The zinc-finger transcription factor Hindsight regulates ovulation competency of Drosophila follicles
- University of Connecticut, United States
Figures
Figure 1
An illustration of Drosophila ovulation process.
The female reproductive system, consisting of two ovaries, oviduct, uterus, seminal receptacle, and a pair of spermathecae and parovaria, was depicted in the cartoon. Two representative ovarioles …
Figure 2 with 1 supplement
Hindsight expression in stage-14 egg chambers.
(A–C) Hnt expression (red in A-C and white in A’-C’) in A/P-Hnt (A), high-Hnt (B), and low-Hnt (C) egg chambers. FC1 expression (FC1 Gal4 driving UAS-eGFP, FC1 > GFP) is shown in green (A–C) and …
Figure 2—figure supplement 1
Egg chambers with different patterns of Hindsight expression.
(A–B’) Hnt (red in A-B and white in A’-B’) is expressed in posterior tip follicle cells (arrowheads) in youngest stage-14 egg chambers, which have faint FC1 (A) but no FC2 (B) expression (green). (C–…
Figure 3 with 2 supplements
Hindsight expression in stage-14 follicle cells is essential for normal ovulation.
(A–D) Hnt expression (green) in control (A) and hntRNAi (B–C) stage-14C egg chambers with FC1. FC1 expression (FC1 > RFP) is shown in red. Inserts are high magnification of Hnt expression in squared …
Figure 3—figure supplement 1
Hnt expression in stage-14A and stage-14B egg chambers.
(A–H) Hnt expression (green in A-C and E-G) in stage-14A (A–D) and stage-14B (E–H) egg chambers with FC1 driving RFP (control, A and E) or hntRNAi (B–C and F–G). Quantification of Hnt expression is …
Figure 3—figure supplement 2
hnt depletion in stage-14 follicle cells does not affect oogenesis in virgin females.
(A–B) Quantification of stage-14 egg chambers in virgin females before egg laying experiment. (A) is using FC1 Gal4, while (B) is using FC2-Gal4.
Figure 4 with 1 supplement
Hindsight is required for OA-induced follicle rupture.
(A–D) Representative images show control (A) and hntRNAi (B–C) egg chambers with FC1 after three-hour culture with 20 μM OA. Quantification of OA-induced follicle rupture is shown in (D). (E–H) …
Figure 4—figure supplement 1
hnt mutant mature follicles are defective for OA-induced follicle rupture.
(A–C) Representative images show hntpeb/+ (A), hntpeb/hntXE81 (B), and hntpeb/hntEH704a (C) mature follicles after three-hour culture with OA. Mature follicles were isolated according to FC2 > RFP …
Figure 5
Hindsight regulates Mmp2 activity in stage-14 follicle cells.
(A–C) Representative images show OA-induced Mmp2 activity (green) in control (A) and hntRNAi (B–C) egg chambers with FC1 after three-hour culture with OA. Egg chambers with posterior Mmp2 activity …
Figure 6 with 2 supplements
Hindsight regulates Mmp2 expression in stage-14 follicle cells.
(A–H) Response of egg chambers isolated according to FC1 (A–D) or FC2 (E–H) to ionomycin-induced rupture in three hours. (A–C and E–G) Representative images show control (A and E) and hntRNAi (B–C …
Figure 6—figure supplement 1
Measurement of intracellular Ca22+in follicle cells after ionomycin or octopamine stimulation.
(A–D) Intracellular Ca2+ depicted by GCaMP5G (green in A-C’) increase in response to ionomycin stimulation in both control (A–A’) and hntRNAi (B–C’) egg chambers with FC2 Gal4, although hntRNAi egg …
Figure 6—figure supplement 2
hnt depletion disrupts Mmp2 expression in posterior follicle cells.
Mmp2::GFP (green in A-C and white in A’-C’) is highly expressed in posterior follicle cells of control stage-14 egg chambers (A–A’), but is weakly detected in posterior follicle cells of hntRNAi egg …
Figure 7
Hnt regulates Oamb expression in stage-14 follicle cells.
(A–B) Quantification of Oamb mRNA levels in hntRNAi egg chambers with FC1 (A) or FC2 (B) Gal4. (C–D) Quantification of egg chambers in response to OA-induced follicle rupture. hntRNAi and/or Oamb …
Figure 8 with 3 supplements
Human RREB-1 can replace Hindsight’s role in regulating follicle’s competency to ovulation.
(A) The quantification of egg-laying capacity of females with FC1 driving hntRNAi and/or RREB-1 overexpression. (B–F) The quantification of OA-induced follicle rupture (B) in egg chambers with hntRNA…
Figure 8—figure supplement 1
Rescue of ovulation defect in hntRNAi females with hnt overexpression.
(A–D) Quantification of OA-induced follicle rupture (A, C) and the egg-laying number (B, D) from females with FC1 (A–B) and FC2 (C–D) Gal4 driving hntRNAi and/or hntEP55 overexpression. (E–L) Hnt …
Figure 8—figure supplement 2
Human RREB-1::GFP does not interfere with Hnt expression in control or hntRNAi egg chambers.
Hnt expression is shown in yellow (A–F) and white (A’–F’), and RREB1::GFP expression is shown in green (D–F) and white (D”–F”). (A–C) Hnt expression is detected in control follicle cells (A) and is …
Figure 8—figure supplement 3
Overexpression of GFP is not sufficient to rescue rupture defect of hntRNAi follicles with FC1 Gal4.
Quantification of OA-induced follicle rupture is shown.
***p<0.001.
Figure 9
A schematic cartoon summarizes the role of Hindsight in stage-14 follicle cells.
Hnt expression is shown in red with different intensity indicating different expression level. Mmp2 expression is shown in green and Oamb expression is shown in blue. OA stands for octopamine.
Videos
Video 1
Signal of GCaMP5G driven by FC2 in Control follicles with ionomycin stimulation (FC2 > GCaMP5G).
https://doi.org/10.7554/eLife.29887.015
Video 2
Signal of GCaMP5G driven by FC2 in hntRNAi1 follicles with ionomycin stimulation (FC2 > GCaMP5G/hntRNAi1).
https://doi.org/10.7554/eLife.29887.016
Video 3
Signal of GCaMP5G driven by FC2 in hntRNAi2 follicles with ionomycin stimulation (FC2 > GCaMP5G/hntRNAi2).
https://doi.org/10.7554/eLife.29887.017
Video 4
Signal of GCaMP5G driven by FC2 in Control follicles with octopamine stimulation (FC2 > GCaMP5G).
https://doi.org/10.7554/eLife.29887.018
Video 5
Signal of GCaMP5G driven by FC2 in hntRNAi1 follicles with octopamine stimulation (FC2 > GCaMP5G/hntRNAi1).
https://doi.org/10.7554/eLife.29887.019
Video 6
Signal of GCaMP5G driven by FC2 in hntRNAi2 follicles with octopamine stimulation (FC2 > GCaMP5G/hntRNAi2).
https://doi.org/10.7554/eLife.29887.020Additional files
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Supplementary file 1
The egg laying, egg distribution within the reproductive tract, and egg-laying time of females with various genotypes.
- https://doi.org/10.7554/eLife.29887.027
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Transparent reporting form
- https://doi.org/10.7554/eLife.29887.028
