Endoplasmic reticulum stress induces autophagy in renal proximal tubular cells.
BACKGROUND
Autophagy, an intracellular self-degradation system conserved throughout eukaryotes, plays an important role in a variety of biological processes, including cell death, development, cancer, defence against infection and neurodegeneration. However, little information about autophagy in renal tubular cells is available. We investigated the relationship of autophagy with endoplasmic reticulum (ER) stress in proximal tubular cells.


METHODS
Immortalized rat proximal tubular cells were exposed to the classic ER stress inducers tunicamycin or brefeldin A. Autophagy was detected mainly by immunoblot analysis of LC3, a widely used marker of autophagy, and also by immunofluorescent cytochemistry of LC3 and electron microscopy. Biological significance of the phenomenon was studied using bafilomycin A1, an inhibitor of autophagosome degradation. Signal transduction pathways following ER stress were also investigated using inhibitors of the MAPK pathway.


RESULTS
Both ER stress inducers significantly increased LC3-II as a marker of autophagy in immunoblot analysis. Immunocytochemistry of LC3 and electron microscopy also showed activation of autophagy by ER stress. Inhibition by bafilomycin A1 showed that autophagy following ER stress fulfilled its intrinsic function, namely degradation of cytoplasmic components. Further, use of the MEK 1/2 inhibitor U0126, which inhibits ER stress-induced autophagy induction and ERK activation, showed that ERK, a MAPK family member, was necessary to the induction of autophagy.


CONCLUSIONS
For the first time, we demonstrate the induction of autophagy following ER stress in renal tubules, and clarify its mechanism. These findings serve as the foundation for further investigation into autophagy in renal diseases.
Kawakami
Takahisa
T
University of Tokyo School of Medicine, Japan.
Inagi
Reiko
R
Takano
Hideki
H
Sato
Shigeru
S
Ingelfinger
Julie R
JR
Fujita
Toshiro
T
Nangaku
Masaomi
M
D000818:Animals; D001343:Autophagy / Q000187:drug effects / Q000502:physiology*; D002460:Cell Line; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000502:physiology*; D007150:Immunohistochemistry; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D020935:MAP Kinase Signaling System / Q000187:drug effects; D018942:Macrolides / Q000494:pharmacology; D046529:Microscopy, Electron, Transmission; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D010588:Phagosomes / Q000648:ultrastructure; D051381:Rats; D013312:Stress, Physiological; D014415:Tunicamycin / Q000494:pharmacology
D018942:Macrolides; D008869:Microtubule-Associated Proteins; D014415:Tunicamycin; C526109:Anxa3 protein, rat; C040929:bafilomycin A1