The Nuclear Envelope in Development, Disease and Ageing

脊椎动物核膜是一种与内核膜核面相关的蛋白质网络。它为染色质结构域提供锚定位点，是间期核结构、DNA复制和染色质组织的重要决定因素。板层的主要成分是中间的丝状蛋白，核板层分为A型和B型两类。B型Lamins由2个基因编码，由2个基因组成表达，而A型lamins是单个基因(Lmna)的剪接变体，其表达受发育调节。它们在早期胚胎或成体干细胞中不表达，其表达与不同谱系的终末分化有关。我们得到的小鼠不表达A-型层蛋白。拉明缺失小鼠的发育是明显正常的，但在4-5周后，它们出现了严重的肌肉营养不良，心脏异常，并在8周时死亡。在人类中，Lamin A基因的不同突变至少导致了8种遗传性疾病。其中包括两种形式的肌营养不良症、扩张型心肌病、两种类型的家族性部分脂肪营养不良，其中一种也会影响骨骼发育，一种周围神经病变，以及最近被称为Hutchinson Gilford Progeria的过早衰老状况。我们已经获得了Lmna基因突变的小鼠，这些突变对应于大多数这些疾病。我们已经培育出了出现肌肉营养不良、扩张型心肌病和孕激素表型的小鼠品系。患有早衰症的小鼠为衰老的基本方面以及细胞核结构如何发育调节细胞增殖提供了新的见解。此外，我们还设计了一种突变小鼠，该突变小鼠携带另一种亚型的Lamin A/C蛋白，称为N195K点突变，在人类中与扩张型心肌病(DCM)有关。尽管纯合子N195K/N195K小鼠似乎比最初衍生的Lamin A/C缺失组小鼠寿命更长，也没有明显的肌肉脂肪消耗，但这些动物最终在4个月大时死于急性心力衰竭，先于心脏异常节律，与人类DCM患者的临床表现相似。我们还能够通过传统的loxP/Cre系统获得一种小鼠系，允许有条件地消除Lamin A/C基因。这些动物通过与一系列表达Cre的转基因品系杂交，提供了一种研究特定组织亚群和/或以诱导方式研究核层功能的替代方案，从而绕过了在Lamin A/C基因缺失小鼠中观察到的早期致死性。我们在实验室中利用该小鼠品系研究了Lamin A/C缺失对毛发周期(通过与K14-Cre转基因小鼠杂交)和骨巢环境中免疫系统细胞成分成熟的影响(通过与Col2a-Cre转基因小鼠交配)。总之，对椎板病变的分析为了解细胞核结构对其功能的重要性提供了新的信息。这一点特别重要，因为除了与椎板相关的疾病(椎板病)外，至少还有两种其他疾病与核膜相关蛋白质的突变有关。此外，最近对核膜成分的蛋白质组学分析表明，另外14种疾病也可能与核膜蛋白的改变有关。除了这些疾病外，我们还在研究Lamin缺乏对细胞复制DNA的能力、染色体分离/定位和染色质组织/基因调节的影响，以及Lamin缺失是否会影响肿瘤的发展。

The vertebrate nuclear lamina is a protein meshwork associated with the nuclear face of the inner nuclear membrane (INM). It provides anchoring sites for chromatin domains, and is an important determinant of interphase nuclear architecture, DNA replication and chromatin organization. The major components of the lamina are the intermediate filament-like proteins, the nuclear lamins, The lamins are grouped into 2 classes, A-type and B-type. The B-type lamins are encoded by 2 genes and are constitutively expressed whereas the A-type lamins are spliced variants from a single gene (Lmna) and their expression is developmentally regulated. They are not expressed in early embryos or adult stem cell and their expression correlates with the terminal differentiation of various lineages. We derived mice that did not express the A-type lamins. Development of the Lamin null mice was overtly normal, but by 4-5 weeks they developed a severe form of muscular dystrophy, had abnormal hearts and were dead by 8 weeks. In humans different mutations in the Lamin A gene are responsible for at least 8 inherited diseases. These include 2 forms of muscular dystrophy, dilated cardiomyopathy, 2 types of Familial Partial Lipodystrohy, one of which also affects skeletal development, a peripheral neuropathy and most recently, the premature ageing condition called Hutchinson Gilford Progeria. We have derived mice with mutations in the Lmna gene that correspond to most of these diseases. We have produced mouse lines that develop muscular dystrophy, dilated cardiomyopathy, and a progeric phenotype. The mice with progeria are providing novel insights into fundamental aspects of aging and how the structure of the nucleus developmentally regulates cell proliferation. In addition, we have engineered a mutant mouse line harboring yet another isoform of Lamin A/C protein, known as N195K point mutation and associated in humans with cases of dilated cardiomyopathy (DCM). Although the homozygous N195K/N195K mice appeared to live longer and developed no visible muscular wasting of fat loss compared to the initially derived Lamin A/C null animals, these animals ultimately succumb by the age of 4 months due to acute heart failure preceded by cardiac aberrant rhythm similar to the clinical findings in human DCM patients. We were also able to derive a mouse line allowing for a conditional elimination of Lamin A/C gene via a conventional loxP/Cre system. These animals provide an alternative of studying the nuclear lamina functioning in specific subsets of tissues and/or in an inducible manner by crossing with a spectrum of Cre-expressing transgenic lines thus bypassing the early lethality observed in Lamin A/C null mice. We exploited this mouse strain in the lab to study the effect of Lamin A/C loss on the hair cycle (via crossing with K14-Cre transgenic line) and on the maturation of immune system cellular components in the bone niche environment harboring the hematopoietic stem cells (via mating with Col2a-Cre transgenic mice). In summary, the analysis of the laminopathies is providing novel information into how the structure of the nucleus is important to its function. This is particularly relevant, as in addition to the diseases associated with the lamins (the laminopathies), at least 2 other disease have been linked to mutations in proteins associated with the nuclear envelope. Furthermore, a recent proteomics analysis of the components of the nuclear envelope suggested that an additional 14 diseases might also be linked to altered nuclear envelope proteins. In addition to these diseases we are also looking at what effects Lamin deficiency has on a cells ability to replicate its DNA, chromosome segregation/location and chromatin organization/gene regulation and whether lamin loss may affect tumor development.