A calmodulin-related light chain from fission yeast that functions with myosin-I and PI 4-kinase

ArticleinJournal of Cell Science 124(Pt 14):2466-77 · July 2011with10 Reads
DOI: 10.1242/jcs.067850 · Source: PubMed
Fission yeast myosin-I (Myo1p) not only associates with calmodulin, but also employs a second light chain called Cam2p. cam2Δ cells exhibit defects in cell polarity and growth consistent with a loss of Myo1p function. Loss of Cam2p leads to a reduction in Myo1p levels at endocytic patches and a 50% drop in the rates of Myo1p-driven actin filament motility. Thus, Cam2p plays a significant role in Myo1p function. However, further studies indicated the existence of an additional Cam2p-binding partner. Cam2p was still present at cortical patches in myo1Δ cells (or in myo1-IQ2 mutants, which lack an intact Cam2p-binding motif), whereas a cam2 null (cam2Δ) suppressed cytokinesis defects of an essential light chain (ELC) mutant known to be impaired in binding to PI 4-kinase (Pik1p). Binding studies revealed that Cam2p and the ELC compete for Pik1p. Cortical localization of Cam2p in the myo1Δ background relied on its association with Pik1p, whereas overexpression studies indicated that Cam2p, in turn, contributes to Pik1p function. The fact that the Myo1p-associated defects of a cam2Δ mutant are more potent than those of a myo1-IQ2 mutant suggests that myosin light chains can contribute to actomyosin function both directly and indirectly (via phospholipid synthesis at sites of polarized growth).
    • "Yeast strains used in this study are listed in supplementary materialTable S1. All strains were constructed by genomic integrations using homologous recombination (Bähler et al., 1998; Sammons et al., 2011) and genetic crosses. The myo1-E1 (G308R) and myo1-S1 (G483D) mutants were generated previously (Stark et al., 2013) based on the sequence homology with myo2-E1 and myo2-S1 mutants (Balasubramanian et al., 1998; Wong et al., 2000). "
    [Show abstract] [Hide abstract] ABSTRACT: Point mutations in the human MYO1E gene, encoding class I myosin Myo1e, are associated with focal segmental glomerulosclerosis (FSGS), a primary kidney disorder that leads to end stage kidney disease. In this study, we used a simple model organism, fission yeast Schizosaccharomyces pombe, to test the effects of FSGS-associated mutations on myosin activity. Fission yeast has only one class I myosin, Myo1, which is involved in actin patch assembly at the sites of endocytosis. The amino acid residues mutated in the FSGS patients are conserved between human Myo1e and yeast Myo1, which allowed us to introduce equivalent mutations into yeast myosin and use the resulting mutant strains for functional analysis. Yeast strains expressing mutant Myo1 exhibited defects in growth and endocytosis similar to those observed in the myo1 deletion strain. These mutations also disrupted Myo1 localization to endocytic actin patches and resulted in mis-localization of Myo1 to eisosomes, linear membrane microdomains found in yeast cells. While both mutants examined in this study exhibited loss of function, one of these mutants was also characterized by the decreased protein stability. Thus, using the yeast model system we were able to determine that the kidney disease-associated mutations impair myosin functional activity and have differential effects on protein stability. © 2015. Published by The Company of Biologists Ltd.
    Full-text · Article · Jun 2015
    • "Scale bar represents 2 mm. genes required for membrane traffic through the Golgi apparatus (Park et al., 2009; Sammons et al., 2011; Walch-Solimena and Novick, 1999). Similarly, mutations in conserved genes required for vesicular transport in the Golgi apparatus (De Matteis and Luini, 2008; Jaulin et al., 2007; Valente et al., 2012 ) cause septation defects in fission yeast (Brazer et al., 2000; Mishra et al., 2005). "
    [Show abstract] [Hide abstract] ABSTRACT: Cdc15p is known to contribute to cytokinesis in fission yeast; however, the protein is not required to assemble the contractile ring of actin and myosin, but it helps to anchor the ring to the plasma membrane. Cdc15p has a lipid-binding F-BAR domain, suggesting that it provides a physical link between the plasma membrane and contractile ring proteins. However, we find that a more important function of Cdc15p during cytokinesis is to help deliver a transmembrane enzyme, Bgs1p (also called Cps1p), from the Golgi apparatus to the plasma membrane, where it appears to anchor the contractile ring. Bgs1p synthesizes the cell wall in the cleavage furrow, but its enzyme activity is not required to anchor the contractile ring. We estimate that ∼2,000 Bgs1p molecules are required to anchor the ring. Without Bgs1p anchors, contractile rings slide along the plasma membrane, a phenomenon that depends on an unconventional type II myosin called Myp2p.
    Full-text · Article · Aug 2014
    • "Indeed, the PtdIns 4-kinase PIK1 is required for late cytokinesis events [Park et al., 2009], endocytosis, post-Golgi trafficking [Walch-Solimena and Novick, 1999], and interacts with the myosin II-light chains Cdc4p and Cam2p [Desautels et al., 2001; Sammons et al., 2011]. Mutant Cdc4p unable to interact with PIK1 cannot sustain cytokinesis, indicating that interaction between the myosin light chains and the PtdIns 4-kinase may be important for actin ring function, perhaps by promoting localized PtdIns4P production at the furrow [Desautels et al., 2001; Sammons et al., 2011]. It is unclear whether this complex is conserved, as a direct interaction between myosin II light chain and Fwd has not been detected in flies [Polevoy et al., 2009]. "
    [Show abstract] [Hide abstract] ABSTRACT: Phosphoinositides [Phosphatidylinositol (PtdIns), phosphatidylinositol 3-monophosphate (PtdIns3P), phosphatidylinositol 4-monophosphate (PtdIns4P), phosphatidylinositol 5-monophosphate (PtdIns5P), phosphatidylinositol 3,4-bisphosphate (PtdIns(3,4)P(2) ), phosphatidylinositol 3,5-bisphosphate (PtdIns(3,5)P(2) ), phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P(2) ), and phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P(3) )] are lowly abundant acidic lipids found at the cytosolic leaflet of the plasma membrane and intracellular membranes. Initially discovered as precursors of second messengers in signal transduction, phosphoinositides are now known to directly or indirectly control key cellular functions, such as cell polarity, cell migration, cell survival, cytoskeletal dynamics, and vesicular traffic. Phosphoinositides actually play a central role at the interface between membranes and cytoskeletons and contribute to the identity of the cellular compartments by recruiting specific proteins. Increasing evidence indicates that several phosphoinositides, particularly PtdIns(4,5)P(2) , are essential for cytokinesis, notably after furrow ingression. The present knowledge about the specific phosphoinositides and phosphoinositide modifying-enzymes involved in cytokinesis will be first presented. The review of the current data will then show that furrow stability and cytokinesis abscission require that both phosphoinositide production and hydrolysis are regulated in space and time. Finally, I will further discuss recent mechanistic insights on how phosphoinositides regulate membrane trafficking and cytoskeletal remodeling for successful furrow ingression and intercellular bridge abscission. This will highlight unanticipated connections between cytokinesis and enzymes implicated in human diseases, such as the Lowe syndrome. © 2012 Wiley Periodicals, Inc.
    Article · Nov 2012
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