Cell Architecture Laboratory • Kimura Group

The mitotic spindle is a diamond-shaped molecular apparatus crucial for chromosomal segregation. Previous studies suggested that the spindle can self-organize to maintain a constant aspect ratio between its length and width against physical perturbations. Here we determine the widths of metaphase spindles of various sizes observed during embryogenesis in Caenorhabditis elegans. The spindle width correlates well with the spindle length, but the aspect ratio between the spindle length and spindle width is not constant, indicating an allometric relationship between these parameters. We characterize how DNA quantity (ploidy) affects spindle shape by using haploid and polyploid embryos. On the basis of the quantitative data, we deduce an allometric equation that describes the spindle width as a function of the length of the hypotenuse and ploidy. On the basis of this equation, we propose a force-balance model to determine the spindle width.

(A) Metaphase spindles at 4 representative cell stages (Bar = 5 μm). (B) The spindle width (pink), half-spindle length (light blue), and hypotenuse length (orange) were shown. (C) The spindle width from diploid (blue diamond), haploid (yellow circle), and polyploid (green triangles) is plotted against the product of 0.36th-power of ploidy and 0.58th-power of hypotenuse length. (Figures were modified from the paper.)

Division of Microbial Genetics • Araki Group

Dpb11/Cut5/TopBP1 is evolutionarily conserved and is essential for the initiation of DNA replication in eukaryotes. The Dpb11 of the budding yeast Saccharomyces cerevisiae has four BRCT domains (BRCT1 to -4). The N-terminal pair (BRCT1 and -2) and the C-terminal pair (BRCT3 and -4) bind to cyclin-dependent kinase (CDK)-phosphorylated Sld3 and Sld2, respectively. These phosphorylation-dependent interactions trigger the initiation of DNA replication. BRCT1 and -2 and BRCT3 and -4 of Dpb11 are separated by a short stretch of ~100 amino acids. It is unknown whether this inter-BRCT region functions in DNA replication. Here, we showed that the inter-BRCT region is a GINS interaction domain that is essential for cell growth and that mutations in this domain cause replication defects in budding yeast. We found the corresponding region in the vertebrate ortholog, TopBP1, and showed that the corresponding region also interacts with GINS and is required for efficient DNA replication. We propose that the inter-BRCT region of Dpb11 is a functionally conserved GINS interaction domain that is important for the initiation of DNA replication in eukaryotes.

Schematic drawings of the initiation of DNA replication in eukaryotes. Dpb11-GINS interaction is important for the assembly of protein complex, which is required for the activation of replicative helicase.