Identification and Characterization of the Lactating Mouse Mammary Gland Citrullinome
Young, Coleman H.
Christensen, Amanda O.
Demoruelle, M. Kristen
Nemmara, Venkatesh V.
Thompson, Paul R.
Rothfuss, Heather M.
Cherrington, Brian D.
UMass Chan AffiliationsThompson Lab
Department of Biochemistry and Molecular Pharmacology
Amino Acids, Peptides, and Proteins
Biochemical Phenomena, Metabolism, and Nutrition
Enzymes and Coenzymes
Nucleic Acids, Nucleotides, and Nucleosides
Reproductive and Urinary Physiology
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AbstractCitrullination is a post-translational modification (PTM) in which positively charged peptidyl-arginine is converted into neutral peptidyl-citrulline by peptidylarginine deiminase (PAD or PADI) enzymes. The full protein citrullinome in many tissues is unknown. Herein, we used mass spectrometry and identified 107 citrullinated proteins in the lactation day 9 (L9) mouse mammary gland including histone H2A, alpha-tubulin, and beta-casein. Given the importance of prolactin to lactation, we next tested if it stimulates PAD-catalyzed citrullination using mouse mammary epithelial CID-9 cells. Stimulation of CID-9 cells with 5 microg/mL prolactin for 10 min induced a 2-fold increase in histone H2A citrullination and a 4.5-fold increase in alpha-tubulin citrullination. We next investigated if prolactin-induced citrullination regulates the expression of lactation genes beta-casein (Csn2) and butyrophilin (Btn1a1). Prolactin treatment for 12 h increased beta-casein and butyrophilin mRNA expression; however, this increase was significantly inhibited by the pan-PAD inhibitor, BB-Cl-amidine (BB-ClA). We also examined the effect of tubulin citrullination on the overall polymerization rate of microtubules. Our results show that citrullinated tubulin had a higher maximum overall polymerization rate. Our work suggests that protein citrullination is an important PTM that regulates gene expression and microtubule dynamics in mammary epithelial cells.
Li G, Young CH, Snow B, Christensen AO, Demoruelle MK, Nemmara VV, Thompson PR, Rothfuss HM, Cherrington BD. Identification and Characterization of the Lactating Mouse Mammary Gland Citrullinome. Int J Mol Sci. 2020 Apr 10;21(7):2634. doi: 10.3390/ijms21072634. PMID: 32290104; PMCID: PMC7177251. Link to article on publisher's site