REGULATION OF MORPHOGENESIS IN EARLY MOUSE DEVELOPMENT

小鼠早期发育中形态发生的调节

• 批准号: 08044283
• 负责人: SHIMADA Kazunori
• 金额: $ 5.63万
• 依托单位: Osaka University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for international Scientific Research
• 财政年份: 1996
• 资助国家: 日本
• 起止时间: 1996 至 1997
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08044283/
• 关键词: bmil; polyhomeotic; protein interactions; rae28; Polycomb group; Hox; segment specification; neural crest; bmil; Pc-G遺伝子群; polyhomeotic遺伝子; Polycomb遺伝子; rae28遺伝子; M33遺伝子; bmi-1遺伝子; CATCH22症候群

l) One of our major areas of interest has been to identify which Polycomb group (PcG) proteins interact, and to determine the interaction domains, both in flies and mammals. We have shown that three fly PcG proteins interact. Psc (Posterior sex combs) interacts with ph (polyhomeotic) and Pc (Polycomb), but ph and Pc do not interact. The same is true for their mouse homologues. RAE28 (ph), BMI1 (Psc), and M33 (Pc) interact homotypically. Both RAE28 and M33 interact with BMI1 , but not with each other. In addition, ph interacts with Scm (Sex comb on midleg) via a conserved domain found in many proteins. The mouse homologues of ph and Scm (RAE28 and mScm) also interact directly. Dr.Brock has further analyzed the conserved domain in more diverged proteins.2) We found that the three mouse PcG genes have overlapping but not identical expression patterns during embryogenesis and in adult tissues. These gene products coimmunoprecipitate from embryonic nuclear extracts. Gel filtration analysis … More of embryonic extracts indicate that these proteins exist in multiple forms as large, multimeric complexes. The heterogeneity of coimmunoprecipitates appears to be cell type-specific, as M33 and RAE28 coimmunoprecipitate and copurify as members of large complexes from F9 cells, which express BMI1 at very low levels. These studies indicate that murine PcG proteins are developmentally regulated, and function as multiple, heterogeneous complexes.3) To study the role of the rae28 gene in mouse development, we generated rae28-deficient mice by gene targeting in embryryonic stem cells. The anterior boundaries of Hoxa-3, a-4, a-5, b-3, b-4 and d-4 expression were shifted in the rostral direction in the paraxial mesoderms of the rae28^<-/-> homozygous embryos, and those of Hoxb-3 and b-4 expression were also similarly altered in the rhombomeres and/or pharyngeal arches. These altered Hox codes were presumed to be correlated with the posterior skeletal transformations and neural crest defects observed in the rae28^<-/-> homozygous mice. These results indicate that the rae28 gene is involved in the regulation of Hox gene expression and segment specification during paraxial mesoderm and neural crest development.4) We are determining if rae28 gene can rescue the corresonding fly mutations (in progress). Less

l）我们感兴趣的主要领域之一是鉴定哪些Polycomb组（PcG）蛋白相互作用，并确定果蝇和哺乳动物中的相互作用结构域。我们已经证明，三个苍蝇PcG蛋白相互作用。Psc（Posterior sex combs）与ph（polyhomeotic）和Pc（Polycomb）相互作用，但ph和Pc不相互作用。它们的小鼠同源物也是如此。RAE 28（ph）、BMI 1（Psc）和M33（Pc）同源地相互作用。RAE 28和M33都与BMI 1相互作用，但彼此不相互作用。此外，ph通过在许多蛋白质中发现的保守结构域与Scm（腿中部的性梳）相互作用。ph和Scm的小鼠同源物（RAE 28和mScm）也直接相互作用。Brock博士进一步分析了更多分歧蛋白质中的保守结构域。2）我们发现三种小鼠PcG基因在胚胎发生和成体组织中具有重叠但不相同的表达模式。这些基因产物从胚胎核提取物中共免疫沉淀。凝胶过滤分析 ...更多信息 胚胎提取物的结果表明，这些蛋白质以多种形式存在，如大的多聚体复合物。免疫共沉淀物的异质性似乎是细胞类型特异性的，因为M33和RAE 28作为来自F9细胞的大复合物的成员进行免疫共沉淀和共纯化，F9细胞以非常低的水平表达BMI 1。这些研究表明，小鼠PcG蛋白是受发育调控的，并且作为多个异质复合体发挥功能。3）为了研究rae 28基因在小鼠发育中的作用，我们通过在胚胎干细胞中进行基因靶向来产生rae 28缺陷小鼠。在rae 28 ^<-/->纯合子胚胎的近轴中胚层中，Hoxa-3、a-4、a-5、B-3、B-4和d-4表达的前边界向吻侧方向移动，在菱形体和/或咽弓中，HoxB-3和B-4表达的前边界也发生类似的改变。推测这些Hox编码的改变与rae 28 ^<-/->纯合子小鼠中观察到的后部骨骼转化和神经嵴缺陷相关。这些结果表明rae 28基因参与了近轴中胚层和神经嵴发育过程中Hox基因表达和片段特化的调控。4）我们正在确定rae 28基因是否可以拯救相应的果蝇突变（正在进行中）。少

项目成果

D.A.R.Sinclair et al.: "The additional sex combs gene of Drosophila encodes a chromatin protein that binds to shared and unique Polycomb group sites on polytene chromosomes." Development. (in press). (1998)

D.A.R.Sinclair 等人：“果蝇的额外性梳基因编码一种染色质蛋白，该蛋白与多线染色体上共享且独特的多梳基团位点结合。”

D.A.R.Sinclair et al.: "Enhancer of Polycomb is a suppressor of position-effect variegation in Drosophila melanogaster." Genetics. (in press). (1998)

D.A.R.Sinclair 等人：“Polycomb 增强剂是果蝇位置效应杂色的抑制因子。”

M.Kyba and H,W.Brock: "The SAM domain encodes a protein interaction domain that is conserved in eukaryotes." Dev.Grent.(in press). (1998)

M.Kyba 和 H,W.Brock：“SAM 结构域编码真核生物中保守的蛋白质相互作用结构域。”

S.Tanaka et al.: "Replication patterns of repetititve DNA sequences on the W-chromosome are altered during development of the chick embryo." Experimental Cell Research. 223. 233-241 (1996)

S.Tanaka 等人：“W 染色体上重复 DNA 序列的复制模式在鸡胚胎发育过程中发生了改变。”

C.Salinas et al.: "Characterization of a Drosophila homologue of the cleavage and polyadenylation specificity factor-160." Mol.Gen.Genet.(in press). (1998)

C.Salinas 等人：“果蝇裂解和聚腺苷酸化特异性因子 160 同源物的表征。”