No abstract

No abstract

Kidney fibrosis is a common pathway that leads to chronic kidney disease. Angiotensin II type-1 receptor (AT1R)-associated protein (ATRAP) was originally identified as an AT1R-binding protein. Previously, we reported that systemic knockout of ATRAP exacerbates kidney fibrosis in aged mice. Although these effects of ATRAP appeared to be AT1R-independent actions, the molecular mechanism remains poorly understood. To elucidate the molecular mechanism of ATRAP independent of AT1R, we explored novel ATRAP-interacting proteins. Mass spectrometric analysis of the immunoprecipitants of a Flag-tagged ATRAP complex revealed 376 candidate proteins that potentially interact with ATRAP. Gene ontology analysis revealed that proteins related to vesicle trafficking, membrane transport, and many membrane proteins, including transferrin receptor 1 (TfR1), were enriched. Because TfR1 promotes cellular iron uptake and iron is a key factor involved in kidney fibrosis, we focused on TfR1 and confirmed that it interacts with ATRAP. In addition, our findings revealed that enhanced ATRAP expression decreased cell-surface TfR1 expression without altering the overall cellular TfR1 expression levels. Furthermore, enhanced ATRAP expression attenuated cellular iron levels. Together, our results highlight the role of ATRAP as a suppressor of TfR1 that functions by facilitating TfR1 internalization, which affects iron metabolism and oxidative stress signaling.

Objective: We previously reported that genetic knockdown of angiotensin II type 1 receptor (AT1R)-associated protein (ATRAP) exacerbated the aging-associated kidney tubulointerstitial fibrosis in mice (Uneda K, et al. J Am Heart Assoc 2017). However, little is known about whether enhancement of ATRAP expression could affect any pathological stimuli-induced kidney fibrosis and inflammation. Recently we proposed that aristolochic acid nephropathy (AAN) might be a useful model of kidney aging along with tubulointerstitial fibrosis (Urate S, et al. Int J Mol Sci 2021). The present study was designed to investigate the functional role of ATRAP in kidney fibrosis and inflammation, using ATRAP transgenic mice subjected to AAN. Design and method:We generated ATRAP transgenic (Tg19) mice under the control of chicken b;-actin promoter. (Wakui H, et al. Am J Physiol Renal Physiol 2010). The level of kidney ATRAP protein expression was about 4-fold higher in the Tg19 mice compared with wild-type littermate control (LC) mice. The Tg19 mice and LC mice were administered either vehicle or aristolochic acid (AA) (3 mg/kg) for 4 weeks, followed by a 4-week remodeling period. Kidney fibrosis was examined by histopathology and macrophage infiltration was determined by immunohistochemistry. Expression levels of genes associated with inflammation, fibrosis, and senescence were determined by quantitative reverse transcriptionpolymerase chain reaction (qRT-PCR) and immuno blot analyses.Results: AA administration provoked kidney fibrosis and inflammation accompanied by decreased kidney function. There were no significant differences in AAinduced kidney fibrosis estimated by qRT-PCR and histological analysis between the Tg19 and LC mice. Creatinine clearance was similarly decreased in the Tg19 and LC mice with AAN. However, AA-induced macrophage infiltration was significantly suppressed in the kidney of Tg19 mice compared with that of the LC mice. In addition, inflammation-, aging-, and oxidative stress-related genes (tumor necrosis factor-a, interleukin-1b;, cyclin-dependent kinase inhibitor 2A, wnt family member 9A, nicotinamide adenine dinucleotide phosphate oxidase 2) in response to AA administration were significantly suppressed in the kidney of the Tg19 mice compared with that of the LC mice. Conclusion:These results indicate that the enhancement of ATRAP expression suppresses kidney inflammation despite no evident effects on kidney fibrosis in a mouse model of AAN. ATRAP may be a therapeutic target for inflammation associated with CKD.

Kidney fibrosis is a common pathway that leads to chronic kidney disease. Angiotensin II type-1 receptor (AT1R)-associated protein (ATRAP) was originally identified as an AT1R-binding protein. Previously, we reported that systemic knockout of ATRAP exacerbates kidney fibrosis in aged mice. Although these effects of ATRAP appeared to be AT1R-independent actions, the molecular mechanism remains poorly understood. To elucidate the molecular mechanism of ATRAP independent of AT1R, we explored novel ATRAP-interacting proteins. Mass spectrometric analysis of the immunoprecipitants of a Flag-tagged ATRAP complex revealed 376 candidate proteins that potentially interact with ATRAP. Gene ontology analysis revealed that proteins related to vesicle trafficking, membrane transport, and many membrane proteins, including transferrin receptor 1 (TfR1), were enriched. Because TfR1 promotes cellular iron uptake and iron is a key factor involved in kidney fibrosis, we focused on TfR1 and confirmed that it interacts with ATRAP. In addition, our findings revealed that enhanced ATRAP expression decreased cell-surface TfR1 expression without altering the overall cellular TfR1 expression levels. Furthermore, enhanced ATRAP expression attenuated cellular iron levels. Together, our results highlight the role of ATRAP as a suppressor of TfR1 that functions by facilitating TfR1 internalization, which affects iron metabolism.