JCS019091 Supplementary Material

Files in this Data Supplement:

• Supplemental Figure S1 -

  Fig. S1. Drug-fed flies show mislocalized grk mRNA. (A) Adult female grk*RFP flies were fed for 24 hours ethanol mixed in with a yeast paste. The stage 9 egg chamber shows properly localized grk*RFP at the dorsal anterior corner. (B,C) grk*RFP flies were fed for 24 hours colchicine and colcemid mixed in with a yeast paste. (B) The stage 8 egg chamber shows mislocalized grk*RFP in the oocyte and an accumulation of fluorescence in the nurse cells. (C) The stage 9 egg chamber shows a misplaced oocyte nucleus with no surrounding grk*RFP.

• Supplemental Figure S2 -

  Fig. S2. FRAP was performed on grk mRNA during different stages of oogenesis. (A-C) Images of egg chambers at stage 6 (A), early 8 (B) and mid-late stage 9 (C) from flies expressing grk^HGFP that correspond to the FRAP movies found in supplementary material Movies 2-4. The black boxes cover the region that is bleached during the FRAP experiments. Panels (D-G) show images of grk*GFP at the dorsal anterior corner (D) pre-bleach, (E) post-bleach, (F) 2 µm up the plane of focus and (G) 2 µm down the plane of focus. A white box outlines the area bleached. OON, oocyte nucleus.

• Supplemental Figure S3 -

  Fig. S3. Mislocalized grk mRNA is photobleached. Image of grk*GFP;sqd¹ egg chamber with a black box covering the ventral-anterior area photobleached during FRAP experiments.

• Supplemental Figure S4 -

  Fig. S4. Recovery of posterior grk mRNA is altered in K10 and sqd¹ mutants FRAP experiments were performed on stage-6 and -7 egg chambers from K10; grk^HGFP and grk*GFP;sqd¹. The X-Y scatter plot shows the data point-spread of recovery from the different fly lines. Posterior-localized grk*GFP in K10 mutants shows a high degree of variability, particularly during stage 6, whereas the sqd¹ mutants show a general increase in recovery during both stages (P=0.004 for sqd¹ stage 7). A minimum of five FRAP experiments were performed for each data point. Error bars represent standard deviations.

• Movie 1 -

  Movie 1. A population of microtubules is resistant to drug treatment. Time-lapse movie of microtubules surrounding the oocyte nucleus of a stage-10 egg chamber from flies expressing tau-GFP. The egg chamber was treated with colchicine and colcemid at time 0′ and subsequently filmed for 40 minutes. There appears to be no obvious depolymerization or breakage of the microtubules.

• Movie 2 -

  Movie 2. There is high level of recovery of grk mRNA fluorescence during young stages of oogenesis. Time-lapse FRAP movie of a stage 6 egg chamber from a grk*GFP-expressing fly under the conditions used for quantification found in the Materials and Methods section. The movie corresponds to supplementary material Fig. S2A.

• Movie 3 -

  Movie 3. There is significant recovery of grk mRNA fluorescence during mid-stages of oogenesis. Time-lapse FRAP movie of an early stage 8 egg chamber from a grk*GFP-expressing fly. The movie corresponds to supplementary material Fig. S2B.

• Movie 4 -

  Movie 4. There is a decrease in recovery of grk mRNA fluorescence as the oocyte develops. Time-lapse FRAP movie of a mid-late stage 9 egg chamber from a grk*GFP-expressing fly. The movie corresponds to supplementary material Fig. S2C. As oogenesis progresses, the amount of fluorescence recovery decreases.

• Movie 5 -

  Movie 5. Mislocalized grk mRNA is dynamic. Time-lapse FRAP movie from the ventral-anterior corner of a grk*GFP;sqd¹ egg chamber showing significant fluorescence recovery of the mislocalized mRNA.