Rho1 activity is necessary for fold and joint formation.

<p>(A-F) Distal region of 3 hrs APF leg discs showing the <i>ap</i> domain that encompasses the fold between the fourth and fifth tarsal segments (GFP, green). Right panels show a detail of the <i>ap</i> domain marked by green dotted lines and the t4-t5 fold (arrows). In A-F Phal is in red and separate channel in A’-F’ and Dcp1 is in blue in A-F and separate channel in A”-F”. Dlg is in blue in E and separate channel in E”. The activity of the <i>ap-Gal4</i> driver was restricted to 24 hrs before dissection using the <i>tubulin-Gal80</i>^<i>ts</i> system, except in A and F that was continuously active (see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1007584#sec012" target="_blank">Materials and methods</a>). (A) <i>ap>GFP</i> control disc. Note the t4-t5 epithelial fold within the <i>ap</i> domain. (B, C) <i>ap>Rho1</i>^<i>N19</i>. B and C correspond to opposite sides of the same leg disc. The t4-t5 fold is lost in B, while the epithelial integrity is severely altered in C. (D, E) <i>ap>Rho1</i>^<i>N19</i>, <i>miRHG</i>. The t4-t5 fold is lost while the epithelial integrity is not compromised as visualized by Dlg staining. (F) <i>ap>miRHG</i>. The t4-t5 fold is formed as in the control. (G-J) Adult legs of <i>ap>GFP</i> (G), <i>ap>Rho1</i>^<i>N19</i> (H), <i>ap>Rho1</i>^<i>N19</i>, <i>miRHG</i> (I) and <i>ap>miRHG</i> (J). The activity of the <i>ap-Gal4</i> driver was restricted to 48 hrs with the <i>tubulin-Gal80</i>^<i>ts</i> system except in G and J that was continuously active (see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1007584#sec012" target="_blank">Materials and methods</a>). In H and I the t4-t5 joint is lost, despite some indentations in the cuticle could still be seen. Note that in <i>ap>miRHG</i> legs (J) the fourth and fifth tarsi are slightly shorter and wider compared to the control (G). Arrows indicate formation of the joint between t4 and t5 while asterisks indicate its loss.