Inscuteable-mediated orientation of the mitotic spindle requires Gαi.

<p>a, current model of Inscuteable function. Insc serves as a link between the apical PAR complex and the spindle-orienting Pins-Gái complex. b, Ed-Insc (green) transfected S2 cells randomly orient the mitotic spindle (red) with respect to the region that is enriched in Ed. Spindle alignment is measured by drawing a vector from the center of the crescent (arrow) to the center of the mitotic spindle and then along the axis (dashes). c, Expression of Gái (blue) with Ed-Insc (green)is able to confer moderate spindle orienting activity. d, Cumulative percentage plot of spindle angles measured in the S2 Echinoid induced-polarity assay for Ed-Insc and Ed-Insc+Gái compared to previously-published data <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0029611#pone.0029611-Johnston1" target="_blank">[17]</a>. In these plots, the cumulative percentage of cells with a spindle angle below a particular value (x-axis) is shown. High spindle orienting activity corresponds to a deflection to lower spindle angles whereas no activity is a line across the diagonal. e, Ed-Insc expression in S2 cells is sufficient to robustly recruit endogenous Pins from the cytoplasm to the region of Ed enrichment. f, Ed alone is unable to polarize endogenous Pins. e, g, Ed-Insc induces colocalization of endogenous Pins with overexpressed Gai. h, Ed-Pins is able to recruit endogenous Dlg. i, Co-expression of Gαi with Ed-Pins results in robust recruitment of endogenous Mud. j,k Ed-GFP is unable to recruit endogenous Dlg or Mud to the induced-polarity cortical domains. l, Ed-Insc is able to recruit Dlg to the cortex, similar to cells expressing Ed-GFP-Pins. m, Ed-Insc is not able to recruit Mud (red) to the Ed-crescent, even in the presence of Gái. Scale bars for all panels represent 5 µm.