Mouse oocytes undergo polarization during meiotic maturation, and this polarization is essential for asymmetric cell divisions that maximize retention of maternal components required for early development. Without con...Mouse oocytes undergo polarization during meiotic maturation, and this polarization is essential for asymmetric cell divisions that maximize retention of maternal components required for early development. Without conventional centrosomes, the meiotic spindle has less focused poles and is barrel-shaped. The migration of meiotic spindles to the cortex is accompanied by a local reorganization and polarization of the cortex. LGN is a conserved protein involved in cell polarity and regulation of spindle organization. In the present study, we characterized the localization dynamics of LGN during mouse oocyte maturation and analyzed the effects of LGN upregulation and downregulation on meiotic spindle organization. At the germinal vesicle stage, LGN is distributed both cytoplasmically and at the cortex. During maturation, LGN localizes to the meiotic spindle apparatus and cortical LGN becomes less concentrated at the actin cap region. Excessive LGN induces meiotic spindle organization defects by elongating the spindle and enhancing pole focusing, whereas depletion of LGN by RNA interference results in meiotic spindle deformation and chromosome misalignment. Furthermore, the N-terminus of LGN has the ability of full-length LGN to regulate spindle organization, whereas the C-terminus of LGN controls cortical localization and polarization. Our results reveal that LGN is cortically polarized in mouse oocytes and is critical for meiotic spindle organization.展开更多
Background and objective Maintenance of genomic integrity is essential to ensure normal organismal development and to prevent diseases such as cancer.PR-Set7(also known as Set8)is a cell cycle regulated enzyme that ca...Background and objective Maintenance of genomic integrity is essential to ensure normal organismal development and to prevent diseases such as cancer.PR-Set7(also known as Set8)is a cell cycle regulated enzyme that catalyses monomethylation of histone 4 at Lys20(H4K20me1)to promote chromosome condensation and prevent DNA damage.Recent studies show that CRL4CDT2-mediated ubiquitylation of PR-Set7 leads to its degradation during S phase and after DNA damage.This might occur to ensure appropriate changes in chromosome structure during the cell cycle or to preserve genome integrity after DNA damage.Methods We developed a new model of lung tumor development in mice harboring a conditionally expressed allele of Cul4A.We have therefore used a mouse model to demonstrate for the first time that Cul4A is oncogenic in vivo.With this model,staining of PR-Set7 in the preneoplastic and tumor lesions in Adeno Cre-induced mouse lungs was performed.Meanwhile we identified higher protein level changes ofγ-tubulin and pericentrin by IHC.Results The level of PR-Set7 down-regulated in the preneoplastic and adenocarcinomous lesions following over-expression of Cul4A.We also identified higher levels of the proteins pericentrin andγ-tubulin in Cul4A mouse lungs induced by Adeno Cre.Conclusion PR-Set7 is a direct target of Cul4A for degradation and involved in the formation of lung tumors in the conditional Cul4A transgenic mouse model.展开更多
微管蛋白是构成微管的主要蛋白,其中α、β亚单位形成异二聚体,而γ-微管蛋白在微管组装中起作用。为了研究小鼠早期孤雌胚中γ-微管蛋白的动态变化,本实验采用了免疫荧光化学染色与激光共聚焦显微镜观察相结合的方法,在SrCl2激活的卵...微管蛋白是构成微管的主要蛋白,其中α、β亚单位形成异二聚体,而γ-微管蛋白在微管组装中起作用。为了研究小鼠早期孤雌胚中γ-微管蛋白的动态变化,本实验采用了免疫荧光化学染色与激光共聚焦显微镜观察相结合的方法,在SrCl2激活的卵母细胞减数分裂以及早期孤雌胚有丝分裂过程中对γ-微管蛋白进行了定位观察。结果显示,SrCl2和细胞松弛素B(cytochalasin B,CB)诱导的第二次减数分裂中期(metaphaseⅡ of meiosis,MⅡ)小鼠卵母细胞恢复减数分裂,并且纺锤体始终与质膜平行,表明纺锤体旋转被抑制,但核分裂不受影响。减数分裂过程中γ-微管蛋白主要定位于中期纺锤体两极和后期分开的染色单体之间;孤雌活化两雌原核形成以后,γ-微管蛋白聚集在两雌原核周围。在早期孤雌胚有丝分裂间期无定形的γ-微管蛋白均匀分布于核;前中期γ-微管蛋白向两极移动,遍布于整个纺锤体区。有丝分裂中期、后期和末期γ-微管蛋白的分布变化与减数分裂相似。结果表明,SrCl2和CB激活的MⅡ卵母细胞产生杂合二倍体;γ-微管蛋白具有促微管负极帽形成和稳定微管的功能,从而促进纺锤体的形成;分裂后期和末期γ-微管蛋白的重新分布可能是由纺锤体牵引同源染色体分离所诱导的;γ-微管蛋白负责两雌原核的迁移靠近。展开更多
