dictyNews Electronic Edition Volume 29, number 15 November 30, 2007 Please submit abstracts of your papers as soon as they have been accepted for publication by sending them to dicty@northwestern.edu or by using the form at http://dictybase.org/db/cgi-bin/dictyBase/abstract_submit. Back issues of dictyNews, the Dicty Reference database and other useful information is available at dictyBase - http://dictybase.org. ========= Abstracts ========= A novel mitosis-specific dynamic actin structure in Dictyostelium cells Go Itoh and Shigehiko Yumura J. Cell Sci., in press Cell division of various animal cells depends on their attachment to a substratum. Dictyostelium cells deficient in type II myosin, analogous to myosin in muscle, can divide on a substratum without the contractile ring. To investigate the mechanism of this substratum-dependent cytokinesis, the dynamics of actin in the ventral cortex were observed by confocal and total internal reflection fluorescence microscopy. Specifically during mitosis, we found novel actin-containing structures (mitosis-specific dynamic actin structures, MiDASes) underneath the nuclei and centrosomes. When the nucleus divided, the MiDAS also split in two and followed the movement of the daughter nuclei. At that time, the distal ends of astral microtubules reached mainly the MiDAS regions of the ventral cortex. An inhibitor of microtubules induced disappearance of MiDASes, leading to aborted cytokinesis, suggesting that astral microtubules are required for the formation and maintenance of MiDASes. Fluorescence recovery after photobleaching experiments revealed that the MiDAS was highly dynamic and comprised small actin-containing dot-like structures. Interference reflection microscopy and assays blowing away the cell bodies by jet streaming showed that MiDASes were major attachment sites of dividing cells. Thus, the MiDASes are strong candidates for scaffolds for substratum-dependent cytokinesis, serving to transmit mechanical force to the substratum. Submitted by: Go Itoh [yumura@yamaguchi-u.ac.jp] -------------------------------------------------------------------------------- Dynacortin facilitates polarization of chemotaxing cells. Kabacoff C., Xiong Y., Musib R., Reichl E.M., Kim J., Iglesias P.A., Robinson D.N. BMC Biol. 2007; 5:53. Background Cell shape changes during cytokinesis and chemotaxis require regulation of the actin cytoskeletal network. Dynacortin, an actin cross-linking protein, localizes to the cell cortex and contributes to cortical resistance, thereby helping to define the cell shape changes of cytokinesis. Dynacortin also becomes highly enriched in cortical protrusions, which are sites of new actin assembly. Results We studied the effect of dynacortin on cell motility during chemotaxis and on actin dynamics in vivo and in vitro. Dynacortin enriches with the actin, particularly at the leading edge of chemotaxing cells. Cells devoid of dynacortin do not become as polarized as wild-type control cells but move with similar velocities as wild-type cells. In particular, they send out multiple pseudopods that radiate at a broader distribution of angles relative to the chemoattractant gradient. Wild-type cells typically only send out one pseudopod at a time that does not diverge much from 0° on average relative to the gradient. Though dynacortin-deficient cells show normal bulk (whole-cell) actin assembly upon chemoattractant stimulation, dynacortin can promote actin assembly in vitro. By fluorescence spectroscopy, co-sedimentation and transmission electron microscopy, dynacortin acts as an actin scaffolder in which it assembles actin monomers into polymers with a stoichiometry of 1 Dyn2:1 actin under salt conditions that disfavor polymer assembly. Conclusion Dynacortin contributes to cell polarization during chemotaxis. By cross-linking and possibly stabilizing actin polymers, dynacortin also contributes to cortical viscoelasticity, which may be critical for establishing cell polarity. Though not essential for directional sensing or motility, dynacortin is required to establish cell polarity, the third core feature of chemotaxis. Submitted by: Doug Robinson [dnr@jhmi.edu] -------------------------------------------------------------------------------- Developmental morphology and chemotactic responses are dependent on Galpha subunit specificity in Dictyostelium Jeffrey A. Hadwiger Developmental Biology (in press) Dictyostelium discoideum expresses multiple Galpha subunits but only a single Gbeta and Ggamma subunit suggesting that the specific response to an external signal depends largely on Galpha subunit function or G protein-independent signaling from the receptor. To test the contribution of Galpha subunit functional specificity, the chimeric Galpha subunits, Galpha2/4 and Galpha5/4, were created and analyzed along with wild-type subunits for the ability to substitute for the Galpha4 subunit in mediating responses from folate receptors. The Galpha2/4 subunit, but not the Galpha2 or Galpha5/4 subunits, partly rescued chemotaxis and cGMP accumulation in folate-stimulated galpha4(-) cells. Expression of the Galpha5/4 or Galpha5 subunits resulted in an inhibition of galpha4(-) and wild-type cell movement and a reduced aggregate size in developing wild-type and galpha5(-) cells suggesting these subunits mediate similar responses. Only the Galpha4 subunit was capable of correcting developmental morphology in galpha4(-) multicellular aggregates suggesting that the chimeric Galpha2/4 or Galpha5/4 subunits were insufficient to provide the Galpha4 function necessary for proper development. These results indicate that Dictyostelium Galpha subunit specificity is not limited to receptor coupling and that Galpha subunit sequences outside of the carboxyl terminus are important for cell movement and developmental processes. Submitted by: Jeff Hadwiger [jeff.hadwiger@okstate.edu] ============================================================== [End dictyNews, volume 29, number 15]