Myosin localization during meiosis I of crane‐fly spermatocytes gives indications about its role in division

Abstract

We showed previously that in crane-fly spermatocytes myosin is required for tubulin flux [Silverman-Gavrila and Forer, 2000a: J Cell Sci 113:597-609], and for normal anaphase chromosome movement and contractile ring contraction [Silverman-Gavrila and Forer, 2001: Cell Motil Cytoskeleton 50:180-197]. Neither the identity nor the distribution of myosin(s) were known. In the present work, we used immunofluorescence and confocal microscopy to study myosin during meiosis-I of crane-fly spermatocytes compared to tubulin, actin, and skeletor, a spindle matrix protein, in order to further understand how myosin might function during cell division. Antibodies to myosin II regulatory light chain and myosin II heavy chain gave similar staining patterns, both dependent on stage: myosin is associated with nuclei, asters, centrosomes, chromosomes, spindle microtubules, midbody microtubules, and contractile rings. Myosin and actin colocalization along kinetochore fibers from prometaphase to anaphase are consistent with suggestions that acto-myosin forces in these stages propel kinetochore fibres poleward and trigger tubulin flux in kinetochore fibres, contributing in this way to poleward chromosome movement. Myosin and actin colocalization at the cell equator in cytokinesis, similar to studies in other cells [e.g., Fujiwara and Pollard, 1978: J Cell Biol 77:182-195], supports a role of actin-myosin interactions in contractile ring function. Myosin and skeletor colocalization in prometaphase spindles is consistent with a role of these proteins in spindle formation. After microtubules or actin were disrupted, myosin remained in spindles and contractile rings, suggesting that the presence of myosin in these structures does not require the continued presence of microtubules or actin. BDM (2,3 butanedione, 2 monoxime) treatment that inhibits chromosome movement and cytokinesis also altered myosin distributions in anaphase spindles and contractile rings, consistent with the physiological effects, suggesting also that myosin needs to be active in order to be properly distributed.

Do nuclear envelope and intranuclear proteins reorganize during mitosis to form an elastic, hydrogel-like spindle matrix?

“…Filamentous actin has been studied and is present near matrix proteins in some species (Fabian et al 2007a;Fabian and Forer 2007) but little is known about total actin in spindles. Myosin is present in spindles (for example, Silverman-Gavrila and Forer 2003;Forer 2005, 2007;Woolner et al 2008;Sandquist and Bement 2010) and phosphorylated (active) myosin is concentrated in the spindle, near kinetochore fibers (Fabian et al 2007a, b). Thus, the limited data are consistent with actin and myosin interacting with or being part of the spindle matrix.…”

Continuous phosphatidylinositol metabolism is required for cleavage of crane fly spermatocytes

“…In control cells, cortical actin filaments are arranged longitudinally around the cell until mid-to lateanaphase; after this, perpendicularly oriented filaments appear in the equatorial region. Subsequently, the filaments form increasingly compact equatorial bundles during telophase as the cells complete cytokinesis (Forer and Behnke, 1972;Silverman-Gavrila and Forer, 2003) (Fig. 9A-C).…”

Gamma-actin is involved in regulating centrosome function and mitotic progression in cancer cells

“…The role of actin and its regulatory proteins in these processes ranges from regulating centrosome separation 18,19 to proper spindle assembly and orientation [6][7][8]12,13,[15][16][17] to elongation of kinetochore microtubules. 11,20 Importantly, disruption of the actin cytoskeleton affects mitotic and/or meiotic spindle structure and function, resulting in mitotic defects [6][7][8][11][12][13]20 and/or meiotic defects. [15][16][17] These studies provide evidence of how critically important actin and its regulatory proteins are in regulating cell division.…”