RNase L represses hair follicle regeneration through altered innate immune signaling
Charles S. Kirby, Nasif Islam, Eric Wier, Martin P. Alphonse, Evan Sweren, Gaofeng Wang, Haiyun Liu, Dongwon Kim, Ang Li, Sam S. Lee, Andrew M. Overmiller, Yingchao Xue, Sashank Reddy, Nathan K. Archer, Lloyd S. Miller, Jianshi Yu, Weiliang Huang, Jace W. Jones, Sooah Kim, Maureen A. Kane, Robert H. Silverman, Luis A. Garza
Charles S. Kirby, Nasif Islam, Eric Wier, Martin P. Alphonse, Evan Sweren, Gaofeng Wang, Haiyun Liu, Dongwon Kim, Ang Li, Sam S. Lee, Andrew M. Overmiller, Yingchao Xue, Sashank Reddy, Nathan K. Archer, Lloyd S. Miller, Jianshi Yu, Weiliang Huang, Jace W. Jones, Sooah Kim, Maureen A. Kane, Robert H. Silverman, Luis A. Garza
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Research Article Dermatology Inflammation

RNase L represses hair follicle regeneration through altered innate immune signaling

Abstract

Mammalian injury responses are predominantly characterized by fibrosis and scarring rather than functional regeneration. This limited regenerative capacity in mammals could reflect a loss of proregeneration programs or active suppression by genes functioning akin to tumor suppressors. To uncover programs governing regeneration in mammals, we screened transcripts in human participants following laser rejuvenation treatment and compared them with mice with enhanced wound-induced hair neogenesis (WIHN), a rare example of mammalian organogenesis. We found that Rnasel–/– mice exhibit an increased regenerative capacity, with elevated WIHN through enhanced IL-36α. Consistent with RNase L’s known role to stimulate caspase-1, we found that pharmacologic inhibition of caspases promoted regeneration in an IL-36–dependent manner in multiple epithelial tissues. We identified a negative feedback loop, where RNase L–activated caspase-1 restrains the proregenerative dsRNA-TLR3 signaling cascade through the cleavage of toll-like adaptor protein TRIF. Through integrated single-cell RNA-seq and spatial transcriptomic profiling, we confirmed OAS & IL-36 genes to be highly expressed at the site of wounding and elevated in Rnasel–/– mouse wounds. This work suggests that RNase L functions as a regeneration repressor gene, in a functional trade off that tempers immune hyperactivation during viral infection at the cost of inhibiting regeneration.

Authors

Charles S. Kirby, Nasif Islam, Eric Wier, Martin P. Alphonse, Evan Sweren, Gaofeng Wang, Haiyun Liu, Dongwon Kim, Ang Li, Sam S. Lee, Andrew M. Overmiller, Yingchao Xue, Sashank Reddy, Nathan K. Archer, Lloyd S. Miller, Jianshi Yu, Weiliang Huang, Jace W. Jones, Sooah Kim, Maureen A. Kane, Robert H. Silverman, Luis A. Garza

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Figure 4

RNase L suppresses IL-36 expression, which is required for and promotes WIHN.

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RNase L suppresses IL-36 expression, which is required for and promotes ...
(A) Wounded tissue from Rnasel–/– mice reveal elevated ILF16/IL-36α protein, as shown by Western blot. (B) Keratinocytes harvested and cultured from Rnasel–/– mice actively secrete more IL-36α than WT controls, as shown by Western blot. (C) Both unwounded and wounded skin show increased expression of IL-36α (green) in Rnasel–/– mice. IL-36α expression peaks in both WT and Rnasel–/– mice at 3 days after wounding. Scale bar: 200 μm. (D) Injection of 50 ng of recombinant IL-36α protein underneath the scab at WD7 promotes WIHN, as shown by CSLM and AP staining (n = 6, 2-tailed unpaired t test, P = 0.0002). (E). Histology of D comparing vehicle or rmIL-36α–treated mouse skin sections. The neogenic hair follicles (purple) are shown aggregated at the center of the scar. Scale bar: 200 μm. (F) Treatment of human keratinocytes with recombinant IL-36α increases WNT7B mRNA expression (n = 4,1-way ANOVA, P < 0.05), as quantified by qRT-PCR. (G) Il36r–/– mice fail to regenerate hair follicles and are not responsive to poly (I:C), as shown by CSLM and AP staining (n = 6, 2-way ANOVA, P = 0.0147, P < 0.0001). (H) Rnasel–/–/Il36r–/– mice lose the ability to regenerate hair follicles compared with Rnasel–/– mice, as shown by CSLM (n = 3 and 7, respectively, 2-tailed unpaired t test, P < 0.0001). (I) siRNA Knockdown of IL-36α and RNase L in human keratinocytes abrogates the increases of both WNT7B and IL6 morphogenesis markers compared with RNase L siRNA alone (n = 3 each, 1-way ANOVA, P < 0.05).