Persistent IKKα phosphorylation induced apoptosis in UVB and Poly I:C co-treated HaCaT cells plausibly through pro-apoptotic p73 and abrogation of IκBα

Toll-like Receptor 3 (TLR3), a member of Pattern Recognition Receptors, is reported to initiate skin inflammation by recognizing double-strand RNA (dsRNA) released from UVB-irradiated cells. Recently, we have discovered the NF-κB pathway activated by TLR3 is involved in Apoptosis of UVB-Poly I:C-treated HaCaT cells. The real culprit for Apoptosis has not been precisely identified since the system of NF-κB pathway is complex. In this study, we silenced main transcriptional factors in NF-κB family, RelA, RelB and c-Rel, but to our surprise the results show that none of them participate in Apoptosis induction in UVB-Poly I:C-treated HaCaT cells. Therefore, we moved to investigate the apoptosis-associated molecules in the upstream of NF-κB pathway. We firstly checked the expression of IκBα, an NF-κB Inhibitor. UVB (4.8 mJ/cm²) and Poly I:C (0.3 μg/mL) co-treatment decreased IκBα expression level in a time-dependent manner. Silencing IκBα with siRNA further enhanced UVB-Poly I:C-induced cell death. We then investigated IκB kinase (IKK) complex that contributes to the degradation of IκBα. IKK is composed of IKKα, IKKβ and NEMO. Treatment with IKK-16, an IKKα/β inhibitor, significantly diminished UVB-Poly I:C-induced IκBα degradation and thus Apoptosis. Silencing either IKKα or NEMO but not IKKβ with corresponding siRNA inhibited Apoptosis. Tumor repressor p73, a homologue of p53, is reported to mediate IKKα-induced Apoptosis in DNA damage response. Silencing p73 reduced cell Apoptosis in UVB-Poly I:C-treated HaCaT cells. In summary, UVB and Poly I:C co-treatment activates IKKα and NEMO, which diminishes anti-apoptotic IκBα, resulting in enhancement of Apoptosis through p73. The findings partially clarify the possible molecular mechanism of pro-apoptotic NF-κB pathway activated by TLR3 in the fate of UVB-irradiated epidermis.