Mechanisms Of Lineage-specific Gene Expression

谱系特异性基因表达机制

• 批准号: 7194124
• 负责人: JOHN H KEHRL
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
• 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/7194124
• 关键词: B lymphocyte; apoptosis; cytogenetics; developmental genetics; developmental immunology; developmental neurobiology; gene expression; gene targeting; genetic transcription; genetically modified animals; histogenesis; human tissue; immunocytochemistry; laboratory mouse; microarray technology; motor neurons; northern blottings; nucleic acid probes; pancreas; polymerase chain reaction; site directed mutagenesis; southern blotting; transcription factor

This project focuses on the transcription factors HB9 and BSAP, originally identified in B lymphocytes. The HB9 gene also referred to as HlxB9 encodes a homeodomain coding protein. Gene targeting in mice has revealed a critical role for HB9 in motor neuron and pancreas development. Abnormalities in the sacral region occurs in humans with one abnormal HB9 allele. Mice heterozygotic for an HB9 null mutation are normal, but mice that lack HB9 die at birth of respiratory failure due to a lack of diaphragm innervation. The motor neuron defects may arise from the inappropriate expression of genes normally expressed in other types of neurons. These mice also lack a portion of their pancreas, the dorsal lobe, and have reduced numbers of insulin producing beta cells in their residual ventral lobe. Because the developmental defects and perinatal lethality makes assesmment of the function of HB9 in adult tissues impossible, we have used a conditional gene targeting approach and have introduced loxP sites on either side of exon 3 of the HB9 gene. We have shown that the introduction of the LoxP sites does not interfere with normal HB9 expression. We have obtained MX-Cre transgenic mice, where the Cre recombinase can be induced by interferon treatment. We have introduced the MX-Cre transgene onto the HB9 loxP background. We have shown that the expression of Cre causes the deletion of exon 3 of HB9, but good deletion only occured in spleen, bone marrow, and intestine. In contrast, we found little deletion in the pancreas. We have found little effect on deleting HB9 in those tissues where Cre is expressed. To determine the consequences of deleting HB9 in the developing and adult pancrease we have crossed the HB9 Cre lox mice with mice expressing Cre from a transgene regulated by the insulin promoter. Mice in which HB9 exon 3 is flanked by loxP sites on both alleles and which express Cre from the insulin promoter should be available this fall for analysis. BSAP is a transcription factor critical for B-lymphocyte development and lineage committment. Mice overexpressing BSAP have B cells which are hyperproliferative, have an apoptosis defect, and a propensity for developing lymphomas. We have largely completed the analysis of these mice.

这个项目的重点是转录因子HB 9和BSAP，最初在B淋巴细胞中鉴定。HB 9基因也称为HlxB 9，编码同源结构域编码蛋白。在小鼠中的基因靶向已经揭示了HB 9在运动神经元和胰腺发育中的关键作用。骶骨区的abbreviation发生在具有一个异常HB 9等位基因的人中。HB 9无效突变的杂合子小鼠是正常的，但缺乏HB 9的小鼠在出生时由于缺乏膈肌神经支配而死于呼吸衰竭。运动神经元的缺陷可能是由于在其他类型的神经元中正常表达的基因的不适当表达引起的。这些小鼠还缺少胰腺的一部分，即背叶，并且在其剩余的腹叶中产生胰岛素的β细胞数量减少。由于发育缺陷和围产期致死性使得不可能评估HB 9在成人组织中的功能，我们使用了条件性基因靶向方法，并在HB 9基因外显子3的两侧引入loxP位点。我们已经证明，LoxP位点的引入不干扰正常HB 9表达。我们已经获得了MX-Cre转基因小鼠，其中Cre重组酶可以通过干扰素治疗来诱导。我们已经将MX-Cre转基因引入HB 9 loxP背景中。我们发现Cre的表达导致HB 9外显子3的缺失，但良好的缺失仅发生在脾脏、骨髓和肠中。相比之下，我们在胰腺中发现了很少的缺失。我们发现在表达Cre的组织中删除HB 9几乎没有影响。为了确定在发育和成年胰腺中删除HB 9的后果，我们将HB 9 Cre lox小鼠与表达Cre的小鼠杂交，Cre来自由胰岛素启动子调节的转基因。HB 9外显子3两侧都有loxP位点的小鼠，以及从胰岛素启动子表达Cre的小鼠，应该可以在今年秋天进行分析。BSAP是B淋巴细胞发育和谱系定型的关键转录因子。过表达BSAP的小鼠具有过度增殖的B细胞，其具有凋亡缺陷和发展淋巴瘤的倾向。我们已经基本完成了对这些小鼠的分析。