HOXA3 activates USP15 to suppress autophagy and promote M2-type macrophage polarization in renal cell carcinoma via facilitating the deubiquitination of SQSTM1.

The disease burden of renal cell carcinoma (RCC) has decreased in recent years with advances in treatment, but its pathogeny still remains elusive. We aim to study the role of homeobox A3 (HOXA3)/ubiquitin-specific peptidase 15 (USP15)/SQSTM1 axis on autophagy and M2-type macrophage polarization in RCC. In this study, cell apoptosis and proliferation were assessed by flow cytometry and CCK-8. Autolysosome fusion was observed by immunofluorescence detection of LC3 and LAMP2. The binding between HOXA3 and USP15 promoter was tested by chromatin immunoprecipitation (ChIP), EMSA, and dual-luciferase reporter assays. Also, the interaction between deubiquitinated enzyme (DUB) USP15 and SQSTM1, and ubiquitinated level of SQSTM1 were determined by co-immunoprecipitation (Co-IP) assay. Expression levels of HOXA3, USP15, C-C motif chemokine 2 (CCL2), CCL2 receptor (CCR2), M2-type macrophages, and autophagy-related markers were measured by Western blot, quantitative reverse transcription PCR (RT-qPCR), ELISA, and immunohistochemistry. Role of HOXA3/USP15 axis was verified by xenograft tumor experiment in vivo. We showed upregulated HOXA3 in RCC tissues and cells, and RCC tissues with metastasis showed higher HOXA3 level. The higher HOXA3 expression was relevant to worse overall survival in patients with RCC. HOXA3 induced RCC cell proliferation, and suppressed autophagy and apoptosis via transcriptionally activating USP15 expression. USP15 then induced deubiquitination modification of SQSTM1 in RCC cells. SQSTM1 supported M2-type macrophage polarization by inducing CCL2 secretion. HOXA3 or USP15 knockdown suppressed tumor growth and M2-type macrophage infiltration in vivo. In conclusion, HOXA3 transcriptionally activates USP15 expression, and upregulated USP15 facilitates the deubiquitination of SQSTM1 in RCC. This process on the one hand suppresses autophagy, on the other hand increases M2-type macrophage polarization through stimulating the secretion of CCL2. We report a novel finding that highly expressed homeobox A3 (HOXA3) transcriptionally activates the expression of ubiquitin-specific peptidase 15 (USP15), resulting in the promotion of deubiquitination of SQSTM1. This process on the one hand suppresses autophagy in renal cell carcinoma (RCC), on the other hand increases M2-type macrophage polarization in the tumor microenvironment through stimulating the secretion of C-C motif chemokine 2 (CCL2).