Fig 1.
Drosophila follicle cells remain in the ovary following ovulation and form a corpus luteum.
(A) A schematic diagram of female reproductive system. The red squared area is the calyx where post-ovulatory follicle cells are located. (B) R47A04-Gal4 driving UAS-GFP expression (R47A04>GFP) is specifically in follicle cells of S14 egg chambers (mature follicles). (C) Follicle cells from post-ovulatory follicles (green; outlined) remain in the ovary after the egg enters the oviduct and form a corpus luteum (CL). (D) Hnt accumulates in S14 follicle cells after decreasing in S13. (E) Hnt expression continues in CL cells. (F and F’) CL cells continue to express Arm, an adherens junction component (Red in F, white in F’) suggesting that they maintain their apical-basal polarity. G) Ovaries of virgin females before egg laying. No pigmentation is found in the ovary. (H, I) Ovaries of mated females after egg laying. Yellow pigmentation (arrows) is found in the CL. A single CL is outlined. (J-L) Shade (Shd) is expressed in CL cells (dash-line outlined in J and J’) and S14 follicle cells (K, K’, and L). Shd expression co-localized with a mitochondria marker (yellow in J and K), but not with an endoplasmic reticulum marker (L).
Fig 2.
Posterior follicle cells are removed to release the oocyte and initiate corpus luteum formation.
(A-D) Schematic (left panels) and real (right panels) expression patterns of Gal4 drivers in follicle cells of mature egg chambers and corpus luteum. Anterior follicle cells (B and D) reside at the anterior part of the corpus luteum, the middle follicle cells (C) reside at the middle portion of the corpus luteum, while the posterior follicle cells (D) are degraded in the corpus luteum. (E) An egg partially extruded into one lateral oviduct. (F-F’) DAPI stained calyx region showing the presence of two stage 14 trimming follicles (outlined) with no follicle cells at the posterior ends. The one with most posterior follicle cells trimmed was half-way in the lateral oviduct. The posterior end of follicle cells (brighter and larger nuclei pointed with an arrowhead) was marked by dashed lines, and the oviduct cells (faint and smaller nuclei) was marked by an arrow. F’ is the higher magnification of the squared region in F. (G) Histogram showing the number of follicles undergone trimming in mated (optimal ovulation) and unmated (no ovulation) females. Student’s T-test is used (*** P<0.001).
Fig 3.
Mmp2 is required for ovulation and CL formation.
(A) In situ zymography showing one preselected follicle with high gelatinase activity at its posterior end in the entire ovary pair. (B-D) Correlation of follicle cell trimming and gelatinase activity (green in B-D). During early (B), middle (C-C’), and late (D-D’) ovulation, gelatinase activity is covering all the egg chamber area lost follicle cells. Posterior leading edge of the follicle cell layer was marked by dashed lines. Smaller nuclei (white in C’ and D’) are oviduct cells. (E-G) Egg laying (E), mature follicles in ovary (F), and the average ovulation time (G) of females of actGal4 or actGal4 driven Mmp1RNAi, Mmp2RNAi, or Timp expression in adult. (H-I) Follicle cell trimming in actGal4 control (H) and actGal4/Mmp2RNAi (I) ovaries. Mature egg chambers with posterior follicle cell trimmed were outlined with solid line and the posterior edge of the residual follicle cells were marked with dashed line. Follicle cell nuclei are elucidated by DAPI signal. (J) A quantification of trimming follicles in ovaries. (K-L) More corpora lutea (arrowheads) are found in actGal4 control ovaries (K) than those in actGal4/Mmp2RNAi ovaries (L) 6 hours after mating. One ovariole with three mature egg chambers was outlined in (K). (M) Number of eggs laid in 6 hours after mating. T-test is used (*** P<0.001, ** P<0.01, * P<0.05).
Table 1.
The effect of Mmp activity on egg laying, egg distribution in reproductive tract, and egg laying time.
Table 2.
The effect of Mmp activity for follicle cell trimming.
Fig 4.
Mmp2 functions in follicle cells to control ovulation.
(A-C) Mmp2 expression in mature egg chambers indicated by Mmp2::GFP fusion protein. Mmp2 is highly expressed in both anterior (B) and posterior (C) follicle cells. Hnt (Red) is used to mark follicle cells. (D) Mmp2::GFP is expressed in anterior and posterior corpus luteum (outlined with dashed line). (E) Mmp2::GFP is expressed in the posterior edge of the trimming follicle cells when the egg is half way in the oviduct. The egg chamber is outlined by solid line, the posterior edge of follicle cells is marked by small dashed lines, and the corpus luteum is outlined by big dashed line. Smaller nuclei without Hnt expression are from oviduct epithelial cells. (F-G) The egg laying (F) and ovulation time (G) of females with 47A04-Gal4 or 47A04-Gal4 driven Mmp2RNAi or Timp expression in mature follicle cells. T-test is used (*** P<0.001, *P<0.05). (H-I) Ectopic Mmp2 in mature follicle cells is sufficient to cause egg release from ovary into abdominal cavity. (J) DAPI staining to indicate the follicle cell trimming of egg chambers released from ovary ectopic Mmp2 expression in (I). (K) Quantification of egg chamber trimming. Egg chambers already released from ovaries were collected from abdominal cavity of females with 47A04 driven Mmp1 or Mmp2 expression. 149 and 144 released egg chambers were collected from Mmp1 and Mmp2 flies respectively.