Abstract
The dynamin-related GTPase protein OPA1, localized in the intermembrane space and tethered to the inner membrane of mitochondria, participates in the fusion of these organelles. Its mutation is the most prevalent cause of Autosomal Dominant Optic Atrophy. OPA1 controls the diameter of the junctions between the boundary part of the inner membrane and the membrane of cristae and reduces the diffusibility of cytochrome c through these junctions. We postulated that if significant Ca2+ uptake into the matrix occurs from the lumen of the cristae, reduced expression of OPA1 would increase the access of Ca2+ to the transporters in the crista membrane and thus would enhance Ca2+ uptake. In intact H295R adrenocortical and HeLa cells cytosolic Ca2+ signals evoked with K+ and histamine, respectively, were transferred into the mitochondria. The rate and amplitude of mitochondrial [Ca2+] rise (followed with confocal laser scanning microscopy and FRET measurements with fluorescent wide-field microscopy) were increased after knockdown of OPA1, as compared with cells transfected with control RNA or mitofusin1 siRNA. Ca2+ uptake was enhanced despite reduced mitochondrial membrane potential. In permeabilized cells the rate of Ca2+ uptake by depolarized mitochondria was also increased in OPA1-silenced cells. The participation of Na+/Ca2+ and Ca2+/H+ antiporters in this transport process is indicated by pharmacological data. Altogether, our observations reveal the significance of OPA1 in the control of mitochondrial Ca2+ metabolism.
Highlights
Recent observations obtained in imaging and electron tomographic studies revealed a dynamically changing structure [1,2] and led to a revised concept of the structure and function of mitochondria
H295R cells may be accounted for by the poor transfectability of this cell line.) To test whether Mfn1-silenced cells are more appropriate controls for OPA1-silenced cells than those transfected with control RNA, the morphology of mitochondria was compared in the three groups
In H295R cells the median value of the length of single mitochondria diminished from 2.30 mm in control RNA-treated cells to 1.08 and 0.98 mm in cells exposed to Mfn1 and OPA1 siRNA, respectively (Figure S3)
Summary
Recent observations obtained in imaging and electron tomographic studies revealed a dynamically changing structure [1,2] and led to a revised concept of the structure and function of mitochondria. The changes in the number and size of mitochondria involve alterations in the inner mitochondrial membrane (IMM). The cristae are connected to the inner boundary membrane (i.e. the part of IMM between two neighboring cristae) by narrow tubular junctions which have a diameter of 15–40 nm [3,4,5,6]. These junctions may impede free diffusion and may induce the formation of a gradient of ions, molecules and macromolecules between the intermembrane space (IMS) and the lumen of the cristae [5,7]. Reduced expression of OPA1 was reported in ischaemic heart failure [15] showing that insufficient expression of the protein may have far-reaching consequences
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