Immunocytochemical and biochemical demonstration of formaldhyde dehydrogenase (class III alcohol dehydrogenase) in the nucleus.

Abstract

Alcohol dehydrogenase (ADH), the major enzyme catalyzing the biological oxidation of ethanol in mammals, includes four classes with very different capacities for ethanol oxidation. Class III ADH is present in all the tissues and is well conserved throughout evolution. This enzyme has a low activity with ethanol, is specific for the glutathione-dependent oxidation of formaldehyde, and is therefore a formaldehyde dehydrogenase (FALDH). Until now there have been few and conflicting studies concerning its intracellular distribution, which is important for the understanding of its role in cell function. In the present work we used biochemical and immunocytochemical methods to assess the distribution of FALDH in rat hepatocytes and astroglial cells. With the glutathione-dependent formaldehyde dehydrogenase assay, we found the highest activity in the cytosol of hepatocytes and brain cells (12 and 2.6 mU/mg protein, respectively), but nuclei also exhibited significant activity (1.16 and 2.1 mU/mg protein, respectively). The immunocytochemical results showed the presence of FALDH binding sites in both the cytoplasm and the nucleus of the different cell types studied. Whereas no specific gold particle labeling was seen associated with any cytoplasmic component, in the nucleus the particles were found mainly over condensed chromatin and interchromatin regions. Finally, the gold particle density over both the nucleus and cytoplasm was greater in differentiated than in proliferating astrocytes in primary culture. In contrast, class I ADH, primarily responsible for ethanol metabolism, was found only in the cytoplasm of hepatocytes. We propose that one of the functions of FALDH is to protect cell structures, including DNA, from the toxic effects of endogenous formaldehyde, which is an intermediate in many metabolic process.

Merging protein, gene and genomic data: the evolution of the MDR-ADH family

“…Indeed, from our data, the widespread expression during zebrafish development was not ubiquitous, but consistent with spatio-temporal regulation (Dasmahapatra et al, 2001;Cañ estro et al, 2003). At the subcellular level, in contrast to other ADHs of strictly cytosolic localization, ADH3 has also been reported in the cell nucleus (Iborra et al, 1992;Fernández et al, 2003), presumably related to DNA protection. The ADH3 expression pattern in invertebrates, far from being widespread, has mainly been found in digestive tissues: in the deuterostomes amphioxus and ascidian, restricted to the posterior portion of the developing gut in the former, and to the anterior endoderm in the latter, which forms the gastric cavity after metamorphosis (Cañ estro et al, 2000(Cañ estro et al, , 2003.…”

Arabidopsis Formaldehyde Dehydrogenase

“…bovine serum albumin (BSA). The membrane was then incubated for 2 h with rabbit anti-(rat class 111 ADH) antiserum (Iborra et al, 1992) diluted 1 :I000 in Tris/NaCl/BSA, washed in Tris/NaCI/BSA for 5 min, incubated with goat anti-(rabbit IgG) alkaline-phosphatase conjugate (Bio-Rad) for 1 .5 h, and washed five times in Tris/NaCI/BSA for 5 min each time. The membrane was further developed by incubation in 100 mM TrisPHCI, 100 mM NaCI, 5 mM MgCl,, pH 9.5, containing 0.165 mglml 5-bromo-4-chloro-3-indolyl phosphate and 0.33 mg/ml nitro-blue tetrazolium.…”

Chronic Formaldehyde-Mediated Impairments and Age-Related Dementia

“…Aldehyde dehydrogenase I, and III (ADH I, ADH III), and alcohol dehydrogenase II (ALDH II) are three important enzymes involved in these reactions (Iborra et al, 1992;Zhiqian Tong, 2008). There are five classes (I-V) of alcohol dehydrogenase located in human tissues.…”