HIRA-like (Hir) proteins are evolutionarily conserved and are implicated in the assembly of repressive chromatin. In Saccharomyces cerevisiae, Hir proteins contribute to the function of centromeres. However, S. cerevisiae has point centromeres that are structurally different from the complex centromeres of metazoans. In contrast, Schizosaccharomyces pombe has complex centromeres whose domain structure is conserved with that of human centromeres. Therefore, we examined the functions of the fission yeast Hir proteins Slm9 and the previously uncharacterised protein Hip1. Deletion of hip1 ؉ resulted in phenotypes that were similar to those described previously for slm9⌬ cells: a cell cycle delay, synthetic lethality with cdc25-22, and poor recovery from nitrogen starvation. However, while it has previously been shown that Slm9 is not required for the periodic expression of histone H2A, we found that loss of Hip1 led to derepression of core histone genes expression outside of S phase. Importantly, we found that deletion of either hip1 ؉ or slm9 ؉ resulted in increased rates of chromosome loss, increased sensitivity to spindle damage, and reduced transcriptional silencing in the outer centromeric repeats. Thus, S. pombe Hir proteins contribute to pericentromeric heterochromatin, and our data thus suggest that Hir proteins may be required for the function of metazoan centromeres.

In eukaryotes the regulation of gene expression plays a key role in controlling cell cycle progression. Here, we demonstrate that a forkhead transcription factor, Fkh2, regulates the periodic expression of cdc15 ؉ and spo12 ؉ in the M and G 1 phases of the cell division cycle in the fission yeast Schizosaccharomyces pombe. We also show that Fkh2 is important for several cell cycle processes, including cell morphology and cell separation, nuclear structure and migration, and mitotic spindle function. We find that the expression of fkh2 ؉ is itself regulated in a cell cycle-dependent manner in G 1 coincident with the expression of cdc18 ؉ , a Cdc10-regulated gene. However, fkh2؉ expression is independent of Cdc10 function. Fkh2 was found to be phosphorylated during the cell division cycle, with a timing that suggests that this posttranslational modification is important for cdc15؉ and spo12 ؉ expression. Related forkhead proteins regulate G 2 and M phase-specific gene expression in the evolutionarily distant Saccharomyces cerevisiae, suggesting that these proteins play conserved roles in regulating cell cycle processes in eukaryotes.Forkhead transcription factors in eukaryotes have been implicated in a number of cellular processes including embryogenesis, development, and the cell division cycle. Forkhead proteins contain a conserved region spanning a monomeric DNA binding (FKH) domain (for reviews, see references 26 and 31). Indeed, a core DNA consensus sequence has been identified to which many forkhead proteins bind (27). Mutations in genes encoding forkhead proteins have been linked with several pathological conditions including thyroid agenesis (11), cleft palate (11), glaucoma (38), speech development defects (30), and tumorigenesis (16,32,39).In eukaryotes the periodic expression of genes plays an important role in the regulation of the cell division cycle. Global approaches in the budding yeast Saccharomyces cerevisiae have provided insight into the groups of genes that are expressed in this manner and also the transcription factors responsible for this cell cycle-dependent expression (10,44,47). The MBF (MluI binding factor), consisting of Mbp1 and Swi6, and the related SBF (Swi4/Swi6 cell cycle box binding factor), consisting of Swi4 and Swi6, transcription factor complexes are important for regulation of genes that are expressed early in the cell division cycle (see reference 51 for a review). In contrast, the forkhead proteins Fkh1 and Fkh2 regulate cell cycle-dependent expression of a group of genes, named the CLB2 gene cluster, that includes CLB1 and CLB2, encoding mitotic cyclins (18, 48), in the G 2 and M phases (28,29,40,53). Regulation of the CLB2 gene cluster involves the formation of a ternary complex between Fkh2 and the MADS-box protein, Mcm1, and the coactivator Ndd1 (28, 29, 40, 53; see references 8, 17, and 25 for reviews). The altered expression of the CLB2 gene cluster in a fkh1⌬ fkh2⌬ mutant is associated with defects in the structure of the mitotic spindle, nuclear migration, cell morp...