How do the cells of an organism, all of which have exactly the same genetic code, adopt such different fates, morphologies and functions? And how do they then respond to the signals and stresses around them in order to make up a living, growing, healthy organism that can adapt to its environment?
Seminal work in the field of Epigenetics has taught us that the answer to the first question lies in the fact that our genome is subject to epigenetic regulation, which ensures its stability and determines when and where genes produce their transcript and protein products. And the answer to the second question lies largely within the fact that these proteins, which then go on to execute most of the cell’s functions, are themselves subject to regulatory mechanisms which determine when, where and how a protein will function. Our lab combines these two fascinating biological questions to understand how genome-regulating proteins are themselves regulated during development.
We employ biochemistry, cell and molecular biology, genomic and epigenetic approaches and mouse model systems to understand the mechanisms that modulate the function of epigenetic regulators, how these mechanisms are perturbed in disease and how they may be targeted for therapeutic effect. We have a particular interest in protein post-translational modifications (PTMs). These are small chemical changes that happen on proteins as a result of cell signalling changes and can quickly alter the activity, stability and sub-cellular localisation of these proteins, as well as their affinity for other molecules. As a result, PTMs add an enormous degree of sophistication to biological systems, beyond what can be achieved by gene regulation.
Our favourite PTM is citrullination, the conversion of an arginine residue to the non-coded amino acid citrulline. Exciting developments in this classically under-explored field have shown that citrullination and the enzymes that catalyse it, the peptidylarginine deiminases (PADIs or PADs), regulate many aspects of cell physiology, while their deregulation contributes to the development of pathologies such as autoimmunity, neurodegeneration and cancer. Understanding the mechanisms that control PADIs and other epigenetic regulators in response to developmental cues and cellular stresses can offer valuable insights into human health, which can be exploited towards therapeutic benefit in a variety of disease conditions.
The post-translational modification of proteins expands the regulatory scope of the proteome far beyond what is achievable through genome regulation. The field of protein citrullination has seen significant progress in the last two decades. The small family of peptidylarginine deiminase (PADI or PAD) enzymes, which catalyse citrullination, have been implicated in virtually all facets of molecular and cell biology, from gene transcription and epigenetics to cell signalling and metabolism. We have learned about their association with a remarkable array of disease states and we are beginning to understand how they mediate normal physiological functions. However, while the biochemistry of PADI activation has been worked out in exquisite detail , we still lack a clear mechanistic understanding of the processes that regulate PADIs within cells, under physiological and pathophysiological conditions. This review summarizes and discusses the current knowledge, highlights some of the unanswered questions of immediate importance and gives a perspective on the outlook of the citrullination field.
Peptidylarginine deiminases (PADIs) are strongly associated with the development of autoimmunity, neurodegeneration and cancer but their physiological roles are ill-defined. The nuclear deiminase PADI4 regulates pluripotency in the mammalian pre-implantation embryo but its function in tissue development is unknown. PADI4 is primarily expressed in the bone marrow, as part of a self-renewal-associated gene signature. It has been shown to regulate the proliferation of multipotent haematopoietic progenitors and proposed to impact on the differentiation of haematopoietic stem cells (HSCs), suggesting that it controls haematopoietic development or regeneration. Using conditional in vivo models of steady state and acute Padi4 ablation, we examined the role of PADI4 in the development and function of the haematopoietic system. We found that PADI4 loss does not significantly affect HSC self-renewal or differentiation potential upon injury or serial transplantation, nor does it lead to HSC exhaustion or premature ageing. Thus PADI4 is dispensable for cell-autonomous HSC maintenance, differentiation and haematopoietic regeneration. This work represents the first study of PADI4 in tissue development and indicates that pharmacological PADI4 inhibition may be tolerated without adverse effects.
Protein post-translational modifications (PTMs) add great sophistication to biological systems. Citrullination, a key regulatory mechanism in human physiology and pathophysiology, is enigmatic from an evolutionary perspective. Although the citrullinating enzymes peptidylarginine deiminases (PADIs) are ubiquitous across vertebrates, they are absent from yeast, worms and flies. Based on this distribution PADIs were proposed to have been horizontally transferred, but this has been contested. Here, we map the evolutionary trajectory of PADIs into the animal lineage. We present strong phylogenetic support for a clade encompassing animal and cyanobacterial PADIs that excludes fungal and other bacterial homologues. The animal and cyanobacterial PADI proteins share functionally relevant primary and tertiary synapomorphic sequences that are distinct from a second PADI type present in fungi and actinobacteria. Molecular clock calculations and sequence divergence analyses using the fossil record estimate the last common ancestor of the cyanobacterial and animal PADIs to be less than one billion years old. Additionally, under an assumption of vertical descent, PADI sequence change during this evolutionary time frame is anachronistically low, even when compared to products of likely endosymbiont gene transfer, mitochondrial proteins and some of the most highly conserved sequences in life. The consilience of evidence indicates that PADIs were introduced from cyanobacteria into animals by horizontal gene transfer (HGT). The ancestral cyanobacterial PADI is enzymatically active and can citrullinate eukaryotic proteins, suggesting that the PADI HGT event introduced a new catalytic capability into the regulatory repertoire of animals. This study reveals the unusual evolution of a pleiotropic protein modification.