Autophagy Impairment Induces Premature Senescence in Primary Human Fibroblasts

BACKGROUND:Recent studies have demonstrated that activation of autophagy increases the lifespan of organisms from yeast to flies. In contrast to the lifespan extension effect in lower organisms, it has been reported that overexpression of unc-51-like kinase 3 (ULK3), the mammalian homolog of autophagy-specific gene 1 (ATG1), induces premature senescence in human fibroblasts. Therefore, we assessed whether the activation of autophagy would genuinely induce premature senescence in human cells. METHODOLOGY/PRINCIPAL FINDINGS:Depletion of ATG7, ATG12, or lysosomal-associated membrane protein 2 (Lamp2) by transfecting siRNA or infecting cells with a virus containing gene-specific shRNA resulted in a senescence-like state in two strains of primary human fibroblasts. Prematurely senescent cells induced by autophagy impairment exhibited the senescent phenotypes, similar to the replicatively senescent cells, such as increased senescence associated β-galactosidase (SA-β-gal) activity, reactive oxygen species (ROS) generation, and accumulation of lipofuscin. In addition, expression levels of ribosomal protein S6 kinase1 (S6K1), p-S6K1, p-S6, and eukaryotic translation initiation factor 4E (eIF4E) binding protein 1 (4E-BP1) in the mammalian target of rapamycin (mTOR) pathway and beclin-1, ATG7, ATG12-ATG5 conjugate, and the sequestosome 1 (SQSTM1/p62) monomer in the autophagy pathway were decreased in both the replicatively and the autophagy impairment-induced prematurely senescent cells. Furthermore, it was found that ROS scavenging by N-acetylcysteine (NAC) and inhibition of p53 activation by pifithrin-α or knockdown of p53 using siRNA, respectively, delayed autophagy impairment-induced premature senescence and restored the expression levels of components in the mTOR and autophagy pathways. CONCLUSION:Taken together, we concluded that autophagy impairment induces premature senescence through a ROS- and p53-dependent manner in primary human fibroblasts

Role for calcium from the sarcoplasmic reticulum in coupling muscle activity to nicotinic acetylcholine receptor gene expression in rat

Neurally evoked muscle electrical activity suppresses nicotinic acetylcholine receptor (nAChR) gene expression in extrajunctional domains of adult muscle fibers. It has been proposed that this regulation is mediated by calcium influx through voltage-dependent L-type calcium channels but bypasses the sarcoplasmic reticulum in chick and mouse C2C12 cells. Here we report that in rat muscle calcium influx through L-type calcium channels preferentially reduced nAChR ε-subunit RNA via a post-transcriptional mechanism. In contrast, calcium release from the sarcoplasmic reticulum (SR) suppressed nAChR subunit RNA levels as a result of decreasing nAChR subunit promoter activity. Finally, we show that this decreased promoter activity is mediated through the same DNA sequences that control activity-dependent gene expression. Therefore, we propose that in rat muscle, calcium release from the SR participates in coupling muscle depolarization to nAChR gene expression. © 1998 John Wiley & Sons, Inc. J Neurobiol 35: 245–257, 1998Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/34476/1/2_ftp.pd