Ca2+ accumulation in the hydrogenosomes of Neocallimastix frontalis L2: A mitochondrial-like physiological role

The anaerobic fungus Neocallimastix frontalis L2 does not carry out oxidative phosphorylation but instead obtains energy from a fermentative metabolism. It does not have mitochondria, but does contain specialised redox organelles, the hydrogenosomes. With the aid of confocal laser scanning microscopy, we were able to follow the compartmentalisation of the Delta pH probe BCECF-AM (bis-(3-propyl-5-oxoisoxazol-4-yl)pentamethine(oxonolVi), 2',7'-bis(2-carboxyethyl)-5-carboxyfluorescein acetomethyl ester) into hydrogenosomes in situ. Association of the Ca2+-specific dye Fluo-3AM (1-[2-amino-5-(2,7-dichloro 6-hydroxy-3-oxy-9-xanthenyl)-phenoxyl]-2-[2-amino-5-methylphenoxy]ethane-N,N,N',N'-tetraacetic acid acetomethyl ester) within the lumen of the hydrogenosomes was also observed. In addition, transmission electron microscopy revealed that the hydrogenosomes contained electron-dense inclusions. X-ray microanalysis of these granules indicated the presence of calcium and phosphate. Our results indicate that hydrogenosomes from N. frontalis L2 maintain an internal alkaline pH and are involved in the calcium regulation of the cell. These physiological features resemble those of mitochondria from aerobic fungi.