Checking the Daughter Cell's Wall

Abstract

Suzuki et al. report a new checkpoint pathway for stopping mitosis if cell wall synthesis is impaired in the emerging daughter cell of the budding yeast, Saccharomyces cerevisiae . The authors constructed yeast with a temperature-sensitive version of the ß(1→3) glucan synthase gene ( fks1-1154 ) and observed lack of full-sized buds and a block in mitosis after DNA replication but before separation of the spindle poles and chromosomes. The abundance of the mitotic cyclin, Clb2, was diminished at the nonpermissive temperature, and overexpression of Clb2 could overrride the defect in spindle pole separation. In a screen for checkpoint-deficient mutants, a gene encoding a subunit of the dynactin complex was identified: ARP1 , which encodes an actin-related protein (the yeast mutant was named wac1 for wall checkpoint defective). The fks1-1154 cells that also carried a mutation in ARP1 exhibited bipolar spindle formation, accumulation of Clb2, and activation of the Clb2-associated cyclin-dependent kinase (CDK), Cdc28. Deletion of the gene encoding the forkhead transcription factor Fkh2 in the fks1-1154, wac1 background resulted in loss of the formation of the bipolar spindles; thus, Fkh2, a known regulator of CLB2 expression, is downstream of ARP1 in the cell wall-integrity checkpoint pathway. Indeed, overexpression of FKH2 permitted bipolar spindle formation and accumulation of Clb2 in the fks1-1154 cells at the nonpermissive temperature. Deletion of genes encoding other subunits of the dynactin complex also resulted in yeast with a deficient checkpoint, which suggests that ARP1 acts within the context of the dynactin complex. However, deletion of genes encoding subunits of dynein, the microtubule motor that is regulated by dynactin, did not exhibit cell wall integrity checkpoint defects, suggesting that dynactin's role in monitoring the cell wall during mitosis is independent from its role in regulating dynein activity. Genetic analysis of various combinations of mutants demonstrated that the cell wall integrity checkpoint pathway was separate from other established checkpoint pathways, such as the morphogenesis checkpoint pathway that acts through the kinase Swe1 to inhibit the CDK, Cdc28. M. Suzuki, R. Igarashi, M. Sekiya, T. Utsugi, S. Morishita, M. Yukawa, Y. Ohya, Dynactin is involved in a checkpoint to monitor cell wall synthesis in Saccharomyces cerevisiae . Nat. Cell Biol. 6 , 861-871 (2004). [Online Journal] M. A. Hoyt, A new checkpoint takes shape. Nat. Cell Biol. 6 , 801-803 (2004). [Online Journal]