Carcinogen inactivation of tumor suppressor genes in p53 haploinsufficient mice.

p53 单倍体不足小鼠中肿瘤抑制基因的致癌灭活。

• 批准号: 6432252
• 负责人: JOHN EDGAR FRENCH
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF ENVIRONMENTAL HEALTH SCIENCES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6432252
• 关键词: bladder neoplasm; carbopolycyclic compound; carcinogen testing; carcinogens; chemical carcinogenesis; gene dosage; genetic models; immunocytochemistry; in situ hybridization; laboratory mouse; lymphoma; mutagen testing; mutagens; neoplasm /cancer genetics; sarcoma; single strand conformation polymorphism; southern blotting; tumor suppressor genes; tumor suppressor proteins

Human and rodent transspecies carcinogens (trans-species carcinogens) often demonstrate similar organotropic patterns of neoplasia and loss of heterozygosity (LOH. Recently, we have observed significant chromosome 11 LOH in N5 C57BL/6:129Sv heterozygous p53 mice using simple sequence length polymorphic loci. Primers specific for known SSLP loci revealed amplicons consistent with the two strains, C57BL/6 and 129Sv, in the line. Heterozygosity was unexpected because the mice were reported to be on a C57BL/6-Trp53 (N5) background. We hypothesize that carcinogen induced DNA damage in the p53 haploinsufficient mouse results in illegitimate mitotic recombination during repair and genomic instability leading to neoplasia. By exploiting the observed heterozygosity on chromosome 11 in the 5th backcross generation, we learned that LOH was not restricted to theTrp53 locus. A complete copy of chromosome 11 was lost during exposure to phenolphthalein and lymphomagenesis. The investigation confirmed an aneugenic mechanism of action for phenolphthalein and revealed allelotypes (germline pattern of SSLP loci) that were not consistent with the reported p53 (+/-) C57BL/6 (N5) produced at Taconic by breeding N4 generation p53 nullizygous males to inbred C57BL/6 wildtype females. Chromosome 11 loss also occurred in benzene and p-cresidine induced p53 (+/-) mouse sarcomas (oral, intubation) and thymic lymphomas (inhalation, whole animal) and bladder tumors (dietary). Allelotype data from the benzene and p-cresidine studies are, like those of the phenolphthalein study; inconsistent with the breeding protocol reported by Taconic. The results establish microsatellite (SSLP loci) mapping as a useful tool for determination of LOH in carcinogenesis studies using p53 haploinsufficient mice, e.g. (C57BL/6 x 129Sv) or (C57BL/6 x C3H) F1. In summary, we have shown that in independent studies that there is sufficient heterozygosity on chromosome 11 in the heterozygous p53 deficient (+/-) N5 generation mouse to use microsatellite markers at 5 cM intervals to demonstrate whole or partial chromosome loss through non-disjunction and homologous recombination. Most striking and novel was the observation of an unexpected pattern of germline recombinants (C57BL/6 N4 males crossed to wildtype C57BL/6Tac females). We aim to investigate rates of homologous recombination and determine loci specific positive and negative interference with recombination on chromosome 11 under exposure to environmental carcinogens inducing genomic instability. Specifically, this would enhance our scientific understanding of how this genetically altered mouse model responds when exposed to environmental carcinogens. Using this model, we will determine meiotic (parental and progeny germline) and mitotic recombinant genotype patterns (established in normal somatic tissues of progeny during embryogenesis as well as cancers that arise sporadically with different and unique recombinant genotypes). With microsatellite mapping, we will be able to fine map chromosome 11 sites and rates of homologous recombination and the effect on genomic instability.

人类和啮齿动物跨物种致癌物（跨物种致癌物）通常表现出相似的器官性肿瘤模式和杂合性丧失（LOH）。最近，我们使用简单序列长度多态性基因座在 N5 C57BL/6:129Sv 杂合 p53 小鼠中观察到显着的染色体 11 LOH。针对已知 SSLP 基因座的特异性引物 显示了与该品系中的两个菌株 C57BL/6 和 129Sv 一致的扩增子。杂合性是出乎意料的，因为据报道这些小鼠具有 C57BL/6-Trp53 (N5) 背景。 我们假设致癌物在 p53 单倍体不足的小鼠中诱导 DNA 损伤，导致修复过程中的非法有丝分裂重组和基因组不稳定，从而导致 瘤形成。通过利用在第 5 代回交中观察到的 11 号染色体杂合性，我们了解到 LOH 并不限于 Trp53 基因座。 11 号染色体的完整副本在接触酚酞和淋巴瘤发生过程中丢失。 该研究证实了酚酞的非优生作用机制，并揭示了等位基因型（种系模式） SSLP 位点）与 Taconic 通过将 N4 代 p53 无效雄性与近交 C57BL/6 野生型雌性进行繁殖而产生的 p53 (+/-) C57BL/6 (N5) 不一致。 11 号染色体缺失也发生在苯和对甲酚胺诱导的 p53 (+/-) 小鼠肉瘤（口服、插管）和 胸腺淋巴瘤（吸入，整个动物）和膀胱肿瘤（饮食）。 苯和对甲酚胺研究的等位基因型数据与酚酞研究的等位基因型数据类似；与 Taconic 报告的育种方案不一致。结果表明，微卫星（SSLP 位点）作图是使用 p53 确定致癌作用研究中 LOH 的有用工具 单倍体不足的小鼠，例如(C57BL/6 x 129Sv) 或 (C57BL/6 x C3H) F1。 总之，我们在独立研究中表明，杂合 p53 缺陷 (+/-) N5 代小鼠的 11 号染色体上存在足够的杂合性，可以使用间隔 5 cM 的微卫星标记来证明全部或部分染色体丢失 通过非分离和同源重组。 最引人注目和新颖的是观察到一种意想不到的种系重组体模式（C57BL/6 N4 雄性与野生型 C57BL/6Tac 雌性杂交）。 我们的目的是研究同源重组率，并确定在暴露于诱导基因组的环境致癌物的情况下，对 11 号染色体重组的位点特异性正干扰和负干扰。 不稳定。具体来说，这将增强我们对这种转基因小鼠模型在暴露于环境致癌物时如何反应的科学理解。 使用该模型，我们将确定减数分裂（亲本和子代种系）和有丝分裂重组基因型模式（在胚胎发生过程中在子代的正常体细胞组织中建立，以及具有不同和独特的重组基因型的散发性癌症）。与 通过微卫星作图，我们将能够精细绘制 11 号染色体位点和同源重组率以及对基因组不稳定性的影响。