ENZYMATIC CONVERSION OF RETINALDEHYDE TO RETINOIC ACID BY CLONED MURINE CYTOSOLIC AND MITOCHONDRIAL ALDEHYDE DEHYDROGENASES

It has previously been reported that retinaldehyde can be converted to retinoic acid by cytosolic aldehyde dehydrogenase (AHD-2) in liver ex tracts [Biochem. Pharmacol. 42: 1279-1285 (1991)]. To determine which enzyme(s) carried out this reaction in murine embryonic stem cells, tw o aldehyde dehydrogenases were cloned; the AHD-2 gene was cloned from a liver cDNA library, and a closely related gene, AHD-M1, was cloned f rom an embryonic F9 cell cDNA library by conserved oligonucleotide seq uence screening. AHD-M1 contained an open reading frame of 1554 base p airs, which encoded 517 amino acids. The AHD-M1 gene encoded a protein with a putative amino acid sequence that was 94% and 97% identical to the mitochondrial aldehyde dehydrogenases of human and rat, respectiv ely, and thus we have cloned the murine cDNA for this enzyme for the f irst time. The AHD-M1 cDNA was only 64% identical to AHD-2. Northern a nalysis showed that AHD-M1 mRNA was constitutively expressed in F9 and P19 embryonic teratocarcinoma stem cells and in AB1 embryonic stem ce lls. There was a 3-5-fold retinoic acid-associated increase in the amo unt of this mRNA during the differentiation of F9 cells into parietal endoderm. In contrast, we could not detect the expression of AHD-2 mRN A in AB1, P19, or F9 cells, even though the F9 cells could convert ret inaldehyde to retinoic acid. When the AHD-M1 and AHD-2 cDNAs were inse rted into the expression vector pSG5 and transfected into cultured COS cells, 3-5-fold and 100-fold increases, respectively, in the conversi on of [H-3]retinaldehyde to [H-3]retinoic acid could be detected by hi gh performance liquid chromatographic assay. We conclude that both enz ymes are capable of converting retinaldehyde to retinoic acid in intac t COS cells. AHD-2 is more active than AHD-M1 in this conversion, but AHD-2 is not the enzyme responsible for this conversion in F9 embryoni c stem cells.