SMAD3 promotes autophagy dysregulation by triggering lysosome depletion in tubular epithelial cells in diabetic nephropathy

Abstract

Macroautophagy/autophagy dysregulation has been noted in diabetic nephropathy; however, the regulatory mechanisms controlling this process remain unclear. In this study, we showed that SMAD3 (SMAD family member 3), the key effector of TGFB (transforming growth factor beta)-SMAD signaling, induces lysosome depletion via the inhibition of TFEB-dependent lysosome biogenesis. The pharmacological inhibition or genetic deletion of SMAD3 restored lysosome biogenesis activity by alleviating the suppression of TFEB, thereby protecting lysosomes from depletion and improving autophagic flux in renal tubular epithelial cells in diabetic nephropathy. Mechanistically, we found that SMAD3 directly binds to the 3ʹ-UTR of TFEB and inhibits its transcription. Silencing TFEB suppressed lysosome biogenesis and resulted in a loss of the protective effects of SMAD3 inactivation on lysosome depletion under diabetic conditions. In conclusion, SMAD3 promotes lysosome depletion via the inhibition of TFEBdependent lysosome biogenesis; this may be an important mechanism underlying autophagy dysregulation in the progression of diabetic nephropathy.

Update on the Mechanisms of Tubular Cell Injury in Diabetic Kidney Disease

“…Theodomir et al confirmed that P2Y2R deficiency increased the expression of sirtuin-1 and FOXO3a, which enhanced autophagy and improved renal insufficiency in DKD (95). In addition, Yang et al found that Smad3, the downstream transcription factor activated by TGFβ (transforming growth factor β), suppressed lysosome biogenesis in a TFEB-dependent manner (96). Furthermore, ATF4 (activating transcription factor 4) (97), TRAIL (TNF related apoptosis inducing ligand) (98), Soluble epoxide hydrolase (sEH) and lys63 UB proteins were also confirmed to be involved in the regulation of autophagy in the kidney cells.…”

Cyclosporine A blocks autophagic flux in tubular epithelial cells by impairing TFEB‐mediated lysosomal function

“…Human proximal tubular HK‐2 cells (ATCC, CRL‐2190™, Manassas, VA, USA) and mouse renal tubular epithelial cells (mTECs, a gift from Dr Jeffrey B. Kopp, NIH, Bethesda, MD, USA) were cultured as previously described 23 . To mimic CsA‐induced nephrotoxicity in vitro, the cells were treated with 5, 7.5 and 10 μmol/L CsA (Selleck, S2286) for 24 h or an equal amount of ethyl alcohol in the control group.…”

“…Western blot analysis was performed as previously described 23 . The membranes were incubated at 4°C overnight with the following primary antibodies: anti‐collagen I (Abcam, ab34710), anti‐TGF‐β1 (Abcam, ab92486), anti‐fibronectin (Abcam, ab23750), anti‐LC3B (Sigma, L7543), anti‐p62/SQSTM1 (Abcam, ab91526), anti‐Cathepsin B (Abcam, ab58802), anti‐phospho‐mTOR (Ser2448) (Cell Signaling, 5536S), anti‐mTOR (Cell Signaling, 2983S), anti‐phospho‐4E‐BP1(Thr37/46) (Cell Signaling, 2855S), anti‐4E‐BP1 (Cell Signaling, 9644S), anti‐phospho‐TFEB (Ser142) (Millipore, ABE1971‐I), anti‐TFEB (BETHYL, A303‐672).…”

Epigenetics and Inflammation in Diabetic Nephropathy

“…TGF-β/SMAD signaling pathway plays a criticial role in diabetic kidney injuries ( Chen et al, 2011 , 2014a ; Liu et al, 2011 ; Lan, 2012 ; Zhong et al, 2013 ; Li et al, 2014 ; Zhang et al, 2019b ; Xu et al, 2020a ; Yang et al, 2020 ). In the diabetic kidney, high glucose and AGEs enhance the phosphorylation of SMAD3 and decrease the phosphorylation SMAD7.…”

“…SMAD3 deficiency protects against diabetes-associated beta cell dysfunction and loss in DN mice ( Sheng et al, 2021 ). SMAD3 also promotes autophagy dysregulation and kidney injury ( Yang et al, 2020 ). SMAD7 inhibits IκBα, an NF-κB inhibitor, suppressing the activation of NF-κB pathway ( Chung et al, 2009 ).…”