Transcriptomic analysis of PADI4 target genes during multi-lineage differentiation of embryonic stem cells

Abstract

During mammalian embryo development, pluripotent epiblast cells diversify into the three primary germ layers, which will later give rise to all fetal and adult tissues. These processes involve profound transcriptional and epigenetic changes that require precise coordination. Peptidylarginine deiminase IV (PADI4) is a transcriptional regulator that is strongly associated with inflammation and carcinogenesis but whose physiological roles are less well understood. We previously found that Padi4 expression is associated with pluripotency. Here, we examined the role of PADI4 in maintaining the multi-lineage differentiation potential of mouse embryonic stem (ES) cells. Using bulk and single-cell transcriptomic analyses of embryoid bodies (EBs) derived from Padi4 knock-out ( Padi4-KO ) mouse ES cells, we find that PADI4 loss impairs mesoderm diversification and differentiation of cardimyocytes and endothelial cells. Additionally, Padi4 deletion leads to concerted downregulation of genes associated with polarized growth, sterol metabolism and the extracellular matrix (ECM). This study indicates a requirement for Padi4 in the specification of the mesodermal lineage and reports the Padi4 associated transcriptome, providing a platform for understanding the physiological functions of Padi4 in development and homeostasis. This article is part of the Theo Murphy meeting issue ‘The virtues and vices of protein citrullination’.

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“…The divergent outcomes in antigen presentation between cells treated with GSK484 or Cl-Amidine and those from animals lacking PAD4 may be attributed to off-target effects of the drugs. Alternatively, the high level of PAD4 expression in mouse bone marrow cells, its role in hematopoiesis [62], and its involvement in the differentiation of other cell types, such as cardiomyocytes and endothelial cells [63], might suggest a role for PAD4 in the differentiation of APCs. This hypothetical impact could potentially drive the differentiation of APCs lacking PAD4 towards subsets with increased MHC expression and enhanced capacity for antigen presentation [64][65][66].…”

Citrullination: new tricks for an old mod

“…Expanding on our understanding of the roles of PADIs in cell differentiation, a study by the Christophorou laboratory probes the role of PADI4 in the regulation of multi-lineage differentiation potential of embryonic stem cells (Singh et al [ 7 ]). PADI4 has been implicated in the regulation of pluripotency, in embryonic stem (ES) and induced pluripotent stem cells [ 8 , 9 ].…”

“…A prominent theme in this issue explores the role of citrullination in the regulation of the extracellular matrix (ECM). The two studies mentioned above, which suggest a role for PADI4 in heart development and demonstrate a non-cell-autonomous role in heart fibrosis, both report that loss of PADI4 leads to the reduction of genes associated with the ECM (Singh et al [ 7 ] ; Van Bruggen et al [ 11 ]). Additionally, an excellent review by Saifi & Ho, details the current knowledge on the mechanisms and functions associated with citrullination of matrisomal proteins [ 12 ].…”

“…While the presence of PADI4 on the cell surface of monocytes was demonstrated previously [ 14 ], and indeed suggested by the fact that extracellular proteins are citrullinated as described above, this systematic study suggests that PADI4 associates with the vesicular organelle fraction of monocytes, presents in vitro evidence that it can bind organelle proteins and identifies organelle-specific PADI4 substrates after monocyte stimulation and PADI4 activation. In light of these findings, it is interesting to consider data presented by Singh et al [ 7 ] , which show that PADI4 loss in ES cells leads to perturbation in the expression of genes associated with vesicular organelles. Collectively, these findings may have important implications regarding PADI4-regulated cellular functions under normal and pathological conditions.…”