Cisplatin Nephrotoxicity Is Critically Mediated by the Availability of BECLIN1.

Cisplatin nephrotoxicity is a critical limitation of solid cancer treatment. Until now, the complex interplay of various pathophysiological mechanisms leading to proximal tubular cell apoptosis after cisplatin exposure has not been fully understood. In our study, we assessed the role of the autophagy-related protein BECLIN1 (ATG6) in cisplatin-induced acute renal injury (AKI)-a candidate protein involved in autophagy and with putative impact on apoptosis by harboring a B-cell lymphoma 2 (BCL2) interaction site of unknown significance. By using mice with heterozygous deletion of Becn1, we demonstrate that reduced intracellular content of BECLIN1 does not impact renal function or autophagy within 12 months. However, these mice were significantly sensitized towards cisplatin-induced AKI, and by using Becn1+/-;Sglt2-Cre;Tomato/EGFP mice with subsequent primary cell analysis, we confirmed that nephrotoxicity depends on proximal tubular BECLIN1 content. Mechanistically, BECLIN1 did not impact autophagy or primarily the apoptotic pathway. In fact, a lack of BECLIN1 sensitized mice towards cisplatin-induced ER stress. Accordingly, the ER stress inhibitor tauroursodeoxycholic acid (TUDCA) blunted cisplatin-induced cell death in Becn1 heterozygosity. In conclusion, our data first highlight a novel role of BECLIN1 in protecting against cellular ER stress independent from autophagy. These novel findings open new therapeutic avenues to intervene in this important intracellular stress response pathway with a promising impact on future AKI management.

BECLIN1 Is Essential for Podocyte Secretory Pathways Mediating VEGF Secretion and Podocyte-Endothelial Crosstalk.

Vascular endothelial growth factor A (VEGFA) secretion from podocytes is crucial for maintaining endothelial integrity within the glomerular filtration barrier. However, until now, the molecular mechanisms underlying podocyte secretory function remained unclear. Through podocyte-specific deletion of BECLIN1 (ATG6 or Becn1), a key protein in autophagy initiation, we identified a major role for this molecule in anterograde Golgi trafficking. The Becn1-deficient podocytes displayed aberrant vesicle formation in the trans-Golgi network (TGN), leading to dramatic vesicle accumulation and complex disrupted patterns of intracellular vesicle trafficking and membrane dynamics. Phenotypically, podocyte-specific deletion of Becn1 resulted in early-onset glomerulosclerosis, which rapidly progressed and dramatically reduced mouse life span. Further, in vivo and in vitro studies clearly showed that VEGFA secretion, and thereby endothelial integrity, greatly depended on BECLIN1 availability and function. Being the first to demonstrate the importance of a secretory pathway for podocyte integrity and function, we identified BECLIN1 as a key component in this complex cellular process. Functionally, by promoting VEGFA secretion, a specific secretory pathway emerged as an essential component for the podocyte-endothelial crosstalk that maintains the glomerular filtration barrier.

A reciprocal regulation of spermidine and autophagy in podocytes maintains the filtration barrier.

Podocyte maintenance and stress resistance are exquisitely based on high basal rates of autophagy making these cells a unique model to unravel mechanisms of autophagy regulation. Polyamines have key cellular functions such as proliferation, nucleic acid biosynthesis and autophagy. Here we test whether endogenous spermidine signaling is a driver of basal and dynamic autophagy in podocytes by using genetic and pharmacologic approaches to interfere with different steps of polyamine metabolism. Translational studies revealed altered spermidine signaling in focal segmental glomerulosclerosis in vivo and in vitro. Exogenous spermidine supplementation emerged as new treatment strategy by successfully activating autophagy in vivo via inhibition of EP300, a protein with an essential role in controlling cell growth, cell division and prompting cells to differentiate to take on specialized functions. Surprisingly, gas chromatography-mass spectroscopy based untargeted metabolomics of wild type and autophagy deficient primary podocytes revealed a positive feedback mechanism whereby autophagy itself maintains polyamine metabolism and spermidine synthesis. The transcription factor MAFB acted as an upstream regulator of polyamine metabolism. Thus, our data highlight a novel positive feedback loop of autophagy and spermidine signaling allowing maintenance of high basal levels of autophagy as a key mechanism to sustain the filtration barrier. Hence, spermidine supplementation may emerge as a new therapeutic to restore autophagy in glomerular disease.