Mass Spectrometric Analysis of High-Mobility Group Proteins and Their Post-Translational Modifications in Normal and Cancerous Human Breast Tissues

Abstract

High-mobility group (HMG) A1 proteins including HMGA1a and HMGA1b are chromosomal proteins that function in a variety of cellular processes such as cell growth, transcription regulation, neoplastic transformation, and progression. Overexpression of HMGA1 proteins has been associated with almost every type of cancer cells. Post-translational modifications (PTMs) of HMGA1 proteins in different types of human cancer cell lines have been extensively explored over the past decade. Here, we extended the identification of PTMs of HMGA1 proteins to human breast tumor tissue specimens with different carcinoma progression stages (metastatic and primary cancer) as well as the paired adjacent normal breast tissues. In this regard, we employed tandem mass spectrometry to examine the nature and sites of PTMs of HMGA1 proteins isolated from cancerous/normal human breast tissues. Novel PTMs of HMGA1a protein, that is, monomethylation at Lys30 and Lys54 as well as monophosphorylation at Ser43 and Ser48, were detected in cancer tissues. In these cancer tissues, we also found C-terminal constitutive phosphorylation in HMGA1a and HMGA1b as well as mono- and dimethylation of Arg25 in HMGA1a, which were previously found to be present in these proteins isolated from human cancer cell lines. Furthermore, a more complex spectrum of PTMs on HMGA1 proteins was correlated with a more aggressive malignancy in human breast cancer tissues.