Insight into Actin Organization and Function in Cytokinesis from Analysis of Fission Yeast Mutants

Abstract

Actin is a key cytoskeletal protein with multiple roles in cellular processes such as polarized growth, cytokinesis, endocytosis, and cell migration. Actin is present in all eukaryotes as highly dynamic filamentous structures, such as linear cables and branched filaments. Detailed investigation of the molecular role of actin in various processes has been hampered due to the multifunctionality of the protein and the lack of alleles defective in specific processes. The actin cytoskeleton of the fission yeast, Schizosaccharomyces pombe, has been extensively characterized and contains structures analogous to those in other cell types. In this study, primarily with the view to uncover actin function in cytokinesis, we generated a large bank of fission yeast actin mutants that affect the organization of distinct actin structures and/or discrete physiological functions of actin. Our screen identified 17 mutants with specific defects in cytokinesis. Some of these cytokinesis mutants helped in dissecting the function of specific actin structures during ring assembly. Further genetic analysis of some of these actin mutants revealed multiple genetic interactions with mutants previously known to affect the actomyosin ring assembly. We also characterize a mutant allele of actin that is suppressed upon overexpression of Cdc8p-tropomyosin, underscoring the utility of this mutant bank. Another 22 mutant alleles, defective in polarized growth and/or other functions of actin obtained from this screen, are also described in this article. This mutant bank should be a valuable resource to study the physiological and biochemical functions of actin.A CTIN is a small globular protein (42 kDa) and is among the most conserved proteins in eukaryotes. In cells, actin transitions between the monomeric G-actin and filamentous F-actin forms that are regulated by the binding of ATP and the hydrolysis of ATP to ADP. The actin cytoskeleton in most eukaryotes comprises two major kinds of filamentous structures: Formin-nucleated unbranched linear filaments and the Arp2/3 complex-nucleated branched filaments (Pollard 2007). Over 100 different accessory proteins function in eukaryotic cells to regulate the assembly, turnover, and function of actin (Pollard and Borisy 2003). The various F-actin structures play important roles in different cellular processes such as polarized growth, targeted secretion, endocytosis, and cytokinesis (Chhabra and Higgs 2007).Although actin has been extensively studied over decades, the precise molecular basis of each distinct function of actin still remains unclear. While inhibitory drugs such as Latrunculin A (LatA), Cytochalasin A (CytA), and Cytochalasin D (CytD) that affect the dynamics of actin monomers and/or polymers have provided valuable insights, these inhibitors affect all aspects of actin function (Cooper 1987;Ayscough et al. 1997;Allingham et al. 2006;Uribe and Jay 2009). One approach to characterize distinct functions of actin involves the generation of mutant alleles specifically defective in ind...

The formins Cdc12 and For3 cooperate during contractile ring assembly in cytokinesis

“…The contractile ring containing actin filaments, myosin II, and many other proteins is required for cytokinesis in fungi and animal cells (Balasubramanian et al, 2004;Barr and Gruneberg, 2007;Pollard and Wu, 2010). Both de novo actin assembly at the division site and cortical transport/flow contribute actin filaments to the contractile ring (White and Borisy, 1983;Bray and White, 1988;Cao and Wang, 1990;Lee et al, 1998;Pelham and Chang, 2002;Chen et al, 2008;Zhou and Wang, 2008;Alsop et al, 2009;Huang et al, 2012;Subramanian et al, 2013). However, the relative importance of these sources of actin filaments for the contractile ring is unknown in any cell type.…”

Proteomic analysis of crevicular fluid of traumatized primary teeth

“…Notwithstanding, oxidative stress can induce not only damage as apoptosis and cell death, but also responses as transformation, adaptation and repair (Forman et al 2004;Navarro-Yepes et al 2014). It is worth highlighting the proteins representative of actin, which are related to several metabolism process as: migration, signaling, endocytosis, cell motility, cellular morphogenesis, cell division, adhesion, cytokinesis, polarized growth, and cell migration (Janji et al 2006;Gandhi and Goode 2013;Subramanian et al 2013;Blanchoin et al 2014). Therefore, all those cellular activities are in accordance to homeostatic process.…”

Interacciones de la GTPasa Rho1 con la ruta de MAPKs de integridad celular y la fosfatasa calcineurina en la levadura de fisión Schizosaccharomyces pombe

“…En esta levadura el citoesqueleto de actina participa en distintos e importantes procesos celulares tales como el crecimiento polarizado, la secreción dirigida, la endocitosis y la citoquinesis (Chhabra y Higgs, 2007;Subramanian, et al, 2013). La actina es una pequeña proteína globular (~42 kDa) y una de las más conservadas en eucariotas.…”

“…La actina es una pequeña proteína globular (~42 kDa) y una de las más conservadas en eucariotas. En las células, la actina aparece tanto en su forma monomérica o G-actina como en su forma filamentosa o F-actina (Subramanian, et al, 2013). En S. pombe, está presente en diferentes estructuras que conforman el citoesqueleto: parches, que contienen filamentos de actina ramificada, cables compuestos por filamentos lineales de actina sin ramificar y el anillo de citoquinesis (Subramanian, et al, 2013) (Figura 6).…”