Localization of Fission Yeast Type II Myosin, Myo2, to the Cytokinetic Actin Ring Is Regulated by Phosphorylation of a C-Terminal Coiled-Coil Domain and Requires a Functional Septation Initiation Network

Abstract

Myo2 truncations fused to green fluorescent protein (GFP) defined a C-terminal domain essential for the localization of Myo2 to the cytokinetic actin ring (CAR). The localization domain contained two predicted phosphorylation sites. Mutation of serine 1518 to alanine (S1518A) abolished Myo2 localization, whereas Myo2 with a glutamic acid at this position (S1518E) localized to the CAR. GFP-Myo2 formed rings in the septation initiation kinase (SIN) mutant cdc7-24 at 25°C but not at 36°C. GFP-Myo2S1518E rings persisted at 36°C incdc7-24 but not in another SIN kinase mutant,sid2-250. To further examine the relationship between Myo2 and the SIN pathway, the chromosomal copy ofmyo2 + was fused to GFP (strainmyo2-gc). Myo2 ring formation was abolished in the double mutants myo2-gc cdc7.24 and myo2-gc sid2-250 at the restrictive temperature. In contrast, activation of the SIN pathway in the double mutant myo2-gc cdc16-116 resulted in the formation of Myo2 rings which subsequently collapsed at 36°C. We conclude that the SIN pathway that controls septation in fission yeast also regulates Myo2 ring formation and contraction. Cdc7 and Sid2 are involved in ring formation, in the case of Cdc7 by phosphorylation of a single serine residue in the Myo2 tail. Other kinases and/or phosphatases may control ring contraction.

Cytokinetic actomyosin ring formation and septation in fission yeast are dependent on the full recruitment of the polo-like kinase Plo1 to the spindle pole body and a functional spindle assembly checkpoint

“…Spg1 and Plo1 inappropriately drive ring formation when overproduced in interphase cells (Ohkura et al, 1995;Schmidt et al, 1997). Recruitment of Myo2 to the CAR is dependent on functional Cdc7 protein kinase (Mulvihill et al, 2000;Mulvihill et al, 2001), a key component of the SIN, which acts downstream of both Plo1 and Spg1 (Sohrmann et al, 1998;Mulvihill et al, 1999;Tanaka et al, 2001). Another component of the SIN is Cdc16, the fission yeast homologue of the spindle assembly checkpoint protein Bub2.…”

Cytokinesis Depends on the Motor Domains of Myosin-II in Fission Yeast but Not in Budding Yeast

“…Point mutations in either the Myo2p head (Naqvi et al, 1999) or tail (Mulvihill et al, 2001;Motegi et al, 2004) can compromise function. Interestingly, a Myo2p Head-GFP fusion localized at contractile rings in wild-type or myo2-E1 cells, despite being unable to substitute functionally for myo2 (Table 3 and Figure 4D).…”

The Fungal Type II Myosin in Penicillium marneffei, MyoB, Is Essential for Chitin Deposition at Nascent Septation Sites but Not Actin Localization

“…Fission yeasts position the nascent septum at the cell midpoint using an interphase negative signal from a concentration gradient of the kinase Pom1p, which concentrates at both poles (1,33,38), and the nucleus as a positional determinant, a process involving counterbalancing microtubule forces (16,51). Localization of Myo2p depends on phosphorylation in the Cterminal coiled coil and occurs in a septation initiation network (SIN)-dependent manner (39). Cdc15p, the formin Cdc12p, ring assembly protein 2 Rng2p, and Mid1p simultaneously localize with Myo2p at the cytokinesis nodes (assemblies serving as the precursors of the contractile ring).…”