OPA1 encodes a large GTPase related to dynamins, anchored to the mitochondrial cristae inner membrane, facing the intermembrane space. OPA1 haplo-insufficiency is responsible for the most common form of autosomal dominant optic atrophy (ADOA, MIM165500), a neuropathy resulting from degeneration of the retinal ganglion cells and optic nerve atrophy. Here we show that down-regulation of OPA1 in HeLa cells using specific small interfering RNA (siRNA) leads to fragmentation of the mitochondrial network concomitantly to the dissipation of the mitochondrial membrane potential and to a drastic disorganization of the cristae. These events are followed by cytochrome c release and caspasedependent apoptotic nuclear events. Similarly, in NIH-OVCAR-3 cells, the OPA1 siRNA induces mitochondrial fragmentation and apoptosis, the latter being inhibited by Bcl2 overexpression. These results suggest that OPA1 is a major organizer of the mitochondrial inner membrane from which the maintenance of the cristae integrity depends. As loss of OPA1 commits cells to apoptosis without any other stimulus, we propose that OPA1 is involved in the cytochrome c sequestration and might be a target for mitochondrial apoptotic effectors. Our results also suggest that abnormal apoptosis is a possible pathophysiological process leading to the retinal ganglion cells degeneration in ADOA patients.

Ribosome biogenesis in eukaryotes depends on the coordinated action of ribosomal and nonribosomal proteins that guide the assembly of preribosomal particles. These intermediate particles follow a maturation pathway in which important changes in their protein composition occur. The mechanisms involved in the coordinated assembly of the ribosomal particles are poorly understood. We show here that the association of preribosomal factors with pre-60S complexes depends on the presence of earlier factors, a phenomenon essential for ribosome biogenesis. The analysis of the composition of purified preribosomal complexes blocked in maturation at specific steps allowed us to propose a model of sequential protein association with, and dissociation from, early pre-60S complexes for several preribosomal factors such as Mak11, Ssf1, Rlp24, Nog1, and Nog2. The presence of either Ssf1 or Nog2 in complexes that contain the 27SB pre-rRNA defines novel, distinct pre-60S particles that contain the same pre-rRNA intermediates and that differ only by the presence or absence of specific proteins. Physical and functional interactions between Rlp24 and Nog1 revealed that the assembly steps are, at least in part, mediated by direct protein-protein interactions.The synthesis of ribosomes is one of the major metabolic pathways of a cell. In Saccharomyces cerevisiae, ribosome assembly begins in the nucleolus after the transcription of two rRNA precursors, the 35S RNA (precursor of the 18S, 5.8S, and 25S rRNAs) and the pre-5S RNA, by RNA polymerases I and III, respectively. The synthesized pre-rRNAs are modified extensively at multiple positions specified by small nucleolar ribonucleoparticles (snoRNPs) or specific enzymes (1,22,33). During rRNA maturation, the 5Ј and 3Ј external transcribed sequences (ETS) and internal transcribed sequence 1 (ITS1) and ITS2 are removed from the 35S precursor RNA by wellordered cleavages and trimming events, which require the enzymatic activities of helicases and endo-and exonucleases (19,37).Cotranscriptional assembly of ribosomal and nonribosomal proteins in the nucleolus gives rise to a large ribonucleoprotein particle corresponding to the 90S preribosomal complexes described more than 20 years ago (35) and recently characterized biochemically (8,14). These early preribosomal complexes are further converted to smaller pre-40S (43S) and pre-60S (66S) particles, precursors of the mature small and large ribosomal subunits. The pre-40S complexes, each containing a precursor of the 18S rRNA, are exported into the cytoplasm, where they give rise to the mature 40S ribosomal particles (36). Most of the large ribosomal subunit proteins are absent from the 90S preribosomes (8,14) and associate in the nucleolus with the pre-rRNA, probably concomitantly with the formation of the pre-60S particles. During pre-60S particle maturation, 27S prerRNA intermediates are converted into 25S and 5.8S mature rRNAs by successive and well-ordered steps. Several pre-60S particles, which differ in their RNA and protein compositions,...