dsRNA Sensing Induces Loss of Cell Identity

Abstract

How cell and tissue identity persist despite constant cell turnover is an important biologic question with cell therapy implications. Although many mechanisms exist, we investigated the controls for site-specific gene expression in skin, given its diverse structures and functions. For example, the transcriptome of in vivo palmoplantar (i.e., volar) epidermis is globally unique, including Keratin 9 (KRT9). Although volar fibroblasts have the capacity to induce KRT9 in nonvolar keratinocytes, we show here that volar keratinocytes continue to express KRT9 in in vitro solo cultures. Despite this, KRT9 expression is lost with volar keratinocyte passaging, despite stable hypomethylation of its promoter. Coincident with KRT9 loss is a gain of the primitive keratin 7 and a signature of dsRNA sensing, including the double-stranded RNA (dsRNA) receptor DExD/H-Box Helicase 58 (DDX58/RIG-I). Exogenous dsRNA inhibits KRT9 expression in early passage volar keratinocytes or in vivo footpads of wild-type mice. Loss of DDX58 in passaged volar keratinocytes rescues KRT9 and inhibits KRT7 expression. Additionally, DDX58-null mice are resistant to the ability of dsRNA to inhibit KRT9 expression. These results show that the sensing of dsRNA is critical for loss of cell-specific gene expression; our results have important implications for how dsRNA sensing is important outside of immune pathways.

Diverse cellular players orchestrate regeneration after wounding

“…This is reminiscent of immune pathways known to be active in WIHN, such as TLR3 and its substrate, dsRNA. 48,49,52 This pro-healing role of the microbiome is in line with other studies showing probiotics as a reliable and safe alternative to antibiotics in some wounds. 53 Clearly, immune signalling is necessary for WIHN; the presence of bacteria, even at pathological levels, is fundamental for regenerative signalling pathways to operate.…”

“…Parallel studies in human subjects show delayed wound healing when wounds are treated with standard antibiotic cream compared to a simple occlusive medium (manuscript in submission). This is reminiscent of immune pathways known to be active in WIHN, such as TLR3 and its substrate, dsRNA 48,49,52 . This pro‐healing role of the microbiome is in line with other studies showing probiotics as a reliable and safe alternative to antibiotics in some wounds 53 .…”

“…Furthermore, experiments highlighting the importance of dsRNA and TLR3 signalling in ablating keratinocyte positional identity also enforce its connection to stemness and regeneration. 52…”

“…Clearly, immune signalling is necessary for WIHN; the presence of bacteria, even at pathological levels, is fundamental for regenerative signalling pathways to operate. Furthermore, experiments highlighting the importance of dsRNA and TLR3 signalling in ablating keratinocyte positional identity also enforce its connection to stemness and regeneration 52 …”

“…Epidermal keratinocytes and fibroblasts maintain a memory of their positional identity through the expression of various markers, including Krt9 in volar epidermis. dsRNA sensing functions to decrease positional identity, increasing the stem‐like nature of the expressing cells 48,52 . After passage 8‐9 Krt‐9 and other differential marker expression is lost 52 .…”