Materials and MethodsĬells grown on poly-L-lysine-coated coverglass were fixed with 4% (w/v) paraformaldehyde in 0.1 M sodium phosphate buffer (pH 7.4) for 20 minutes at room temperature, or with ice cold acetone-methanol (1:1) solution for 2 minutes. Here, we report that condensin I and II differ in their intracellular location, the timing of their association with mitotic chromosomes, and that these complexes have distinct functions in chromosome condensation, dissociation of cohesin from chromosomes, chromosome shortening and mitotic progression. Condensin I and II are enriched at different sites along chromatid axes, and their depletion causes different morphological changes when chromosomes are treated with hypotonic buffers ( Ono et al., 2003), but their relative contribution to chromosome structure and function is poorly understood. In addition to Smc2 and Smc4, condensin II contains CAP-D3/hHCP-6, CAP-G2 and Kleisin-β/CAP-H2, proteins that are distantly related to the non-Smc subunits of the originally identified complex, now called condensin I. Condensin is also essential for cell division in budding yeast, where only a small degree of mitotic chromosome condensation is observed ( Guacci et al., 1994 Lavoie et al., 2004), and where condensin function is required for the organization and segregation of ribosomal DNA and other repetitive DNA sequences ( D'Amours et al., 2004 Freeman et al., 2000 Machin et al., 2004 Sullivan et al., 2004 Wang et al., 2004).Ĭondensin is composed of five proteins, Smc2, Smc4, CAP-D2/Eg7, CAP-G and Kleisin-γ/CAP-H (reviewed in Gassmann et al., 2004 Hagstrom and Meyer, 2003 Swedlow and Hirano, 2003), but recent work has shown that Smc2 and Smc4 are also subunits of a separate complex, called condensin II ( Ono et al., 2003 Schleiffer et al., 2003 Yeong et al., 2003). However, condensin mutants largely show normal degrees of chromosome condensation at later stages of mitosis, and the major defect observed in these mutants is abnormal anaphase ( Saka et al., 1994 Bhat et al., 1996 Giet and Glover, 2001 Steffensen et al., 2001 Hagstrom et al., 2002 Kaitna et al., 2002 Hudson et al., 2003). Consistent with this hypothesis, it has been observed that worm and chicken cells, which lack functional condensin, have chromosome-condensation defects during prophase ( Hagstrom et al., 2002 Kaitna et al., 2002 Hudson et al., 2003). This observation and the finding that depletion of condensin from Xenopus leavis egg extracts prevents chromosome assembly ( Hirano et al., 1997) implies that condensin is required for chromosome condensation. These results reveal that condensin II and I associate with chromosomes sequentially and have distinct functions in mitotic chromosome assembly.Ī different Smc-Kleisin complex, called condensin, becomes specifically enriched in axial structures of sister chromatids during mitosis ( Hirano and Mitchison, 1994 Maeshima and Laemmli, 2003 Saitoh et al., 1994). After depletion of both condensin complexes, the onset of chromosome condensation is delayed until the end of prophase, but is then initiated rapidly before nuclear envelope breakdown. ![]() By contrast, condensin I is required for the complete dissociation of cohesin from chromosome arms, for chromosome shortening and for normal timing of progression through prometaphase and metaphase, whereas normal condensin II levels are dispensable for these processes. RNA interference experiments in conjunction with imaging of live and fixed cells revealed that condensin II is required for chromosome condensation in early prophase, whereas condensin I appears to be dispensable at this stage. Here, we show that in mammalian cells condensin II associates with chromatin in prophase, in contrast to condensin I which is cytoplasmic and can thus interact with chromosomes only after nuclear envelope breakdown. In vertebrates, two types of condensin complexes have recently been identified, called condensin I and II. Condensin is a protein complex associated with mitotic chromosomes that has been implicated in chromosome condensation.
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