Toll-like receptor 4 deletion promotes bacterial burden and cutaneous tumorigenesis in mice lacking one Ikkalpha allele in keratinocytes

Toll-like receptor 4 (TLR4) is an extracellular pathogen recognition receptor (PRR) that identifies a variety of pathogens and damage-associated molecular patterns. Analysis of human TCGA database reveals many mutations of the Toll-Like Receptor 4 ( TLR4 ) gene in human skin cancers. The exact function of PRRs in response to elevated ambient and commensal bacteria for carcinogenesis is still unknown. Recently, we found a low number of spontaneous skin squamous cell carcinomas (SCCs) in 1-year-old Ikka f/+ K5 Cre mice with specific Ikka reduction in keratinocytes. Interestingly, ablation of Tlr4 enhanced SCC numbers and sizes but reduced the latency of SCC development in Ikkalpha Δ KC /+ ; Tlr4 -/- mice compared to Ikka Δ KC/+ mice. Both spontaneous skin SCCs obtained from Ikka Δ KC/+ and Ikka Δ KC/+ ; Tlr4 -/- mice showed loss of wild-type (WT) Ikka allele. Thus, we hypothesize that Ikka reduction in keratinocytes alters skin microbes and Tlr4 deletion promote this condition, which lead to increased spontaneous skin SCCs. Indeed, the bacterial burden were increased in the skin of Ikka Δ KC/+ and Ikka Δ KC/+ ; Tlr4 -/- mice compared to the skin of WT and Tlr4 -/- mice predominately Gram (+) firmicutes. Further, we explore the interactions between bacteria and their hosts using skin organoids derived from these mice. Organoids are three dimensional structures recapitulating the organ's epithelial layer, linking the gap between in vitro and in vivo models. Skin organoids infected with heat killed gram positive bacteria reveals more DNA damage in the Ikka Δ KC/+ ; Tlr4 -/- along with impaired bacteria clearance in these mice . Together, these findings suggest that increased bacteria cause genomic instability which leads to DNA damage. Additionally, we are investigating which bacterial pathways compromise the integrity of the skin genome through Ikka reduction or Ikka reduction/Tlr4 deletion, as well as whether Tlr4 deletion affects immune cell function in combat against bacterial infections.