Simultaneous order buildup during myofibril initiation and subsarcomeric order during sarcomere maturation.

<p>(A) Flight muscle at 32-h APF stained with rhodamine-Alexa488. Overlaid longitudinal orange lines (from one fiber end to the other) indicate positions at which molecular actin order was determined. Scale bar represents 50 μm. (B) Molecular actin order angle longitudinal profile for flight muscles at 32-h APF. Each color line corresponds to an average profile obtained on an individual pupa (the orange one corresponds to the pupa shown in panel A; the others are displayed in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.2004718#pbio.2004718.s001" target="_blank">S1 Fig</a>). Bars indicate the mean values of 6 different pupae and error bars the SDs. Note that there is no significant difference between the center and the ends of the muscle fiber. Values are relative to the averaged value per pupa. (C–E) Grey maps showing the actin intensity of an averaged sarcomere with the Z-disc (green arrowhead) shown in the middle and the M-lines (red arrowhead) at 48-h (panel C), 72-h (panel D), and 90-h APF (panel E) as shown in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.2004718#pbio.2004718.g002" target="_blank">Fig 2G–2I</a>. Red sticks represent the local actin orientation angle ρ. Stick angles have been amplified 50 times relative to horizontal axis to visualize the small differences. Scale bar represents 1 μm. (F) Molecular actin order angle profiles along the averaged sarcomere length at 48-h (light grey), 72-h (dark grey), and 90-h APF (black). Values are relative to the minimum of the 90-h APF profile. Red arrows show M-line positions and green ones the Z-disc position. Note the relatively higher actin order at the developing Z-disc of 48-h APF sarcomeres. **<i>P</i> < 0.01; (Wilcoxon signed-rank test; see <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.2004718#pbio.2004718.s003" target="_blank">S1 Data</a> for primary data). APF, After Puparium Formation.