Translational control in human keratinocytes following UVB irradiation

UVB 照射后人类角质形成细胞的翻译控制

• 批准号: 9199053
• 负责人: Annie Collier
• 金额: $ 1.63万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-01-01 至 2017-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9199053
• 关键词: Address; Affect; Alpha Cell; Apoptosis; Cell Cycle; Cell Cycle Arrest; Cell Cycle Regulation; Cell Death; Cell Line; Cell Survival; Cell physiology; Cells; Cytoprotection; DNA Damage; DNA Repair; Data; Development; Disease; Eczema; Environmental Carcinogens; Epidermis; Eukaryotic Cell; Eukaryotic Initiation Factor-2; Exhibits; Feedback; Flow Cytometry; G1 Phase; Gene Expression; Gene Targeting; Genes; Genetic Translation; Goals; Healthcare Systems; Human; Immunoblot Analysis; Incidence; Individual; Label; Messenger RNA; Metabolism; Molecular; Pathway interactions; Phosphorylation; Phosphotransferases; Photosensitization; Phototherapy; Play; Process; Protein Biosynthesis; Protein Dephosphorylation; Proteins; Psoriasis; Regulation; Repression; Resistance; Ribosomes; Risk Factors; Role; Skin; Skin Cancer; Skin Carcinoma; Stress; Techniques; The Sun; Thymine Dimers; Transcript; Transcription Coactivator; Translation Initiation; Translational Regulation; Translations; UV protection; UVB induced; United States; Vitiligo; Work; biological adaptation to stress; cancer diagnosis; cell type; experience; genome-wide; genome-wide analysis; improved; innovation; insight; irradiation; keratinocyte; novel; nutrient deprivation; overexpression; protein folding; public health relevance; remediation; repaired; response; ribosome profiling; skin disorder; skin organogenesis; small molecule; small molecule inhibitor; synthetic nucleotide; targeted treatment; tool; transcription factor; transcriptome sequencing; ultraviolet

 DESCRIPTION (provided by applicant): Ultraviolet B (UVB) irradiation is an environmental carcinogen and the most common risk factor for the development of skin cancers, which account for the highest percentage of all cancers diagnosed in the United States each year. Narrow band UVB has also been approved for the treatment of several skin diseases such as psoriasis, vitiligo, and eczema. Therefore, a better understanding of how human skin cells respond to UVB at the molecular level is required for improvement of skin disease therapies. One way that cells protect themselves from environmental stress is by rapid modulation of protein synthesis through phosphorylation of eukaryotic initiation factor 2 (eIF2~P). There are four eIF2 kinases that are each activated by different environmental stresses. Because multiple stresses elicit translational control this pathway is referred to as the Integrated Stress Response (ISR). eIF2~P causes a global repression of translation initiation, coincident with preferential translation of select gene transcripts, such as the transcription factors ATF4 and downstream CHOP. We previously showed that repression of protein synthesis through eIF2~P provides keratinocytes resistance to UVB-induced cell death. However ATF4 is repressed following UVB, and forced ATF4 expression with the small molecule salubrinal results in increased UVB-induced apoptosis. Therefore, the protective aspects of eIF2~P and translational control are independent of ATF4. We hypothesize that eIF2~P elicits positive effects on cell viability following UVB through translational control of mRNAs involved in key protective pathways, such as DNA repair and cell cycle control. In order to investigate this idea, we will subject ISR-deficient keratinocytes to UVB irradiation, followed by immunoblot analysis of thymine dimers as well as cell cycle analysis via synthetic nucleotide labeling followed by flow cytometry. We predict that ISR-deficient keratinocytes will exhibit a reduced ability to repair DNA damage and arrest in the G1 phase of the cell cycle. Secondly, we also propose a genome-wide study of UVB irradiated keratinocytes with ribosome profiling, which will determine changes in ribosome occupancies of individual mRNAs and provide a snapshot of translational changes in the cell following UVB. We predict that ribosome profiling will identify translationally regulated mRNAs that have not previously been investigated in the context of UVB irradiation and skin disease. Together the proposed aims will determine mechanisms by which eIF2~P and translational provides for cytoprotection to UVB irradiation, with the goal of identifying targets for improved skin disease therapies.

 描述（应用程序提供）：紫外线B（UVB）辐射是一种环境致癌物，也是皮肤癌发展的最常见危险因素，这是每年在美国诊断出的所有癌症中最高百分比。狭窄的UVB狭窄，也已被批准用于治疗几种皮肤病，例如牛皮癣，白癜风和湿疹。因此，需要更好地了解人类皮肤细胞在分子水平上如何反应UVB以改善皮肤病疗法。细胞保护自己免受环境压力的一种方法是通过真核启动因子2（EIF2〜p）的磷酸化快速调节蛋白质合成。有四种EIF2激酶都被不同的环境应力激活。由于多种压力会引起翻译控制，因此该途径称为综合应力响应 （ISR）。 EIF2〜p引起翻译启动的全局表达，与选定基因转录物的首选翻译相吻合，例如转录因子ATF4和下游CHOP。我们先前表明，通过EIF2〜P的蛋白质合成的表达提供了角质形成细胞对UVB诱导的细胞死亡的抗性。但是，在UVB之后反映了ATF4，并强迫ATF4表达较小的分子salubrinal会导致增加。 UVB诱导的凋亡。因此，EIF2〜P和翻译控制的受保护方面与ATF4无关。我们假设EIF2〜P通过翻译参与关键保护途径的mRNA（例如DNA修复和细胞周期控制）的mRNA的转换对UVB后的细胞活力产生积极影响。为了研究这一想法，我们将对缺乏ISR缺乏的角质形成细胞进行UVB辐射，然后通过合成的核肽标记进行免疫印迹分析以及细胞周期分析，然后进行流式细胞仪。我们预测，ISR缺陷型角质形成细胞将在细胞周期的G1阶段中降低修复DNA损伤和停滞的能力。其次，我们还提出了一项针对核糖体分析的UVB辐照角质形成细胞的全基因组研究，该研究将确定单个mRNA的核糖体占用率变化，并提供UVB后细胞转换变化的快照。我们预测，核糖体分析将鉴定在UVB辐射和皮肤疾病的背景下以前尚未研究的翻译受调节的mRNA。拟议的目的共同确定了EIF2〜P和翻译为UVB辐射提供细胞保护的机制，其目的是确定改善皮肤病疗法的靶标。