RESUMO
Lamins are the major components of the nuclear lamina, a filamentous layer underlying the inner nuclear membrane and attached to the peripheral chromatin. Lamins are required for maintaining nuclear shape and are involved in most nuclear activities. Here, we studied the 3D organization of the nuclear lamina formed upon the expression of Caenorhabditis elegans lamin (Ce-lamin) within the nucleus of a Xenopus laevis oocyte. We show that Ce-lamin forms an intricate 3D meshwork of 5-6 nm lamin protofilaments. The diverse protofilament interactions and organization may shed light upon the unique mechano-elastic properties of the nuclear lamina scaffold supporting the nuclear envelope. The Q159K Hutchinson-Gilford Progeria Syndrome-linked mutation alters interactions between protofilaments within the lamina, leading to the formation of more bundled arrays of less isotropically-oriented protofilaments. Using this system, we show for the first time the organization of lamin proteins that were translated and assembled within the environment of a living cell.
Assuntos
Caenorhabditis elegans/química , Caenorhabditis elegans/genética , Citoesqueleto/química , Laminas/química , Oócitos/química , Animais , Citoesqueleto/genética , Feminino , Regulação da Expressão Gênica , Processamento de Imagem Assistida por Computador , Laminas/genética , Microscopia Eletrônica de Varredura , Lâmina Nuclear/química , Lâmina Nuclear/genética , Estrutura Terciária de Proteína , Xenopus laevis/genética , Xenopus laevis/metabolismoRESUMO
The nuclear pore complex (NPC) is the sole gateway between the nucleus and the cytoplasm. NPCs fuse the inner and outer nuclear membranes to form aqueous translocation channels that allow the free diffusion of small molecules and ions, as well as receptor-mediated transport of large macromolecules. The NPC regulates nucleocytoplasmic transport of macromolecules, utilizing soluble receptors that identify and present cargo to the NPC, in a highly selective manner to maintain cellular functions. The NPC is composed of multiple copies of approximately 30 different proteins, termed nucleoporins, which assemble to form one of the largest multiprotein assemblies in the cell. In this review, we address structural and functional aspects of this fundamental cellular machinery.
Assuntos
Complexo de Proteínas Formadoras de Poros Nucleares/metabolismo , Poro Nuclear/química , Poro Nuclear/metabolismo , Transporte Ativo do Núcleo Celular , Animais , Núcleo Celular/metabolismo , Citoplasma/metabolismo , Humanos , Membrana Nuclear/metabolismo , Poro Nuclear/ultraestrutura , Complexo de Proteínas Formadoras de Poros Nucleares/química , Transporte ProteicoRESUMO
Visualization of cellular processes at a resolution of the individual protein should involve integrative and complementary approaches that can eventually draw realistic functional and cellular landscapes. Electron tomography of vitrified but otherwise unaltered cells emerges as a central method for three-dimensional reconstruction of cellular architecture at a resolution of 2-6 nm. While a combination of correlative light-based microscopy with cryo-electron tomography (cryo-ET) provides medium-resolution insight into pivotal cellular processes, fitting high-resolution structural approaches, for example, X-ray crystallography, into reconstructed macromolecular assemblies provides unprecedented information on native protein assemblies. Thus, cryo-ET bridges the resolution gap between cellular and structural biology. In this article, we focus on the study of eukaryotic cells and macromolecular complexes in a close-to-life-state. We discuss recent developments and structural findings enabling major strides to be made in understanding complex physiological functions.