No abstract

Respiratory chain (RC) complexes are organized into supercomplexes forming 'respirasomes'. The mechanism underlying the interdependence of individual complexes is still unclear. Here, we show in human patient cells that the presence of a truncated COX1 subunit leads to destabilization of complex IV (CIV) and other RC complexes. Surprisingly, the truncated COX1 protein is integrated into subcomplexes, the holocomplex and even into supercomplexes, which however are all unstable. Depletion of the m-AAA protease AFG3L2 increases stability of the truncated COX1 and other mitochondrially encoded proteins, whereas overexpression of wild-type AFG3L2 decreases their stability. Both full-length and truncated COX1 proteins physically interact with AFG3L2. Expression of a dominant negative AFG3L2 variant also promotes stabilization of CIV proteins as well as the assembled complex and rescues the severe phenotype in heteroplasmic cells. Our data indicate that the mechanism underlying pathogenesis in these patients is the rapid clearance of unstable respiratory complexes by quality control pathways, rather than their impaired assembly.

Methionine restriction (MetR) extends lifespan in animal models including rodents. Using human diploid fibroblasts (HDF), we report here that MetR significantly extends their replicative lifespan, thereby postponing cellular senescence. MetR significantly decreased activity of mitochondrial complex IV and diminished the accumulation of reactive oxygen species. Lifespan extension was accompanied by a significant decrease in the levels of subunits of mitochondrial complex IV, but also complex I, which was due to a decreased translation rate of several mtDNA-encoded subunits. Together, these findings indicate that MetR slows down aging in human cells by modulating mitochondrial protein synthesis and respiratory chain assembly.

No abstract

This work explored the mechanism of augmented stress‐induced vascular reactivity of senescent murine femoral arteries (FAs). Mechanical and pharmacological reactivity of young (12–25 weeks, y‐FA) and senescent (>104 weeks, s‐FAs) femoral arteries was measured by wire myography. Expression and protein phosphorylation of selected regulatory proteins were studied by western blotting. Expression ratio of the Exon24 in/out splice isoforms of the regulatory subunit of myosin phosphatase, MYPT1 (MYPT1‐Exon24 in/out), was determined by polymerase chain reaction (PCR). While the resting length–tension relationship showed no alteration, the stretch‐induced‐tone increased to 8.3 ± 0.9 mN in s‐FA versus only 4.6 ± 0.3 mN in y‐FAs. Under basal conditions, phosphorylation of the regulatory light chain of myosin at S19 was 19.2 ± 5.8% in y‐FA versus 49.2 ± 12.6% in s‐FA. Inhibition of endogenous NO release raised tone additionally to 10.4 ± 1.2 mN in s‐FA, whereas this treatment had a negligible effect in y‐FAs (4.8 ± 0.3 mN). In s‐FAs, reactivity to NO donor was augmented (pD2 = −4.5 ± 0.3 in y‐FA vs. ‐5.2 ± 0.1 in senescent). Accordingly, in s‐FAs, MYPT1‐Exon24‐out‐mRNA, which is responsible for expression of the more sensitive to protein‐kinase G, leucine‐zipper‐positive MYPT1 isoform, was increased. The present work provides evidence that senescent murine s‐FA undergoes vascular remodelling associated with increases in stretch‐activated contractility and sensitivity to NO/cGMP/PKG system.