TY - JOUR
T1 - SIRT5 regulation of ammonia-induced autophagy and mitophagy
AU - Polletta, Lucia
AU - Vernucci, Enza
AU - Carnevale, Ilaria
AU - Arcangeli, Tania
AU - Rotili, Dante
AU - Palmerio, Silvia
AU - Steegborn, Clemens
AU - Nowak, Theresa
AU - Schutkowski, Mike
AU - Pellegrini, Laura
AU - Sansone, Luigi
AU - Villanova, Lidia
AU - Runci, Alessandra
AU - Pucci, Bruna
AU - Morgante, Emanuela
AU - Fini, Massimo
AU - Mai, Antonello
AU - Russo, Matteo A.
AU - Tafani, Marco
PY - 2015
Y1 - 2015
N2 - In liver the mitochondrial sirtuin, SIRT5, controls ammonia detoxification by regulating CPS1, the first enzyme of the urea cycle. However, while SIRT5 is ubiquitously expressed, urea cycle and CPS1 are only present in the liver and, to a minor extent, in the kidney. To address the possibility that SIRT5 is involved in ammonia production also in nonliver cells, clones of human breast cancer cell lines MDA-MB-231 and mouse myoblast C2C12, overexpressing or silenced for SIRT5 were produced. Our results show that ammonia production increased in SIRT5-silenced and decreased in SIRT5- overexpressing cells. We also obtained the same ammonia increase when using a new specific inhibitor of SIRT5 called MC3482. SIRT5 regulates ammonia production by controlling glutamine metabolism. In fact, in the mitochondria, glutamine is transformed in glutamate by the enzyme glutaminase, a reaction producing ammonia. We found that SIRT5 and glutaminase coimmunoprecipitated and that SIRT5 inhibition resulted in an increased succinylation of glutaminase. We next determined that autophagy and mitophagy were increased by ammonia by measuring autophagic proteolysis of long-lived proteins, increase of autophagy markers MAP1LC3B, GABARAP, and GABARAPL2, mitophagy markers BNIP3 and the PINK1-PARK2 system as well as mitochondrial morphology and dynamics. We observed that autophagy and mitophagy increased in SIRT5-silenced cells and in WT cells treated with MC3482 and decreased in SIRT5-overexpressing cells. Moreover, glutaminase inhibition or glutamine withdrawal completely prevented autophagy. In conclusion we propose that the role of SIRT5 in nonliver cells is to regulate ammonia production and ammonia-induced autophagy by regulating glutamine metabolism.
AB - In liver the mitochondrial sirtuin, SIRT5, controls ammonia detoxification by regulating CPS1, the first enzyme of the urea cycle. However, while SIRT5 is ubiquitously expressed, urea cycle and CPS1 are only present in the liver and, to a minor extent, in the kidney. To address the possibility that SIRT5 is involved in ammonia production also in nonliver cells, clones of human breast cancer cell lines MDA-MB-231 and mouse myoblast C2C12, overexpressing or silenced for SIRT5 were produced. Our results show that ammonia production increased in SIRT5-silenced and decreased in SIRT5- overexpressing cells. We also obtained the same ammonia increase when using a new specific inhibitor of SIRT5 called MC3482. SIRT5 regulates ammonia production by controlling glutamine metabolism. In fact, in the mitochondria, glutamine is transformed in glutamate by the enzyme glutaminase, a reaction producing ammonia. We found that SIRT5 and glutaminase coimmunoprecipitated and that SIRT5 inhibition resulted in an increased succinylation of glutaminase. We next determined that autophagy and mitophagy were increased by ammonia by measuring autophagic proteolysis of long-lived proteins, increase of autophagy markers MAP1LC3B, GABARAP, and GABARAPL2, mitophagy markers BNIP3 and the PINK1-PARK2 system as well as mitochondrial morphology and dynamics. We observed that autophagy and mitophagy increased in SIRT5-silenced cells and in WT cells treated with MC3482 and decreased in SIRT5-overexpressing cells. Moreover, glutaminase inhibition or glutamine withdrawal completely prevented autophagy. In conclusion we propose that the role of SIRT5 in nonliver cells is to regulate ammonia production and ammonia-induced autophagy by regulating glutamine metabolism.
KW - Ammonia
KW - Autophagy
KW - Glutaminase
KW - Glutamine
KW - Mitochondrial dynamics sirtuin 5
KW - Mitophagy
KW - Molecular rehabilitation
KW - Succinylation
UR - http://www.scopus.com/inward/record.url?scp=84943265499&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84943265499&partnerID=8YFLogxK
U2 - 10.1080/15548627.2015.1009778
DO - 10.1080/15548627.2015.1009778
M3 - Article
AN - SCOPUS:84943265499
VL - 11
SP - 253
EP - 270
JO - Autophagy
JF - Autophagy
SN - 1554-8627
IS - 2
ER -