Aldehyde dehydrogenase, Ald4p, is a major component of mitochondrial fluorescent inclusion bodies in the yeast Saccharomyces cerevisiae

Abstract

When Saccharomyces cerevisiae strain 3626 was cultured to the stationary phase in a medium that contained glucose, needle-like structures that emitted autofluorescence were observed in almost all cells by fluorescence microscopy under UV excitation. The needle-like structures completely overlapped with the profile of straight elongated mitochondria. Therefore, these structures were designated as mitochondrial fluorescent inclusion bodies (MFIBs). The MFIB-enriched mitochondrial fractions were successfully isolated and 2D-gel electrophoresis revealed that a protein of 54 kDa was only highly concentrated in the fractions. Determination of the N-terminal amino acid sequence of the 54-kDa protein identified it as a mitochondrial aldehyde dehydrogenase, Ald4p. Immunofluorescence microscopy showed that anti-Ald4p antibody specifically stained MFIBs. Freeze-substitution electron microscopy demonstrated that cells that retained MFIBs had electron-dense filamentous structures with a diameter of 10 nm in straight elongated mitochondria. Immunoelectron microscopy showed that Ald4p was localized to the electron-dense filamentous structures in mitochondria. These results together showed that a major component of MFIBs is Ald4p. In addition, we demonstrate that MFIBs are common features that appear in mitochondria of many species of yeast.

Organization and dynamics of yeast mitochondrial nucleoids

“…We isolated mitochondria that included fluorescent inclusion bodies and then identified the major component of mitochondrial fluorescent inclusion body as a mitochondrial aldehyde dehydrogenase, Ald4. 106 ) Subsequently, we confirmed the localization of Ald4 in the inclusion body by means of both immunoelectron microscopy and immunofluorescence microscopy. 106 ) Mitochondria with mitochondrial inclusion bodies are very long in length and elongate straightforward.…”

A quantitative screen for metabolic enzyme structures reveals patterns of assembly across the yeast metabolic network

“…This is particularly problematic because the compartmentalization of several metabolic pathways to mitochondria is a key organizing principle of metabolic biochemistry. Furthermore, the yeast metabolic enzyme Ald4p has recently been found to form structures within the mitochondria, suggesting that a subset of metabolic enzymes might be forming within other compartments (Misonou et al , 2014; Noree, 2018). The fact that our screen identified 24 putative mitochondrial metabolic enzymes presented us with a unique opportunity to explore the extent of metabolic enzyme self-assembly within mitochondria.…”

Extramitochondrial Assembly of Mitochondrial Targeting Signal Disrupted Mitochondrial Enzyme Aldehyde Dehydrogenase

“…From re-screening the yeast GFP collection to search for any GFP-tagged metabolic enzyme capable of supramolecular assembly, I found that yeast mitochondrial enzyme aldehyde dehydrogenase (Ald4p), involved in acetate biosynthesis, and also, ethanol and pyruvate metabolism 16 , seemed likely to possess this property. Through the literature search, this enzyme has recently been reported that it is a component of the needle-like structures, present in fused mitochondria 18 .…”