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.
Highlights
Lysine acetylation is a reversible and precisely regulated posttranslational modification (PTM) which plays a key role in a wide range of cellular processes such as gene transcription, DNA binding, cell metabolism and signal transduction
By SILAC labeling and mass spectrometry-based quantitative proteomics, we have identified acetylome affected by ING5 overexpression, which reveal that ING5 differentially regulates lysine acetylation of proteins involved in DNA binding and metabolism
ING5 has been identified as a component of HBO1 and MOZ/MORF histone acetyltransferases www.impactjournals.com/oncotarget (HATs) complexes [11] and associates with p300 HAT [12], may participate in regulation of protein acetylation, chromatin remodeling and gene expression
Summary
Lysine acetylation is a reversible and precisely regulated posttranslational modification (PTM) which plays a key role in a wide range of cellular processes such as gene transcription, DNA binding, cell metabolism and signal transduction. Deregulation of protein acetylation has been associated with several diseases, especially cancer [1,2,3]. The acetylation of histones and non-histone proteins is catalyzed by histone acetyltransferases www.impactjournals.com/oncotarget (HATs), which transfer the acetyl group from acetylCoA to the ε-amino group of a lysine residue. The reverse reaction is catalyzed by histone deacetylases (HDACs). A growing number of cellular proteins have been identified to be acetylated by proteomic studies [4]. The functions and regulating mechanisms of this modification in diverse cellular proteins remain largely unknown
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