RESUMO
It is considered that sister chromatids are held together immediately after replication by special protein complex--cohesin that consists of Smc1--Smc3 core dimer and two additional subunits, Scc1 and Scc3. This process is called cohesion. We have characterized binding of cohesin complex to early- and late-replicated chromatin at different stages of the cell cycle in human cells HeLa and HT1080 using superresolution microscopy (based on Structural ilumination microscopy--SIM) and immunoelectron microscopy. It has been shown that cohesins do not play important role in cohesion of heterochromatic domains, but they provide cohesion and organization of subdomains in euchromatic regions.
Assuntos
Proteínas de Ciclo Celular/química , Proteoglicanas de Sulfatos de Condroitina/química , Cromátides/metabolismo , Proteínas Cromossômicas não Histona/química , Eucromatina/metabolismo , Heterocromatina/metabolismo , Proteínas Nucleares/química , Fosfoproteínas/química , Ciclo Celular/genética , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Proteoglicanas de Sulfatos de Condroitina/genética , Proteoglicanas de Sulfatos de Condroitina/metabolismo , Cromátides/ultraestrutura , Proteínas Cromossômicas não Histona/genética , Proteínas Cromossômicas não Histona/metabolismo , Proteínas de Ligação a DNA , Eucromatina/ultraestrutura , Expressão Gênica , Células HeLa , Heterocromatina/ultraestrutura , Humanos , Microscopia Imunoeletrônica , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Ligação Proteica , Multimerização ProteicaRESUMO
Tight association of peripheral chromatin with nuclear lamina unavoidably creates topological constraints during replication. Additional complications are associated with high stability of lamina meshwork, which may hinder an access of replication factors to the sites of DNA synthesis in highly condensed template with limited mobility. In the current work we studied structural organization and dynamics of lamina as a function of replicative status of associated peripheral heterochromatin. The studies of molecular mobility of laminas at various stages of S-phase in vivo and using super-resolution microscopy showed no correlation between lamina dynamics and replicative status of attached heterochromatin. These data support the hypothesis that lamina-chromatin interactions during S-phase are regulated at the level of adapter proteins. Ultrastructural studies have demonstrated that temporal break of lamina-chromatin connections during replication does not cause noticeable spatial separation of replicating domains from nuclear periphery.
Assuntos
Replicação do DNA , DNA/metabolismo , Fibroblastos/metabolismo , Heterocromatina/metabolismo , Lâmina Nuclear/metabolismo , Animais , Células CHO , Linhagem Celular Tumoral , Cricetulus , Fibroblastos/citologia , Expressão Gênica , Heterocromatina/ultraestrutura , Humanos , Lamina Tipo A/genética , Lamina Tipo A/metabolismo , Lamina Tipo B/genética , Lamina Tipo B/metabolismo , Lâmina Nuclear/ultraestruturaRESUMO
The non-coding and repetitive sequences constitute a great amount of higher eukaryotes genomes, but the elucidation of its role and mechanisms of action is now at the very beginning. Here we found, that internal telomeric repeats in Danio rerio are colocalized with some repetitive elements, namely, hAT and EnSpm repeats, which are highly represented in vertebrate genome. While investigating one of genome regions, containing two pairs of such repeats in close proximity we found, that it is transcribed. RNA-dependent structures, containing this sequence, were revealed in D. rerio fibroblast nuclei, which may serve as evidence of functional relevance of repetitive elements in genomes or of their transcripts.
