Lack of functional and expression homology between human and mouse aldo-keto reductase 1C enzymes: implications for modelling human cancers

<p>Abstract</p> <p>Background</p> <p>Over recent years, enzymes of the aldo-keto reductase (AKR) 1C subfamily have been implicated in the progression of prostate, breast, endometrial and leukemic cancers. This is due to the ability of AKR1C enzymes to modify androgens, estrogens, progesterone and prostaglandins (PGs) in a tissue-specific manner, regulating the activity of nuclear receptors and other downstream effects. Evidence supporting a role for AKR1C enzymes in cancer derives mostly from studies with isolated primary cells from patients or immortalized cell lines. Mice are ideal organisms for <it>in vivo </it>studies, using knock-out or over-expression strains. However, the functional conservation of AKR1C enzymes between human and mice has yet to be described.</p> <p>Results</p> <p>In this study, we have characterized and compared the four human (AKR1C1,-1C2, -1C3 and -1C4) and the eight murine (AKR1C6, -1C12, -1C13, -1C14, -1C18, -1C19, -1C20 and -1C21) isoforms in their phylogeny, substrate preference and tissue distribution. We have found divergent evolution between human and murine AKR1C enzymes that was reflected by differing substrate preference. Murine enzymes did not perform the 11β-ketoreduction of prostaglandin (PG) D₂, an activity specific to human AKR1C3 and important in promoting leukemic cell survival. Instead, murine AKR1C6 was able to perform the 9-ketoreduction of PGE₂, an activity absent amongst human isoforms. Nevertheless, reduction of the key steroids androstenedione, 5α-dihydrotestosterone, progesterone and estrone was found in murine isoforms. However, unlike humans, no AKR1C isoforms were detected in murine prostate, testes, uterus and haemopoietic progenitors.</p> <p>Conclusions</p> <p>This study exposes significant lack of phylogenetic and functional homology between human and murine AKR1C enzymes. Therefore, we conclude that mice are not suitable to model the role of AKR1C in human cancers and leukemia.</p

Cortisone-reductase deficiency associated with heterozygous mutations in 11β-hydroxysteroid dehydrogenase type 1

In peripheral target tissues, levels of active glucocorticoid hormones are controlled by 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1), a dimeric enzyme that catalyzes the reduction of cortisone to cortisol within the endoplasmic reticulum. Loss of this activity results in a disorder termed cortisone reductase deficiency (CRD), typified by increased cortisol clearance and androgen excess. To date, only mutations in H6PD, which encodes an enzyme supplying cofactor for the reaction, have been identified as the cause of disease. Here we examined the HSD11B1 gene in two cases presenting with biochemical features indicative of a milder form of CRD in whom the H6PD gene was normal. Novel heterozygous mutations (R137C or K187N) were found in the coding sequence of HSD11B1. The R137C mutation disrupts salt bridges at the subunit interface of the 11β-HSD1 dimer, whereas K187N affects a key active site residue. On expression of the mutants in bacterial and mammalian cells, activity was either abolished (K187N) or greatly reduced (R137C). Expression of either mutant in a bacterial system greatly reduced the yield of soluble protein, suggesting that both mutations interfere with subunit folding or dimer assembly. Simultaneous expression of mutant and WT 11β-HSD1 in bacterial or mammalian cells, to simulate the heterozygous condition, indicated a marked suppressive effect of the mutants on both the yield and activity of 11β-HSD1 dimers. Thus, these heterozygous mutations in the HSD11B1 gene have a dominant negative effect on the formation of functional dimers and explain the genetic cause of CRD in these patients

The interface between subunits in the head-to-tail arrangement.

<p><b>A</b>) Human frataxin interface with residues D112, D115, D122, and D124 marked in magenta. The N-terminus is colored in pink and the different monomers within the dimer are shown in yellow (head) and green (tail). A crystal structure (PDB entry 1EKG) was used for preparing the figure. <b>B</b>) CyaY interface and the residues making up the potential metal binding sites are shown. Residues H7, E19, D22, and D23, which may build up the first metal binding site, are shown as red sticks; D3, H58, and D25 may participate in the second site (yellow sticks); and H70, D29, and E44 (blue sticks) in the third. Gray spheres show Europium ions bound in the 2P1X crystal structure. Residues involved in metal binding in the crystal structures of CyaY in complex with Co and/or Eu are labeled. The different monomers in the dimer are shown in green (head) and blue (tail). Crystal structures (PDB entry 2P1X, 2EFF, and 1EW4) were used in the preparation of the figue.</p