ING5 differentially regulates protein lysine acetylation and promotes p300 autoacetylation

// Tao Zhang 1, * , Jin Meng 2, 3, * , Xinli Liu 2 , Xutao Zhang 2 , Xiaojun Peng 4 , Zhongyi Cheng 4 and Feng Zhang 2 1 Department of Thoracic Surgery, Tangdu Hospital, Fourth Military Medical University, Xi’an 710038, China 2 Department of Pharmacology, Key Laboratory of Gastrointestinal Pharmacology of Chinese Materia Medica of the State Administration of Traditional Chinese Medicine, School of Pharmacy, Fourth Military Medical University, Xi’an 710032, China 3 Department of Pharmacy, No. 309 Hospital of PLA, Beijing 100091, China 4 Department of Bioinformatics, Jingjie PTM Biolab (Hangzhou) Co. Ltd, Hangzhou 310018, China * These authors contributed equally to this work Correspondence to: Tao Zhang, email: zhangft@fmmu.edu.cn Feng Zhang, email: zhangf037@163.com Keywords: inhibitor of growth 5 (ING5); quantitative proteomics; acetylation; p300 autoacetylation; lung cancer Received: November 30, 2016      Accepted: October 11, 2017      Published: October 31, 2017 ABSTRACT ING5 belongs to the Inhibitor of Growth (ING) candidate tumor suppressor family. Previously, we have shown that ING5 inhibits invasiveness of lung cancer cells by downregulating EMT-inducing genes. However, the underlying mechanisms remain unclear. The aim of the study was to use integrated approach involving SILAC labeling and mass spectrometry-based quantitative proteomics to quantify dynamic changes of acetylation regulated by ING5 in lung cancer cells. Here, we have found that ING5 has a profound influence on protein lysine acetylation with 163 acetylation peptides on 122 proteins significantly upregulated and 100 acetylation peptides on 72 proteins downregulated by ING5 overexpression. Bioinfomatic analysis revealed that the acetylated proteins upregulated by ING5 located preferentially in nucleus to cytoplasm and were significantly enriched in transcription cofactor activity, chromatin binding and DNA binding functions; while those downregulated by ING5 located preferentially in cytoplasm rather than nucleus and were functionally enriched in metabolism, suggesting diverse functions of ING5 through differentially regulating protein acetylation. Interestingly, we found ING5 overexpression promotes p300 autoacetylation at K1555, K1558 and K1560 within p300 HAT domain, and two novel sites K1647 and K1794, leading to activation of p300 HAT activity, which was confirmed by accelerated acetylation of p300 target proteins, p53 at k382 and histone H3 at K18. A specific p300 HAT inhibitor C646 impaired ING5-increased acetylation of H3K18 and p53K382, and subsequent expression of p21 and Bax. In conclusion, our results reveal the lysine acetylome regulated by ING5 and provide new insights into mechanisms of ING5 in the regulation of gene expression, metabolism and other cellular functions.