The role of AID/APOBEC3 proteins in genome instability in multiple myeloma

总结 多发性骨髓瘤（MM）是一种骨髓浆细胞恶性肿瘤， 和过渡阶段。所有阶段的细胞都表现出显著的基因组畸变，其来源是 没有很好地理解。这在我们对MM的驱动机制的理解上留下了很大的空白 启动和进展，这是本提案旨在填补的空白。最近的证据表明 胞苷脱氨酶--在DNA中将胞嘧啶转化为尿嘧啶的酶--是基因组中的重要罪魁祸首。 一个重要的例子是激活诱导的脱氨酶，AID，它启动体细胞的 超突变和类别转换重组，但也使B细胞基因组的许多区域突变 并导致类似于MM中发现的易位。 脱氨酶也可以使DNA突变，许多发现将AID和APOBEC 3酶与基因组联系起来， 我们的初步数据表明MM细胞表达APOBEC 3B，C， D、F和G，其中APOBEC 3B表达特别强。有趣的是，APOBEC 3B最近 与乳腺癌的基因组不稳定性有关。我们进一步发现，AID的表达和某些 APOBEC 3酶增加MM细胞中DNA链断裂的水平。基于此，我们假设 AID和APOBEC 3家族酶是基因组畸变和疾病进展的主要原因 最近的研究结果提供了脂质失调，免疫激活和MM之间的密切联系， 多达三分之一的克隆性丙种球蛋白病在MM患者中发现对脂质有反应。因此，重要的 我们工作的总体指导假设是，慢性B细胞活化是由高水平的 炎性脂质导致AID/APOBEC 3活性增加和MM疾病进展。我们 系统地测试这些假设在三个目标。在目标1中，我们确定 AID/APOBEC 3酶，单独和协同，有助于生物化学和分子测量， 核脱氨酶活性和基因组损伤。在目标2中，我们确定这些脱氨酶是如何 调节，揭示了MM基因组中易受突变、DNA断裂和 因其行为而引起的转移。最后，在目标3中，我们使用已建立的和新的小鼠模型系统 和MM及其在人源化小鼠中增殖的癌前阶段，以评估 炎症脂质对AID/APOBEC 3因子的表达/活性、DNA损伤和 MM基因组和MM的克隆进化。总之，拟议的实验将提供一个 胞苷脱氨酶在MM中作用的活性、靶向和结局的综合情况， 对疾病发病机制的广泛影响。

SUMMARY Multiple Myeloma (MM) is a malignancy of bone marrow plasma cells preceded by a series of premalignant and transitional stages. Cells at all stages exhibit significant genomic aberrations, the sources of which are not well understood. This has left a major gap in our understanding of the mechanisms that drive MM initiation and progression, a gap that this proposal is designed to fill. Recent evidence suggests that cytidine deaminases—enzymes that convert cytosine to uracil in DNA—are important culprits in genomic instability in MM. One important example is the activation induced deaminase, AID, which initiates somatic hypermutation and class switch recombination but which also mutates many regions of the B cell genome and causes translocations similar to those found in MM. The related APOBEC3 family of cytidine deaminases can also mutate DNA, and numerous findings link AID and APOBEC3 enzymes to genome instability and mutations in MM. Our preliminary data demonstrate that MM cells express APOBEC3B, C, D, F and G, with APOBEC3B being expressed particularly strongly. Interestingly, APOBEC3B has recently been implicated in genome instability in breast cancer. We further find that expression of AID and certain APOBEC3 enzymes increases levels of DNA strand breaks in MM cells. Based on this, we hypothesize that AID and APOBEC3 family enzymes are a major cause of genomic aberrations and disease progression in MM. Recent findings provide a strong link between lipid disregulation, immune activation, and MM, with as much as a third of the clonal gammopathies found in MM patients reacting to lipids. Hence, an important overall guiding hypothesis of our work is that chronic B cell activation arising from elevated levels of inflammatory lipids contributes to increased activity of AID/APOBEC3 and MM disease progression. We systematically test these hypotheses in three Aims. In Aim 1, we determine the extent to which AID/APOBEC3 enzymes, alone and in concert, contribute to biochemical and molecular measures of nuclear deaminase activity and genomic damage. In Aim 2, we determine how these deaminases are regulated, revealing those regions of the MM genome that are susceptible to mutation, DNA breaks, and translocations due to their action. Finally, in Aim 3 we use established and novel mouse model systems and MM and its premalignant stage propagated in humanized mice to assess the in vivo effects of inflammatory lipids on the expression/activity of AID/APOBEC3 factors, DNA damage and mutation of the MM genome, and clonal evolution of MM. Together, the proposed experiments will provide a comprehensive picture of the activity, targeting, and outcome of cytidine deaminase action in MM, with broad implications for disease pathogenesis.