Potential role for PADI-mediated histone citrullination in preimplantation development.
BACKGROUND
The peptidylarginine deiminases (PADIs) convert positively charged arginine residues to neutrally charged citrulline on protein substrates in a process that is known as citrullination or deimination. Previous reports have documented roles for histone citrullination in chromatin remodeling and gene regulation in several tissue types, however, a potential role for histone citrullination in chromatin-based activities during early embryogenesis has not been investigated.


RESULTS
In the present study, we tested by laser scanning confocal indirect immunofluorescence microscopy whether specific arginine residues on the histone H3 and H4 N-terminal tails (H4R3, H3R2 + 8 + 17, and H3R26) were citrullinated in mouse oocytes and preimplantation embryos. Results showed that all of the tested residues were deiminated with each site showing a unique localization pattern during early development. Given these findings, we next tested whether inhibition of PADI activity using the PADI-specific inhibitor, Cl-amidine, may affect embryonic development. We found that treatment of pronuclear stage zygotes with Cl-amidine reduces both histone H3 and H4 tail citrullination and also potently blocks early cleavage divisions in vitro. Additionally, we found that the Cl-amidine treatment reduces acetylation at histone H3K9, H3K18, and H4K5 while having no apparent effect on the repressive histone H3K9 dimethylation modification. Lastly, we found that treatment of zygotes with trichostatin A (TSA) to induce hyperacetylation also resulted in an increase in histone citrullination at H3R2 + 8 + 17.


CONCLUSIONS
Given the observed effects of Cl-amidine on embryonic development and the well documented correlation between histone acetylation and transcriptional activation, our findings suggest that histone citrullination may play an important role in facilitating gene expression in early embryos by creating a chromatin environment that is permissive for histone acetylation.
Baker Institute for Animal Health, Cornell University, Ithaca, NY 14850, USA.
Rui
R
Kan
Mei
M
Jin
Venkataraman
V
Subramanian
Corey P
CP
Causey
Paul R
PR
Thompson
Scott A
SA
Coonrod
D000818:Animals; D001755:Blastocyst / Q000187:drug effects / Q000378:metabolism*; D002956:Citrulline / Q000378:metabolism*; D047108:Embryonic Development / Q000187:drug effects; D005260:Female; D006657:Histones / Q000378:metabolism*; D006867:Hydrolases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D006877:Hydroxamic Acids / Q000494:pharmacology; D051379:Mice; D008810:Mice, Inbred C57BL; D008815:Mice, Inbred Strains; D009865:Oocytes / Q000187:drug effects / Q000378:metabolism; D009952:Ornithine / Q000031:analogs & derivatives / Q000494:pharmacology; D000080002:Protein-Arginine Deiminase Type 4; D000080065:Protein-Arginine Deiminase Type 6; D000076342:Protein-Arginine Deiminases
D002956:Citrulline; D006657:Histones; D006867:Hydrolases; D006877:Hydroxamic Acids; D009952:Ornithine; D000080002:Protein-Arginine Deiminase Type 4; D000080065:Protein-Arginine Deiminase Type 6; D000076342:Protein-Arginine Deiminases; C558727:N-alpha-benzoyl-N5-(2-chloro-1-iminoethyl)-L-ornithine amide; C012589:trichostatin A; C523190:PADI6 protein, mouse; C552151:peptidylarginine deiminase 4, mouse