Generation of a plasma cell-specific inducible Cre transgenic mouse

项目概要/摘要 浆细胞在感染或接种疫苗时产生抗体，但也发挥致病作用 自身免疫和器官移植尽管浆细胞在体液免疫中起关键作用， 目前还没有允许特异性地在浆细胞中条件性缺失基因的转基因小鼠模型。 该提议旨在产生并验证小鼠模型，该小鼠模型特异性地表达诱导型Cre， 浆细胞这一目标将通过使用piggyBac转座酶插入含有转座酶的BAC盒来实现。 Tnfrsf 17基因座上游的他莫昔芬诱导型CreERT 2基因。Tnfrsf 17编码蛋白质B细胞 成熟抗原（BCMA），并且仅在浆细胞中表达。由于BCMA对浆细胞至关重要， 存活和敲入或CRISPR/Cas9技术可以导致无效等位基因，BAC技术提供了 这是有利的替代方案，因为它保持内源基因表达完整。目标1将着手产生 小鼠模型和Aim 2将致力于验证模型的特异性和效率。测试 由于对浆细胞谱系的特异性，BCMA-CreERT 2小鼠将与谱系追踪小鼠杂交 R26R-EYFP。当这些小鼠用他莫昔芬治疗时，浆细胞将激活Cre重组酶， 缺失Rosa 26驱动的EYFP上游的loxP侧翼终止密码子，不可逆地标记浆细胞。 由于Rosa 26基因座处的loxP位点对Cre是众所周知的敏感，我们还将测试BCMA-100的能力。 CreERT 2小鼠中有效地从Mcl-1fl/fl小鼠中删除Mcl-1基因。长寿的浆细胞需要Mcl-1 因此有效的删除将导致长寿命浆细胞的固定库的损失。 在完成这些目标后，BCMA-CreERT 2小鼠将成为分析长寿命肿瘤的有价值的工具。 浆细胞的研究将促进各种疾病的治疗。

Project Summary/Abstract Plasma cells produce antibodies in response to infection or vaccination, but also play pathogenic roles in autoimmunity and organ transplant. Despite the critical role of plasma cells in humoral immunity, there is currently no transgenic mouse model that allows for conditional deletion of genes specifically in plasma cells. This proposal aims to generate and validate a mouse model that expresses an inducible Cre specifically in plasma cells. This goal will be achieved by using piggyBac transposase to insert a BAC cassette containing the tamoxifen inducible CreERT2 gene upstream of the Tnfrsf17 locus. Tnfrsf17 encodes the protein B cell maturation antigen (BCMA) and is expressed exclusively in plasma cells. Since BCMA is critical for plasma cell survival and knock-in or CRISPR/Cas9 techniques can result in a null allele, BAC technology provides advantageous alternative because it leaves endogenous gene expression intact. Aim 1 will set out to generate the mouse model and Aim 2 will be dedicated to validating the specificity and efficiency of the model. To test specificity to the plasma cell lineage, the BCMA-CreERT2 mouse will be crossed to the lineage-tracing mouse R26R-EYFP. When these mice are treated with tamoxifen the plasma cells will activate Cre recombinase and delete a loxP flanked stop codon upstream of Rosa26 driven EYFP, irreversibly labeling the plasma cells. Since loxP sites at the Rosa26 locus are notoriously sensitive to Cre, we will also test the ability of the BCMA- CreERT2 mouse to efficiently delete the Mcl-1 gene from Mcl-1fl/fl mice. Long-lived plasma cells require Mcl-1 for survival and therefore efficient deletion will result in the loss of the standing pool of long-lived plasma cells. Upon completion of these aims, the BCMA-CreERT2 mouse will be a valuable tool for the analysis of long-lived plasma cells leading to research that will advance the treatment of a variety of diseases.