Potential Role for PAD2 in Gene Regulation in Breast Cancer Cells

Abstract

The peptidylarginine deiminase (PAD) family of enzymes post-translationally convert positively charged arginine residues in substrate proteins to the neutral, non-standard residue citrulline. PAD family members 1, 2, 3, and 6 have previously been localized to the cell cytoplasm and, thus, their potential to regulate gene activity has not been described. We recently demonstrated that PAD2 is expressed in the canine mammary gland epithelium and that levels of histone citrullination in this tissue correlate with PAD2 expression. Given these observations, we decided to test whether PAD2 might localize to the nuclear compartment of the human mammary epithelium and regulate gene activity in these cells. Here we show, for the first time, that PAD2 is specifically expressed in human mammary gland epithelial cells and that a portion of PAD2 associates with chromatin in MCF-7 breast cancer cells. We investigated a potential nuclear function for PAD2 by microarray, qPCR, and chromatin immunoprecipitation analysis. Results show that the expression of a unique subset of genes is disregulated following depletion of PAD2 from MCF-7 cells. Further, ChIP analysis of two of the most highly up- and down-regulated genes (PTN and MAGEA12, respectively) found that PAD2 binds directly to these gene promoters and that the likely mechanism by which PAD2 regulates expression of these genes is via citrullination of arginine residues 2–8–17 on histone H3 tails. Thus, our findings define a novel role for PAD2 in gene expression in human mammary epithelial cells.

Downregulation of the Deiminase PADI2 Is an Early Event in Colorectal Carcinogenesis and Indicates Poor Prognosis

“…2D). A similar expression of PADI2 was previously described in other epithelial cells from kidney, endometrium, and mammary gland (28).…”

PAD2 Overexpression in Transgenic Mice Promotes Spontaneous Skin Neoplasia

“…In addition, we found strong nuclear anti-pan-citrulline staining in these lesions (Fig. 2D, inset bottom image), which is in line with our previous studies showing that PAD2-mediated histone deimination regulates gene expression (9,10,14,21). Because the anti-human PAD2 antibody likely cross-reacts with endogenous mouse PAD2, we also stained these tissues with an anti-FLAG antibody to specifically stain the PAD2 fusion protein.…”

“…Therefore, while purely speculative, it is possible that the relatively low levels of FLAG-PAD2 that we observed in the skin promotes the expression of a specific ratio of PAD family members that is conducive to inflammation and tumorigenesis, whereas the relatively higher level of FLAG-PAD2 in the mammary gland alters this ratio, leading to reduced inflammatory mediator expression and an absence of neoplastic growth. Regarding the potential mechanisms by which PAD2 overexpression regulates gene activity, we have recently found that PAD2 functions as a transcription factor co-factor and modulates target gene expression via histone deimination at transcription factor-binding sites (9,10,14,21). Therefore, we are currently testing the hypothesis that PAD2 regulates the expression of target genes via similar mechanisms.…”

“…While the presumptive function of PAD activity in most diseases is linked to inflammation, the role that PADs play in cancer progression is still under investigation (4)(5)(6)(7). Our recent studies suggest a role for PAD2 in the oncogenic progression of breast cancer (8)(9)(10)(11) and provide preclinical evidence showing that the PAD inhibitor, Cl-amidine, could be used as a therapeutic agent for the treatment of tumors in vivo (11). To further investigate the involvement of PAD2 in the oncogenesis of epithelial tumors, we generated FVB/N mice expressing FLAG-PAD2 under the control of the mammary tumor virus (MMTV) promoter.…”

Origins of cancer symposium 2015: posttranslational modifications and cancer

“…The deregulation of PAD enzymes is involved in cancers and autoimmune disorders [9]. Dr. Coonrod's lab has shown that PAD2 expression is up-regulated in tamoxifen-resistant breast cancer cells [10]. While they predicted that PAD2 might be a driver of breast cancer, unexpectedly, overexpression of PAD2 in mice by the murine mammary tumor virus promoter led to dysplastic tumors in skin cells, but not in mammary glands [11].…”