title: Peptidylarginine Deiminase 3 (PAD3) Is Upregulated by Prolactin Stimulation of CID-9 Cells and Expressed in the Lactating Mouse Mammary Gland.
abstract: Peptidylarginine deiminases (PADs) post-translationally convert arginine into neutral citrulline residues. Our past work shows that PADs are expressed in the canine and murine mammary glands; however, the mechanisms regulating PAD expression and the function of citrullination in the normal mammary gland are unclear. Therefore, the first objective herein was to investigate regulation of PAD expression in mammary epithelial cells. We first examined PAD levels in CID-9 cells, which were derived from the mammary gland of mid-pregnant mice. PAD3 expression is significantly higher than all other PAD isoforms and mediates protein citrullination in CID-9 cells. We next hypothesized that prolactin regulates PAD3 expression. To test this, CID-9 cells were stimulated with 5 μg/mL of prolactin for 48 hours which significantly increases PAD3 mRNA and protein expression. Use of a JAK2 inhibitor and a dominant negative (DN)-STAT5 adenovirus indicate that prolactin stimulation of PAD3 expression is mediated by the JAK2/STAT5 signaling pathway in CID-9 cells. In addition, the human PAD3 gene promoter is prolactin responsive in CID-9 cells. Our second objective was to investigate the expression and activity of PAD3 in the lactating mouse mammary gland. PAD3 expression in the mammary gland is highest on lactation day 9 and coincident with citrullinated proteins such as histones. Use of the PAD3 specific inhibitor, Cl4-amidine, indicates that PAD3, in part, can citrullinate proteins in L9 mammary glands. Collectively, our results show that upregulation of PAD3 is mediated by prolactin induction of the JAK2/STAT5 signaling pathway, and that PAD3 appears to citrullinate proteins during lactation.
authors.affiliation: University of Wyoming, Department of Zoology and Physiology, Laramie, WY, United States of America.
authors.forename: Guangyuan
authors.initials: G
authors.lastname: Li
authors.affiliation: University of Wyoming, Department of Zoology and Physiology, Laramie, WY, United States of America.
authors.forename: Isaac N
authors.initials: IN
authors.lastname: Hayward
authors.affiliation: University of Wyoming, Department of Zoology and Physiology, Laramie, WY, United States of America.
authors.forename: Brittany R
authors.initials: BR
authors.lastname: Jenkins
authors.affiliation: University of Wyoming, Department of Zoology and Physiology, Laramie, WY, United States of America.
authors.forename: Heather M
authors.initials: HM
authors.lastname: Rothfuss
authors.affiliation: University of Wyoming, Department of Zoology and Physiology, Laramie, WY, United States of America.
authors.forename: Coleman H
authors.initials: CH
authors.lastname: Young
authors.affiliation: Medical College of Wisconsin, Department of Pathology, Milwaukee, WI, United States of America.
authors.forename: Marja T
authors.initials: MT
authors.lastname: Nevalainen
authors.affiliation: University of Massachusetts Medical School, Department of Biochemistry and Molecular Pharmacology, Worcester, MA, United States of America.
authors.forename: Aaron
authors.initials: A
authors.lastname: Muth
authors.affiliation: University of Massachusetts Medical School, Department of Biochemistry and Molecular Pharmacology, Worcester, MA, United States of America.
authors.forename: Paul R
authors.initials: PR
authors.lastname: Thompson
authors.affiliation: University of Wyoming, Department of Zoology and Physiology, Laramie, WY, United States of America.
authors.forename: Amy M
authors.initials: AM
authors.lastname: Navratil
authors.affiliation: University of Wyoming, Department of Zoology and Physiology, Laramie, WY, United States of America.
authors.forename: Brian D
authors.initials: BD
authors.lastname: Cherrington
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D005260:Female; D006867:Hydrolases / Q000502:physiology*; D053614:Janus Kinase 2 / Q000502:physiology; D007774:Lactation / Q000502:physiology*; D008321:Mammary Glands, Animal / Q000166:cytology / Q000201:enzymology / Q000378:metabolism* / Q000502:physiology; D051379:Mice; D011247:Pregnancy; D000080063:Protein-Arginine Deiminase Type 3; D000076342:Protein-Arginine Deiminases; D050799:STAT5 Transcription Factor / Q000502:physiology; D015398:Signal Transduction / Q000502:physiology; D015854:Up-Regulation
chemical_list: D050799:STAT5 Transcription Factor; C507926:Jak2 protein, mouse; D053614:Janus Kinase 2; D006867:Hydrolases; C000609098:PAD3 protein, mouse; D000080063:Protein-Arginine Deiminase Type 3; D000076342:Protein-Arginine Deiminases