Sporogenesis in the hepatic Marchantia polymorpha L. provides an outstanding example of the pleiomorphic nature of the plant microtubule organizing center （MTOC）. Microtubules are nucleated from γ-tubuUn in MTOCs t...Sporogenesis in the hepatic Marchantia polymorpha L. provides an outstanding example of the pleiomorphic nature of the plant microtubule organizing center （MTOC）. Microtubules are nucleated from γ-tubuUn in MTOCs that change form during mitosis and meiosis. Following entry of cells into the reproductive pathway of sporogenesis, successive rounds of mitosis give rise to packets of 4-16 sporocytes. Mitotic spindles are organized at discrete polar organizers （POs）, a type of MTOC that is unique to this group of early divergent land plants. An abrupt and radical transformation in microtubule organization occurs when sporocytes enter meiosis： POs are lost and γ-tubulin is closely associated with surfaces of two large elongated plastids that subsequently divide into four. Migration of the four plastid MTOCs into a tetrahedral arrangement establishes the future spore domains and the division polarity of meiosis. As is typical of many bryophytes, cones of microtubules from the four plastid MTOCs initiate a quadripolar microtubule system （QMS） in meiotic prophase. At this point a transformation in the organization of the MTOCs occurs. The γ-tubulin detaches from plastids and forms a diffuse spheroidal pole in each of the spore domains. The plastids, which are no longer MTOCs, continue to divide. The diffuse MTOCs continue to nucleate cones of microtubules during transformation of the QMS to a bipolar spindle. Following meiosis I, γ-tubulin is associated with nuclear envelopes, and the spindles of meiosis II are organized from diffuse MTOCs at the tetrad poles. At simultaneous cytokinesis, radial microtubule systems are organized at nuclear envelope MTOCs in each of the tetrad members.展开更多
文摘Mouse oocytes undergo polarization during meiotic maturation, and this polarization is essential for asymmetric cell divisions that maximize retention of maternal components required for early development. Without conventional centrosomes, the meiotic spindle has less focused poles and is barrel-shaped. The migration of meiotic spindles to the cortex is accompanied by a local reorganization and polarization of the cortex. LGN is a conserved protein involved in cell polarity and regulation of spindle organization. In the present study, we characterized the localization dynamics of LGN during mouse oocyte maturation and analyzed the effects of LGN upregulation and downregulation on meiotic spindle organization. At the germinal vesicle stage, LGN is distributed both cytoplasmically and at the cortex. During maturation, LGN localizes to the meiotic spindle apparatus and cortical LGN becomes less concentrated at the actin cap region. Excessive LGN induces meiotic spindle organization defects by elongating the spindle and enhancing pole focusing, whereas depletion of LGN by RNA interference results in meiotic spindle deformation and chromosome misalignment. Furthermore, the N-terminus of LGN has the ability of full-length LGN to regulate spindle organization, whereas the C-terminus of LGN controls cortical localization and polarization. Our results reveal that LGN is cortically polarized in mouse oocytes and is critical for meiotic spindle organization.
