Autophagy maturation associated with CD38-mediated regulation of lysosome function in mouse glomerular podocytes.
Podocytes are highly differentiated glomerular epithelial cells that contribute to the glomerular barrier function of kidney. A role for autophagy has been proposed in maintenance of their cellular integrity, but the mechanisms controlling autophagy in podocytes are not clear. The present study tested whether CD38-mediated regulation of lysosome function contributes to autophagic flux or autophagy maturation in podocytes. Podocytes were found to exhibit a high constitutive level of LC3-II, a robust marker of autophagosomes (APs), suggesting a high basal level of autophagic activity. Treatment with the mTOR inhibitor, rapamycin, increased LC3-II and the content of both APs detected by Cyto-ID Green staining and autophagolysosomes (APLs) measured by acridine orange staining and colocalization of LC3 and Lamp1. Lysosome function inhibitor bafilomycin A1 increased APs, but decreased APLs content under both basal and rapamycin-induced conditions. Inhibition of CD38 activity by nicotinamide or silencing of CD38 gene produced the similar effects to that bafilomycin A1 did in podocytes. To explore the possibility that CD38 may control podocyte autophagy through its regulation of lysosome function, the fusion of APs with lysosomes in living podocytes was observed by co-transfection of GFP-LC3B and RFP-Lamp1 expression vectors. A colocalization of GFP-LC3B and RFP-Lamp1 upon stimulation of rapamycin became obvious in transfected podocytes, which could be substantially blocked by nicotinamide, CD38 shRNA, and bafilomycin. Moreover, blockade of the CD38-mediated regulation by PPADS completely abolished rapamycin-induced fusion of APs with lysosomes. These results indicate that CD38 importantly control lysosomal function and influence autophagy at the maturation step in podocytes.
Department of Pharmacology and Toxicology, Virginia Commonwealth University School of Medicine, Richmond, VA, USA.
Jing
J
Xiong
Min
M
Xia
Ming
M
Xu
Yang
Y
Zhang
Justine M
JM
Abais
Guangbi
G
Li
Christopher R
CR
Riebling
Joseph K
JK
Ritter
Krishna M
KM
Boini
Pin-Lan
PL
Li
D051997:ADP-ribosyl Cyclase 1 / Q000378:metabolism*; D000246:Adenosine Diphosphate Ribose / Q000378:metabolism; D000818:Animals; D001343:Autophagy / Q000187:drug effects; D020013:Calcium Signaling / Q000187:drug effects; D020868:Gene Silencing / Q000187:drug effects; D007678:Kidney Glomerulus / Q000166:cytology*; D008247:Lysosomes / Q000187:drug effects / Q000378:metabolism*; D008561:Membrane Fusion / Q000187:drug effects; D051379:Mice; D009249:NADP / Q000031:analogs & derivatives / Q000378:metabolism; D010588:Phagosomes / Q000187:drug effects / Q000378:metabolism; D050199:Podocytes / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D021381:Protein Transport / Q000187:drug effects; D059748:Proteolysis / Q000187:drug effects; D020123:Sirolimus / Q000494:pharmacology; D051758:Transcription Factor TFIIH; D014157:Transcription Factors / Q000378:metabolism; D057149:Ubiquitinated Proteins / Q000378:metabolism
C490981:Gtf2h1 protein, mouse; D014157:Transcription Factors; D057149:Ubiquitinated Proteins; D051758:Transcription Factor TFIIH; D000246:Adenosine Diphosphate Ribose; D009249:NADP; C024376:NAADP; D051997:ADP-ribosyl Cyclase 1; D020123:Sirolimus
ADP-ribosylcyclase; CD38; autophagic flux; glomeruli; lysosome trafficking