NRIP1 regulates cell proliferation in lung adenocarcinoma cells.

Nuclear receptor interacting protein 1 (NRIP1) is a transcription cofactor that regulates the activity of nuclear receptors and transcription factors. Functional expression of NRIP1 has been identified in multiple cancers. However, the expression and function of NRIP1 in lung adenocarcinoma have remained unclear. Thus, we aimed to clarify the NRIP1 expression and its functions in lung adenocarcinoma cells. NRIP1 and Ki-67 were immunostained in the tissue microarray section consisting of 64 lung adenocarcinoma cases, and the association of NRIP1 immunoreactivity with clinical phenotypes was examined. Survival analysis was performed in lung adenocarcinoma data from The Cancer Genome Atlas (TCGA). Human A549 lung adenocarcinoma cell line with an NRIP1-silencing technique was used in vitro study. Forty-three of 64 cases were immunostained with NRIP1. Ki-67-positive cases were more frequent in NRIP1-positive cases as opposed to NRIP1-negative cases. Higher NRIP1 mRNA expression was associated with poor prognosis in the TCGA lung adenocarcinoma data. NRIP1 was mainly located in the nucleus of A549 cells. NRIP1 silencing significantly reduced the number of living cells, suppressed cell proliferation, and induced apoptosis. These results suggest that NRIP1 participates in the progression and development of lung adenocarcinoma. Targeting NRIP1 may be a possible therapeutic strategy against lung adenocarcinoma.

Elevated (Pro)renin Receptor Expression by Anti-Cancer Drugs, Carboplatin and Paclitaxel, in Cultured Cancer Cells: Possible Involvement of Apoptosis and Autophagy.

(Pro)renin receptor [(P)RR] is a component of the renin-angiotensin system and plays an essential role in the activity of vacuolar H+-ATPase and autophagy. (P)RR is expressed in cancer cells. However, the relationship among (P)RR, apoptosis and autophagy in the treatment of anti-cancer drugs has not been clarified. The aim of this study was to clarify the effects of anti-cancer drugs with autophagy-promoting activity on (P)RR expression in cancer cells. MCF-7 breast cancer cells and A549 lung cancer cells were treated with carboplatin or paclitaxel, and the expression of (P)RR, apoptosis markers and autophagy markers were assessed by RT-qPCR, western blot analysis and immunocytochemistry. Expression levels of (P)RR mRNA and soluble (P)RR protein were increased by carboplatin or paclitaxel in a dose-dependent manner. Immunofluorescence staining of (P)RR was increased in both MCF-7 and A549 cells treated by carboplatin or paclitaxel. Apoptosis induction was shown by elevated BAX/BCL2 mRNA levels and increased active caspase3-positive cells. Moreover, autophagy induction was confirmed by increased levels of autophagy-associated mRNAs and LC3B-II proteins. (P)RR knockdown by (P)RR-specific siRNA suppressed the cell viability in MCF-7 cells and A549 cells under the treatment of carboplatin or paclitaxel, suggesting that (P)RR deficiency inhibits the proliferation of cancer cells in a pathway different from carboplatin or paclitaxel. The present study showed that the expression of (P)RR mRNA and soluble (P)RR was increased by anti-cancer drugs with autophagy-promoting activity. Upregulated (P)RR and autophagy may constitute a stress adaptation that protects cancer cells from apoptosis.

(Pro)renin receptor/ATP6AP2 is required for autophagy and regulates proliferation in lung adenocarcinoma cells.

(Pro)renin receptor ((P)RR)/ ATP6AP2 (ATPase, H+ transporting, lysosomal accessory protein 2) functions as an essential accessory subunit of vacuolar H+ -ATPase (V-ATPase). V-ATPase is necessary for lysosome function and autophagy. Autophagy is related to cell proliferation, migration and invasion of various cancer cells. In this study, we aim to clarify the relationship between (P)RR and autophagy in lung adenocarcinoma. Expression of (P)RR and Ki-67 (a proliferation marker) was studied in sixty-four adenocarcinoma cases by immunohistochemistry. Lung adenocarcinoma cell line, A549, was transfected with (P)RR-specific siRNA. Autophagy inhibitors, bafilomycin A1 and chloroquine, were used as positive controls. Cell proliferation and migration were measured by WST-8 assay and wound healing assay. Autophagosome markers, p62 and LC3, were analyzed by RT-qPCR, Western blot and immunocytochemistry. Immunohistochemistry showed that (P)RR was expressed in all adenocarcinoma tissues. The intensity of (P)RR immunoreactivity was significantly associated with Ki-67. Treatment of (P)RR-specific siRNA suppressed (P)RR expression and significantly reduced cell proliferation and migration as did the autophagy inhibitors. Western blot and immunocytochemistry showed that (P)RR-specific siRNA, as well as the autophagy inhibitors, induced p62 and LC3 accumulation in cytoplasmic granules. These results suggest that (P)RR is involved in cell proliferation and progression of lung adenocarcinoma via regulating autophagy.

Increased soluble (pro)renin receptor protein by autophagy inhibition in cultured cancer cells.

(Pro)renin receptor ((P)RR) regulates the renin-angiotensin system and functions as an essential accessory subunit of vacuolar H+ -ATPase. There is accumulating evidence that shows close relationship between (P)RR and autophagy. Soluble (P)RR consisting of the extracellular domain of (P)RR is generated from (P)RR by proteolytic enzymes. The aim of the present study was to clarify the influence of autophagy inhibition on soluble (P)RR expression in cancer cells. Autophagy was inhibited by treatment of bafilomycin A1 or chloroquine in MCF-7 and A549 cells for 72 hr. Western blot analysis showed that protein levels of soluble (P)RR were markedly elevated by autophagy inhibition, whereas no noticeable increases were observed in full-length (P)RR. Secretion of soluble (P)RR into the medium was increased dose-dependently by bafilomycin A1 or chloroquine. Autophagy inhibition was confirmed by enhanced accumulation of autophagy-related proteins, LC3, p62 and LAMP1 in intracellular vesicles. Increased amount of soluble (P)RR by autophagy inhibition was decreased by site-1 protease inhibitor, whereas no noticeable increase in site-1 protease immunoreactivity was observed in cells with autophagy inhibition by immunocytochemistry. These findings suggest that soluble (P)RR protein accumulates by autophagy inhibition, possibly because of the reduced degradation of soluble (P)RR in the intracellular vesicles during autophagy inhibition.