A Hairy Cituation – PADIs in Regeneration and Alopecia

In this Review article, we focus on delineating the expression and function of Peptidyl Arginine Delminases (PADIs) in the hair follicle stem cell lineage and in inflammatory alopecia. We outline our current understanding of cellular processes influenced by protein citrullination, the PADI mediated posttranslational enzymatic conversion of arginine to citrulline, by exploring citrullinomes from normal and inflamed tissues. Drawing from other stem cell lineages, we detail the potential function of PADIs and specific citrullinated protein residues in hair follicle stem cell activation, lineage specification and differentiation. We highlight PADI3 as a mediator of hair shaft differentiation and display why mutations in PADI3 are linked to human alopecia. Furthermore, we propose mechanisms of PADI4 dependent fine-tuning of the hair follicle lineage progression. Finally, we discuss citrullination in the context of inflammatory alopecia. We present how infiltrating neutrophils establish a citrullination-driven self-perpetuating proinflammatory circuitry resulting in T-cell recruitment and activation contributing to hair follicle degeneration. In summary, we aim to provide a comprehensive perspective on how citrullination modulates hair follicle regeneration and contributes to inflammatory alopecia.

Intracellular pH dynamics regulates intestinal stem cell fate

Emerging evidence is revealing critical roles of intracellular pH (pHi) in development, but it remains unclear whether pHi regulates stem cell fate specification. We find that pHi dynamics is a key regulator of cell fate in the mouse intestinal stem cell lineage. We identify a pHi gradient along the intestinal crypt axis and find that dissipating this gradient inhibits crypt budding due to loss Paneth cell differentiation. Mechanistically, decreasing pHi biases intestinal stem cell fate toward the absorptive and away from the secretory lineage, by regulating the activity of the lineage transcription factor Atoh1. Our findings reveal a previously unrecognized role for pHi dynamics in the specification of cell fate within an adult stem cell lineage.

Engineering hydrogels with homogeneous mechanical properties for controlling stem cell lineage specification

The extracellular matrix (ECM) is mechanically inhomogeneous due to the presence of a wide spectrum of biomacromolecules and hierarchically assembled structures at the nanoscale. Mechanical inhomogeneity can be even more pronounced under pathological conditions due to injury, fibrogenesis, or tumorigenesis. Although considerable progress has been devoted to engineering synthetic hydrogels to mimic the ECM, the effect of the mechanical inhomogeneity of hydrogels has been widely overlooked. Here, we develop a method based on host–guest chemistry to control the homogeneity of maleimide–thiol cross-linked poly(ethylene glycol) hydrogels. We show that mechanical homogeneity plays an important role in controlling the differentiation or stemness maintenance of human embryonic stem cells. Inhomogeneous hydrogels disrupt actin assembly and lead to reduced YAP activation levels, while homogeneous hydrogels promote mechanotransduction. Thus, the method we developed to minimize the mechanical inhomogeneity of hydrogels may have broad applications in cell culture and tissue engineering.

Keratin Profiling by Single-Cell RNA-Sequencing Identifies Human Prostate Stem Cell Lineage Hierarchy and Cancer Stem-Like Cells

Single prostate stem cells can generate stem and progenitor cells to form prostaspheres in 3D culture. Using a prostasphere-based label retention assay, we recently identified keratin 13 (KRT13)-enriched prostate stem cells at single-cell resolution, distinguishing them from daughter progenitors. Herein, we characterized the epithelial cell lineage hierarchy in prostaspheres using single-cell RNA-seq analysis. Keratin profiling revealed three clusters of label-retaining prostate stem cells; cluster I represents quiescent stem cells (PSCA, CD36, SPINK1, and KRT13/23/80/78/4 enriched), while clusters II and III represent active stem and bipotent progenitor cells (KRT16/17/6 enriched). Gene set enrichment analysis revealed enrichment of stem and cancer-related pathways in cluster I. In non-label-retaining daughter progenitor cells, three clusters were identified; cluster IV represents basal progenitors (KRT5/14/6/16 enriched), while clusters V and VI represent early and late-stage luminal progenitors, respectively (KRT8/18/10 enriched). Furthermore, MetaCore analysis showed enrichment of the “cytoskeleton remodeling–keratin filaments” pathway in cancer stem-like cells from human prostate cancer specimens. Along with common keratins (KRT13/23/80/78/4) in normal stem cells, unique keratins (KRT10/19/6C/16) were enriched in cancer stem-like cells. Clarification of these keratin profiles in human prostate stem cell lineage hierarchy and cancer stem-like cells can facilitate the identification and therapeutic targeting of prostate cancer stem-like cells.