Condensin-dependent localisation of topoisomerase II to an axial chromosomal structure is required for sister chromatid resolution during mitosis.
Assembly of compact mitotic chromosomes and resolution of sister chromatids are two essential processes for the correct segregation of the genome during mitosis. Condensin, a five-subunit protein complex, is thought to be required for chromosome condensation. However, recent genetic analysis suggests that condensin is only essential to resolve sister chromatids. To study further the function of condensin we have depleted DmSMC4, a subunit of the complex, from Drosophila S2 cells by dsRNA-mediated interference. Cells lacking DmSMC4 assemble short mitotic chromosomes with unresolved sister chromatids where Barren, a non-SMC subunit of the complex is unable to localise. Topoisomerase II, however, binds mitotic chromatin after depletion of DmSMC4 but it is no longer confined to a central axial structure and becomes diffusely distributed all over the chromatin. Furthermore, cell extracts from DmSMC4 dsRNA-treated cells show significantly reduced topoisomerase II-dependent DNA decatenation activity in vitro. Nevertheless, DmSMC4-depleted chromosomes have centromeres and kinetochores that are able to segregate, although sister chromatid arms form extensive chromatin bridges during anaphase. These chromatin bridges do not result from inappropriate maintenance of sister chromatid cohesion by DRAD21, a subunit of the cohesin complex. Moreover, depletion of DmSMC4 prevents premature sister chromatid separation, caused by removal of DRAD21, allowing cells to exit mitosis with chromatin bridges. Our results suggest that condensin is required so that an axial chromatid structure can be organised where topoisomerase II can effectively promote sister chromatid resolution.
Coelho
Paula A
PA
Instituto de Biologia Molecular e Celular, Universidade do Porto, Rua do Campo Alegre 823, 4150-180 Porto, Portugal.
Queiroz-Machado
Joana
J
Sunkel
Claudio E
CE
D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000818:Animals; D018797:Cell Cycle Proteins / Q000378:metabolism; D002478:Cells, Cultured; D002868:Chromosomal Proteins, Non-Histone / Q000235:genetics / Q000378:metabolism*; D002875:Chromosomes / Q000378:metabolism*; D004250:DNA Topoisomerases, Type II / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D004330:Drosophila / Q000378:metabolism; D029721:Drosophila Proteins / Q000378:metabolism; D018386:Kinetochores / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008938:Mitosis / Q000502:physiology; D046912:Multiprotein Complexes; D021122:Protein Subunits / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D008941:Spindle Apparatus / Q000378:metabolism
D018797:Cell Cycle Proteins; D002868:Chromosomal Proteins, Non-Histone; D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D046912:Multiprotein Complexes; D021122:Protein Subunits; D034741:RNA, Small Interfering; C108312:condensin complexes; C410884:vtd protein, Drosophila; D000251:Adenosine Triphosphatases; D004250:DNA Topoisomerases, Type II