steroidogenesis. We demonstrated that zFdx1b is essential for de novo steroidogenesis in zebrafish larvae, and that zFdx1 cannot compensate for Fdx1b function. Overall, our data establishes zebrafish as a model to study inborn errors of adrenal steroidogenesis.

Declaration of funding: This work was supported by the Elite Interntation Student Scholarship from the University of Birmingham (awarded to AG) and an Wellcome Trust ISSF grant (awarded to N K).