AT-Rich Interaction Domain-containing protein 5a (ARID5A; also known as Modulator Recognition Factor 1(MRF1) ) was first cloned in our laboratory by screening a cDNA library of Tera-2 mRNA with an oligonucleotide probe from the major immediate early gene of human cytomegalo virus. It has been reported that ARID5A forms a complex with androgen receptor (AR) in yeast two hybrid assays and is a cosuppressor of AR in transient transfection assays. In this study, we report that the knockdown of ARID5a expression inhibits proliferation of LNCaP cells on the contrary to the expectation from the transient transfection assay. We found that proliferation of cells stimulated by the treatment with dihydro-testosterone (DHT) was suppressed when ARID5a expression was down-regulated by siRNA technology. Androgen stimulates cell proliferation through activation of mammalian target of rapamycin complex 1 (mTORC1). However, the suppression of ARID5a does not affect androgen-activated mTOR activity. Furthermore, the expression of androgen-inducible and androgen-suppressive genes were not affected by suppression of ARID5a. We concluded that ARID5a is not a cosuppressor of AR.

Meanwhile, flow cytometer analysis showed that DHT-stimulated cell cycle was arrested at G1 phase after the knockdown of ARID5a expression. Oil Red O staining assays showed that the inhibition of ARID5a expression led to decreased DHT-induced lipid accumulation. Our data further proved that the knockdown of ARID5a expression inhibited the protein expression of DHT-induced sterol regulatory element binding transcription factor 1 (SREBP-1). In addition, Western blot analysis revealed that the down-regulation of ARID5a decreased the expression of cell survival / proliferation and cell cycle regulation proteins, such as hypoxia inducible factor 1 alpha (HIF1a), cyclin D1 and cyclin D3. Real time PCR data revealed that the knockdown of ARID5a expression did not affect the steady level of HIF1a, cyclin D1 and cyclin D3 mRNA. These suggested that knockdown of ARID5a expression affected protein expression at translational and/or post-translational level. Our data suggested that knockdown of ARID5a expression increased the phosphorylation of eukaryotic translation initiation factor 2a (eIF2a) which inhibited translation initiation. S35 labeling assay confirmed that the knockdown of ARID5a expression suppressed the global protein synthesis. We

showed that one of the four eIF2a kinases, general control non-derepressible 2 (GCN2) was activated by the suppression of ARID5a expression. We are currently studying on the mechanisms by which ARID5A regulates GCN2 kinase.

Citation Format: Chao Sun, Viktor Chesnokov, Keiichi Itakura. Knockdown ARID5A suppresses proliferation of LNCaP prostate cancer cell through the inhibition of global protein synthesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4489. doi:10.1158/1538-7445.AM2017-4489