PIWI - Transposon regulation by epithelial adherens junctions

PIWI - 上皮粘附连接的转座子调节

• 批准号: 9979311
• 负责人: Antonis Kourtidis
• 金额: $ 20.97万
• 依托单位: MEDICAL UNIVERSITY OF SOUTH CAROLINA
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-07-01 至 2022-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9979311
• 关键词: Adherens Junction; Apical; Architecture; Area; Biological; Biological Assay; Biology; Breast; Breast Cancer Cell; Breast Epithelial Cells; CRISPR/Cas technology; Cadherins; Carcinoma; Cell Differentiation process; Cell Line; Cell-Cell Adhesion; Cells; Chemicals; Colon; Complex; DNA Damage; DNA Double Strand Break; DNA Transposable Elements; DNA Transposons; Data; Development; Disease; E-Cadherin; Elements; Epithelial; Epithelial Cells; Epithelium; Family; Foundations; Future; Gene Mutation; Genomic Instability; Genomics; Goals; Human Genome; Inherited; Kidney; Knowledge; Lead; Ligation; Link; Malignant Neoplasms; MicroRNAs; Microscopy; Modeling; Mutagenesis; Mutation; Normal tissue morphology; Oncogenes; Organ; Pathway interactions; Phenotype; RNA; RNA Interference; Radiation; Regulation; Research; Resolution; Retrotransposition; Role; Small RNA; Structure; Technology; Testing; Tissue Differentiation; Tissue Sample; Tissues; Tumor Promoters; Tumor Suppressor Proteins; Up-Regulation; Work; base; cancer cell; cell transformation; de novo mutation; genome editing; genome integrity; human tissue; innovation; insight; kidney epithelial cell; malignant breast neoplasm; mortality; novel; novel therapeutics; piRNA; prevent; recruit; targeted treatment; tool; transcriptome sequencing; transposon sequencing; tumor; tumorigenesis; tumorigenic

PROJECT SUMMARY Transposon activity accounts for genomic instability in more than 50% of epithelial cancers. However, the reasons of this activity are still unclear. The E-cadherin-based adherens junctions are essential structural components of the epithelial cells and frequently compromised in tumors. We have found association of cadherin junctions with PIWIL2, a key component of the piRNA-processing pathway that is responsible for silencing of transposable elements. piRNAs comprise the largest class of small RNAs and have been extensively studied in the germline; however, their roles in somatic tissues are unclear. Our preliminary data reveal localization of PIWIL2 at the mature apical adherens junctions of well-differentiated breast, kidney and colon epithelial cells, whereas this localization is lost in cancer cells. Interestingly, E-cadherin depletion results in loss of junctional localization of PIWIL2, in upregulation of a transposable element, and increased levels of γ-H2AX, which is an indicator of DNA damage. A hallmark of increased transposon activity is DNA double-stranded breaks. We hypothesize that the adherens junctions recruit PIWIL2 to suppress transposon activity in differentiated epithelial cells to maintain genomic integrity and the normal epithelial phenotype. We will test this hypothesis under the following Aims: 1) examine whether E-cadherin suppress transposon levels and activity by enabling formation of a PIWI-piRNA complex in well-differentiated epithelial cells; 2) investigate whether the junction-associated PIWIL2 suppresses pro-tumorigenic transformation. This work is significant, since it will fill a gap in the knowledge of the role of the PIWI-transposon regulation in differentiated epithelial tissues and in cancer. The proposal is innovative, since it provides an unexpected mechanistic link between cell-cell adhesion, PIWI- transposon biology and genomic integrity. In addition, it employs cutting-edge technologies, such as piRNA- transposon sequencing, CRISPR/Cas9 genome editing, and super-resolution microscopy. The long-term goal of this study is to identify a new mode of regulation of transposon silencing, coordinated by the adherens junctions, which could be critical for suppression of transposon-driven mutagenesis and tumorigenesis. Successful completion of the above Aims will help us gain insights into a new mechanism that tethers cell architecture to genomic integrity and generate significant data for subsequent R01-level proposals.

项目摘要 转座子活性解释了超过50％的上​​皮癌的基因组不稳定性。但是， 此活动的原因尚不清楚。基于电子 - 钙粘蛋白的粘附连接是必不可少的结构 上皮细胞的成分，并经常在肿瘤中损害。我们发现了钙粘蛋白的关联 与Piwil2的交界处，Piwil2是PIRNA加工途径的关键组成部分，该途径负责沉默 转座元素。 PIRNA是最大的小型RNA类，并且已广泛研究 种系；但是，它们在体细胞中的作用尚不清楚。我们的初步数据揭示了 piwil2处于成熟的顶端粘附剂连接良好的乳房，肾脏和结肠上皮细胞的连接 而这种定位在癌细胞中丢失。有趣的是，电子 - 钙黏着蛋白的部署会导致连接损失 piwil2的定位，在可转座元件的上调和γ-H2AX的水平增加时，这是一种 DNA损伤的指标。转座子活性增加的标志是DNA双链断裂。我们 假设粘附器连接募集了piwil2以抑制分化上皮的转座子活性 维持基因组完整性和正常上皮表型的细胞。我们将在 以下目的：1）检查E-钙粘蛋白是否通过形成抑制转座子的水平和活动 在分化良好的上皮细胞中的piwi-piRNA复合物； 2）调查连接相关是否相关 PIWIL2抑制促肿瘤的转化。这项工作很重要，因为它将填补 了解Piwi-Transposon调节在分化上皮组织和癌症中的作用。这 提案具有创新性，因为它在细胞 - 细胞粘合剂，piwi-之间提供了意外的机械联系。 转座子生物学和基因组完整性。此外，它采用了尖端技术，例如pirna- 转座子测序，CRISPR/CAS9基因组编辑和超分辨率显微镜。长期目标 这项研究是为了确定一种新的转座子沉默调节模式，由粘附连接协调， 这对于抑制转座子驱动的诱变和肿瘤发生至关重要。 以上目的的完成将有助于我们深入了解一种新机制，将细胞体系结构推向 基因组完整性并为随后的R01级建议生成重要数据。