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Schizosaccharomyces pombe divides by means of a centrally placed division septum. The initiation of septation must be tightly coordinated with events in mitosis, as premature formation of the septum can lethally cut the undivided nucleus. The Spg1p GTPase and the Cdc7p kinase, with which it interacts, play a central role in signaling the initiation of septum formation. Loss-of-function mutations in either gene prevent septation, whereas inappropriate activation of Spg1p can induce septum formation from G1 or G2 interphase cells. Increased expression of either gene leads to multiple rounds of septation without cell cleavage, emphasizing the need for precise cell cycle regulation of their activity. To understand the mechanisms underlying this regulation, we have investigated whether these key initiators of septum formation are controlled by changes in their activity and/or location during mitosis and cytokinesis. We demonstrate that Spg1p localizes to the spindle pole body in interphase and to both spindle poles during mitosis. In contrast, Cdc7p shows no discrete localization during interphase, but early in mitosis it associates with both spindle pole bodies and, as the spindle extends, is seen on only one pole of the spindle during anaphase B. Spg1p activity is required for localization of Cdc7p in vivo but not for its kinase activity in vitro. Staining with an antiserum that recognizes preferentially GDP-Spg1p indicates that activated GTP-Spg1p predominates during mitosis when Cdc7p is associated with the spindle pole body. Furthermore, staining with this antibody shows that asymmetric distribution of Cdc7p may be mediated by inactivation of Spg1p on one spindle pole. Deregulated septation in mutant cells correlates with segregation of Cdc7p to both spindle poles.

The spg1 gene (septum-promoting GTPase) was cloned as a multicopy suppressor of a dominant-negative mutant of the Cdc7p kinase. It encodes a small GTPase of the Ras superfamily. spg1 is an essential gene. Null or heat-sensitive alleles do not make a division septum, but growth, S-phase, and mitosis continue in the absence of cell division, producing elongated, multinucleate cells. Increased expression of Spg1p induces septum formation in G2, S-phase, and pre-Start G1-arrested cells. This requires the activity of Cdc7p kinase, but not p34(cdc2). Increased expression of Cdc7p bypasses the requirement for Spg1p. Spg1p and Cdc7p can be coimmunoprecipitated from cell extracts, and interact in the two-hybrid system. These data indicate that Spg1p is a key element in controlling the onset of septum formation in Schizosaccharomyces pombe, and that it acts through the Cdc7p kinase.

Little is known about the mechanisms that establish the position of the division plane in eukaryotic cells. Wild-type fission yeast cells divide by forming a septum in the middle of the cell at the end of mitosis. Dmf1 mutants complete mitosis and initiate septum formation, but the septa that form are positioned at random locations and angles in the cell, rather than in the middle. We have cloned the dmf1 gene as a suppressor of the cdc7-24 mutant. The dmf1 mutant is allelic with mid1. The gene encodes a novel protein containing a putative nuclear localization signal, and a carboxy-terminal PH domain. In wild-type cells, Dmf1p is nuclear during interphase, and relocates to form a medial ring at the cell cortex coincident with the onset of mitosis. This relocalization occurs before formation of the actin ring and is associated with increased phosphorylation of Dmf1p. The Dmf1p ring can be formed in the absence of an actin ring, but depends on some of the genes required for actin ring formation. When the septum is completed and the cells separate, Dmf1p staining is once again nuclear. These data implicate Dmf1p as an important element in assuring correct placement of the division septum in Schizosaccharomyces pombe cells.