An exPADItion for citrullination in the developing hair follicle

During epidermal development, to assure proper tissue structure, highly complex transcriptional networks interact within the stem cell compartments of the epidermis and hair follicles (HFs) to balance the choice between self-renewal or differentiation. The full characterisation of the protein profiles resulting from those transcriptional networks, within the compartments of the HF, remains, however, incomplete. Moreover, the proteins themselves can be regulated via posttranslational modification (PTMs). One such PTM is citrullination, carried out by the peptidylarginine deiminase (PADI) family of enzymes. Although, PADIs have been described in other stem and progenitor cells, their role in hair follicle stem cell (HFSC) and progenitor lineages have remained elusive.

The main objectives of this thesis are to address the functional consequences of PADI expression in HFSCs during development.

Paper I identifies Padi4 expression in the developing HF, where it is found to participate in restricting proliferation and lineage commitment of HF progenitors, as well as playing a role in the central mechanism for translational control, and by doing so altering the distinct sequential events that mark HF differentiation progression. As a result, we identify citrullination as a means to assert regulation of protein function in HFSCs and progenitors.

Paper II identifies alternative isoforms of PADI2 and PADI3, in oligodendrocytes and HF differentiated cells, respectively, and show that the alternative isoforms have an incumbering effect on the enzymatic activity and stability of their conventional counterparts.

Paper III is a review paper in which meta-analysis of published human citrullinomes in health and inflammatory disease reveals that citrullination is a commonplace yet highly dynamic molecular regulator of protein function. A strong case is made for the involvement of PADIs and citrullination in hair follicle stem cell biology and inflammatory alopecia.

Paper IV addresses the involvement of transcription factor ID1 in self-renewal and differentiation of epidermal progenitor cells during development. This study describes how ID1 facilitates synchronisation of progenitor proliferation and differentiation via TCF3- binding, and establishes a novel axis of coordination for how BMP-induction of Id1 expression via pSMAD1/5 is supressed by CEBPa.

The combined efforts within this thesis demonstrate the clear and overarching importance of PADIs and citrullination in skin developmental physiology.

List of scientific papers

I. Vikhe Patil, K., Genander, M. PADI4 restricts proliferation of hair follicle progenitor cells by rewiring translation. [Manuscript]

II. Vikhe Patil, K., Meijer, M., Mak, K. H. M., Yang, W., Falcão A. M., Castelo-Branco, G., Genander, M. Co-expression of PADI isoforms during progenitor differentiation enables functional diversity. Philosophical Transactions B. 2023. [Accepted]
https://doi.org/10.1098/rstb.2022.0451

III. Vikhe Patil, K., Mak, K. H. M., Genander, M. A Hairy Cituation – PADIs in Regeneration and Alopecia. Front Cell Dev Biol. (2021) 9, 789676.
https://doi.org/10.3389/fcell.2021.789676

IV. Kantzer, C. G*., Yang, W*., Grommisch, D., Vikhe Patil, K., Mak, K. H. M., Shirokova, V., & Genander, M. ID1 and CEBPA coordinate epidermal progenitor cell differentiation. Development. (2022) 149(22), dev201262. *These authors contributed equally to this work.
https://doi.org/10.1242/dev.201262