基金support from the Kazan, Mc Clain, Abrams, Fernandez, Lyons, Greenwood, Harley & Oberman Foundation, Incthe Estate of Robert Griffiths+3 种基金the Jeffrey and Karen Peterson Family FoundationPaul and Michelle Zygielbaumthe Estate of Norman Mancinithe Barbara Isackson Lung Cancer Research Fund
文摘Background and objective Maintenance of genomic integrity is essential to ensure normal organismal development and to prevent diseases such as cancer.PR-Set7(also known as Set8)is a cell cycle regulated enzyme that catalyses monomethylation of histone 4 at Lys20(H4K20me1)to promote chromosome condensation and prevent DNA damage.Recent studies show that CRL4CDT2-mediated ubiquitylation of PR-Set7 leads to its degradation during S phase and after DNA damage.This might occur to ensure appropriate changes in chromosome structure during the cell cycle or to preserve genome integrity after DNA damage.Methods We developed a new model of lung tumor development in mice harboring a conditionally expressed allele of Cul4A.We have therefore used a mouse model to demonstrate for the first time that Cul4A is oncogenic in vivo.With this model,staining of PR-Set7 in the preneoplastic and tumor lesions in Adeno Cre-induced mouse lungs was performed.Meanwhile we identified higher protein level changes ofγ-tubulin and pericentrin by IHC.Results The level of PR-Set7 down-regulated in the preneoplastic and adenocarcinomous lesions following over-expression of Cul4A.We also identified higher levels of the proteins pericentrin andγ-tubulin in Cul4A mouse lungs induced by Adeno Cre.Conclusion PR-Set7 is a direct target of Cul4A for degradation and involved in the formation of lung tumors in the conditional Cul4A transgenic mouse model.
基金supported by the National Natural Science Foundation of China(No.30671025)the Returned Oversea Chinese Researcher Foundation of Science and Technology Department of Heilongjiang Province(No.LC07C17)
文摘微管蛋白是构成微管的主要蛋白,其中α、β亚单位形成异二聚体,而γ-微管蛋白在微管组装中起作用。为了研究小鼠早期孤雌胚中γ-微管蛋白的动态变化,本实验采用了免疫荧光化学染色与激光共聚焦显微镜观察相结合的方法,在SrCl2激活的卵母细胞减数分裂以及早期孤雌胚有丝分裂过程中对γ-微管蛋白进行了定位观察。结果显示,SrCl2和细胞松弛素B(cytochalasin B,CB)诱导的第二次减数分裂中期(metaphaseⅡ of meiosis,MⅡ)小鼠卵母细胞恢复减数分裂,并且纺锤体始终与质膜平行,表明纺锤体旋转被抑制,但核分裂不受影响。减数分裂过程中γ-微管蛋白主要定位于中期纺锤体两极和后期分开的染色单体之间;孤雌活化两雌原核形成以后,γ-微管蛋白聚集在两雌原核周围。在早期孤雌胚有丝分裂间期无定形的γ-微管蛋白均匀分布于核;前中期γ-微管蛋白向两极移动,遍布于整个纺锤体区。有丝分裂中期、后期和末期γ-微管蛋白的分布变化与减数分裂相似。结果表明,SrCl2和CB激活的MⅡ卵母细胞产生杂合二倍体;γ-微管蛋白具有促微管负极帽形成和稳定微管的功能,从而促进纺锤体的形成;分裂后期和末期γ-微管蛋白的重新分布可能是由纺锤体牵引同源染色体分离所诱导的;γ-微管蛋白负责两雌原核的迁移靠近。
文摘Sporogenesis in the hepatic Marchantia polymorpha L. provides an outstanding example of the pleiomorphic nature of the plant microtubule organizing center （MTOC）. Microtubules are nucleated from γ-tubuUn in MTOCs that change form during mitosis and meiosis. Following entry of cells into the reproductive pathway of sporogenesis, successive rounds of mitosis give rise to packets of 4-16 sporocytes. Mitotic spindles are organized at discrete polar organizers （POs）, a type of MTOC that is unique to this group of early divergent land plants. An abrupt and radical transformation in microtubule organization occurs when sporocytes enter meiosis： POs are lost and γ-tubulin is closely associated with surfaces of two large elongated plastids that subsequently divide into four. Migration of the four plastid MTOCs into a tetrahedral arrangement establishes the future spore domains and the division polarity of meiosis. As is typical of many bryophytes, cones of microtubules from the four plastid MTOCs initiate a quadripolar microtubule system （QMS） in meiotic prophase. At this point a transformation in the organization of the MTOCs occurs. The γ-tubulin detaches from plastids and forms a diffuse spheroidal pole in each of the spore domains. The plastids, which are no longer MTOCs, continue to divide. The diffuse MTOCs continue to nucleate cones of microtubules during transformation of the QMS to a bipolar spindle. Following meiosis I, γ-tubulin is associated with nuclear envelopes, and the spindles of meiosis II are organized from diffuse MTOCs at the tetrad poles. At simultaneous cytokinesis, radial microtubule systems are organized at nuclear envelope MTOCs in each of the tetrad members.
