Correction: Zhu et al. A Feedback Loop Formed by ATG7/Autophagy, FOXO3a/miR-145 and PD-L1 Regulates Stem-like Properties and Invasion in Human Bladder Cancer. Cancers 2019, 11, 349.

In the original article [...].

A Feedback Loop Formed by ATG7/Autophagy, FOXO3a/miR-145 and PD-L1 Regulates Stem-Like Properties and Invasion in Human Bladder Cancer.

Programmed cell death protein 1 (PD-1) and its ligand PD-L1 blockade have been identified to target immune checkpoints to treat human cancers with durable clinical benefit. Several studies reveal that the response to PD-1-PD-L1 blockade might correlate with PD-L1 expression levels in tumor cells. However, the mechanistic pathways that regulate PD-L1 protein expression are not understood. Here, we reported that PD-L1 protein is regulated by ATG7-autophagy with an ATG7-initiated positive feedback loop in bladder cancer (BC). Mechanistic studies revealed that ATG7 overexpression elevates PD-L1 protein level mainly through promoting autophagy-mediated degradation of FOXO3a, thereby inhibiting its initiated miR-145 transcription. The lower expression of miR-145 increases pd-l1 mRNA stability due to the reduction of its direct binding to 3'-UTR of pd-l1 mRNA, in turn leading to increasing in pd-l1 mRNA stability and expression, and finally enhancing stem-like property and invasion of BC cells. Notably, overexpression of PD-L1 in ATG7 knockdown cells can reverse the defect of autophagy activation, FOXO3A degradation, and miR-145 transcription attenuation. Collectively, our results revealed a positive feedback loop to promoting PD-L1 expression in human BC cells. Our study uncovers a novel molecular mechanism for regulating pd-l1 mRNA stability and expression via ATG7/autophagy/FOXO3A/miR-145 axis and reveals the potential for using combination treatment with autophagy inhibitors and PD-1/PD-L1 immune checkpoint blockade to enhance therapeutic efficacy for human BCs.

CD44s is a crucial ATG7 downstream regulator for stem-like property, invasion, and lung metastasis of human bladder cancer (BC) cells.

Over half a million US residents are suffering with bladder cancer (BC), which costs a total $4 billion in treatment annually. Although recent studies report that autophagy-related gene 7 (ATG7) is overexpressed in BCs, the regulatory effects of ATG7 on cancer stem-like phenotypes and invasion have not been explored yet. Current studies demonstrated that the deficiency of ATG7 by its shRNA dramatically reduced sphere formation and invasion in vitro, as well as lung metastasis in vivo in human invasive BC cells. Further studies indicated that the knockdown of ATG7 attenuated the expression of CD44 standard (CD44s), while ectopic introduction of CD44s, was capable of completely restoring sphere formation, invasion, and lung metastasis in T24T(shATG7) cells. Mechanistic studies revealed that ATG7 overexpression stabilized CD44s proteins accompanied with upregulating USP28 proteins. Upregulated USP28 was able to bind to CD44s and remove the ubiquitin group from CD44s' protein, resulting in the stabilization of CD44s protein. Moreover, ATG7 inhibition stabilized AUF1 protein and thereby reduced tet1 mRNA stability and expression, which was able to demethylate usp28 promoter, reduced USP28 expression, finally promoting CD44s degradation. In addition, CD44s was defined to inhibit degradation of RhoGDIß, which in turn promotes BC invasion. Our results demonstrate that CD44s is a key ATG7 downstream regulator of the sphere formation, invasion, and lung metastasis of BCs, providing significant insight into understanding the BC invasions, metastasis, and stem-like properties.

Decreased c-Myc mRNA Stability via the MicroRNA 141-3p/AUF1 Axis Is Crucial for p63α Inhibition of Cyclin D1 Gene Transcription and Bladder Cancer Cell Tumorigenicity.

Bladder cancer (BC) ranks as the sixth most common cancer in the United States and is the leading cause of death in patients with urinary malignancies. p63 is a member of the p53 family and is believed to function as a tumor suppressor in human BCs. Our most recent studies revealed a previously unknown function of the RING of XIAP in promoting microRNA 4295 (miR-4295) transcription, thereby reducing p63α protein translation and enhancing normal urothelial transformation, whereas p63α upregulates hsp70 transcription, subsequently activating the HSP70/Wasf3/Wave3/matrix metalloproteinase 9 (MMP-9) axis and promoting BC cell invasion via initiating the transcription factor E2F1. In this study, we found that p63α inhibited cyclin D1 protein expression, subsequently decreasing the ability of BC cell anchorage-independent growth in vitro and tumorigenicity in vivo Mechanistic studies demonstrated that p63α expression is able to downregulate cyclin D1 gene transcription through attenuation of c-Myc mRNA stability. We further show that the reduction of miR-141-3p expression by p63α directly releases its inhibition of 3' untranslated region (UTR) activity of AU-rich element RNA-binding factor 1 (AUF1) mRNA, thereby increasing AUF1 protein translation and further resulting in degradation of c-Myc mRNA, which, in turn, reduces cyclin D1 gene transcription and BC cell anchorage-independent growth. Collectively, our results demonstrate that p63α is a negative regulator of BC cell tumorigenic growth, a distinctly different function than its promotion of BC invasion, thus providing further new insight into the "two faces" of p63α in regulation of BC cell tumorigenic growth and progression/invasion.

MicroRNA-411 Downregulation Enhances Tumor Growth by Upregulating MLLT11 Expression in Human Bladder Cancer.

Although several previous studies have reported the implication of various microRNAs (miRNAs) in regulation of human bladder cancer (BC) development, alterations and function of many miRNAs in bladder cancer growth are not explored yet at present. Here, we screened 1,900 known miRNAs and first discovered that miR-411 was one of the major miRNAs, which was down-regulated in n-butyl-N-(4-hydroxybutyl)-nitrosamine (BBN)-induced BCs. This miR-411 down-regulation was also observed in human BC tissues and cell lines. The results from evaluating the relationship between miR-411 and patient survival in BC using the TCGA (The Cancer Genome Atlas) database indicated that miR-411 was positively correlated with DFS (disease-free survival). Our studies also showed that miR-411 inhibited tumor growth of human BC cells in a xenograft animal model. Mechanistic studies revealed that overexpression of miR-411 repressed the expression of ALL1-fused gene from the chromosome 1q (AF1q) (MLLT11) by binding to the 3' untranslated region (UTR) of mllt11 mRNA and in turn induced p21 expression and caused cell cycle arrest at the G2/M phase, further inhibiting BC tumor growth. Collectively, our results improve our understanding of the role of miR-411 in BC tumor growth and suggest miR-411 and MLLT11 as potential new targets for the treatment of BC patients.