C/EBPβ Regulation of the Type 1 IFN Response; Sensitizing Keratinocytes to Direct Activators of Cytosolic PRRs and DNA Damage-Induced Cell Death

细胞应激、DNA损伤、病原体伤害和免疫机制可以激活分子通路， 导致正常的细胞死亡。增加我们对调节细胞死亡决定的途径的理解 对细胞应激的反应至关重要，因为调控不当的细胞死亡与癌症、神经退行性变和 自身免疫性疾病。表皮是抵御皮肤微生物、病毒和细菌的第一道防线。 环境的侮辱，角质形成细胞在宿主的先天免疫反应中起着关键作用。 I型干扰素(干扰素-I)。除了抗微生物和抗病毒反应外，这种相同的干扰素-I反应还 调节多种细胞和生物反应，如增殖、凋亡、衰老和DNA 损坏反应。来自外部实验室的最新数据表明，CCAAT/增强子结合蛋白-β (C/eBPβ)转录因子是一种新的角质形成细胞型干扰素-I反应调节因子。我们观察到 条件性缺失小鼠表皮C/EBPβ(CKOβ)导致IFNb和IFNb表达增加 大量的ISG，包括胞质模式识别受体(CPRRs)。CPRRs感知病毒/病原体 RNA和DNA以及受损的宿主胞浆DNA和RNA，以触发干扰素-I反应。CKOβ 经胞浆PRRs直接激活剂处理的角质形成细胞显示出显著增强的干扰素-I反应 令人惊讶的是，通过激活caspase-8和caspase-3，导致细胞凋亡增加。简明扼要地 提到，DNA损伤可以激活先天免疫反应。DNA损伤剂可能导致 核糖核酸和脱氧核糖核酸的修饰和结构变化，这些修饰和结构变化可能被胞质PRRs感知。我们 观察到ckoβ角质形成细胞受到dna损伤后，caspase8介导的表达增加。 依赖干扰素a/b受体(INFAR)的细胞凋亡。I型干扰素反应的激活可以 导致具有凋亡功能的ISGs表达增加，从而激活caspase-8介导的细胞凋亡 通过死亡受体信号。这项建议的目的是了解C/EBPβ的分子基础 负向调节干扰素-I和半胱氨酸天冬氨酸蛋白酶-8介导的细胞凋亡 体内小鼠表皮和原代角质形成细胞胞浆PRRs和DNA损伤。我们假设 C/eBPβ负性调节角质形成细胞的干扰素-I反应及C/eBPβ的缺失增敏 角质形成细胞通过病原体RNA/DNA或受损的宿主RNA/DNA直接激活胞浆PRRs 诱导促凋亡的ISGs和caspase 8介导的细胞凋亡。这些实验将增加我们的 C/EBPβ调节cPRRs激活和干扰素-I应答的新途径的理解 控制细胞死亡决定，以应对细胞压力，包括DNA损伤。这增加了 了解这一点可以确定新的治疗靶点，以恢复细胞死亡的适当调节来治疗皮肤 癌症、皮肤传染病和皮肤自身免疫性疾病。

Cellular stress, DNA damage, pathogen insult, and immune mechanisms can activate molecular pathways that result in regulated cell death. Increasing our understanding of the pathways that regulate cell death decisions in response to cellular stress is critical as misregulated cell death is linked to cancer, neurodegeneration and autoimmune diseases. The epidermis is the first line of defense to cutaneous microbes, viruses and environmental insults, and keratinocytes play a critical role in the host innate immune response mediated by type I interferons (IFN-I). In addition to anti-microbial and anti-viral responses this same IFN-I response also mediates diverse cellular and biological responses such as proliferation, apoptosis, senescence and the DNA damage response. Recent data from out laboratory suggest that the CCAAT/enhancer-binding protein-β (C/EBPβ) transcription factor is a novel regulator the keratinocyte type IFN-I response. We observed the conditional deletion of C/EBPβ in mouse epidermis (CKOβ) resulted in increased expression of IFNb and numerous ISGs which included cytosolic pattern recognition receptors (cPRRs). cPRRs sense viral/pathogen RNA and DNA as well as damaged host cytosolic DNA and RNA to trigger a IFN-I response. CKOβ keratinocytes treated with direct activators of cytosolic PRRs displayed a greatly increased IFN-I response that surprisingly resulted in increased apoptosis via the activation of caspase-8 and caspase-3. As breifly mentioned, DNA damage can activate the innate immune response. DNA damaging agents can result in modifications and structural changes in RNA and DNA that could potentially be sensed by cytosolic PRRs. We observed that CKOβ keratinocytes challenged with DNA damage displayed increase caspase 8-mediated apoptosis that was dependent on the interferon a/b receptor (INFAR). Activation of a type IFN-I response can result in the increased expression of ISGs with apoptotic functions that activate caspase-8 mediated apoptosis via death receptor signaling. The objective of this proposal is to understand molecular basis for how C/EBPβ negatively regulates the IFN-I response and caspase-8 mediated apoptosis in response to direct activators of cytosolic PRRs and DNA damage in vivo in mouse epidermis and in primary keratinocytes. We hypothesize that C/EBPβ negatively regulates the IFN-I response in keratinocytes and the loss of C/EBPβ sensitizes keratinocytes to the direct activation of cytosolic PRRs by pathogen RNA/DNA or damaged host RNA/DNA to induce pro-apoptotic ISGs and caspase 8-mediated apoptosis. These experiments will increase our understanding of a novel pathway by which C/EBPβ regulates activation of cPRRs and the IFN-I response to control cell death decisions in response to cellular stress including DNA damage. This increased understanding could identify new therapeutic targets to restore proper regulation of cell death to treat skin cancer, skin infectious diseases, and skin autoimmune diseases.