(A to E) Confocal section through the testis apex immunostained with [(A) and (B)] antibody to Vasa (germline, red), antibody to Fasciclin III (hub, green), antibody to Hts (1B1 fusome, green), and 4′,6′-diamidino-2-phenylindole (DAPI) (DNA, blue) or with [(D) and (E)] antibody to Vasa (red), antibody to ZFH-1 (CPCs and their immediate daughters, green), and DAPI (blue). The hub is outlined. (A) In wild-type testes, a rosette of GSCs (one indicated, arrow) surrounds the hub; Vasa-negative CPCs (one indicated, arrowhead) make minimal contact with the hub. (B) socs36E^PZ1647 testes have fewer GSCs (arrow); instead, somatic cells (arrowhead) surround the hub. (C) Bar graph showing number of GSCs per testis. Homozygous mutant socs36E^PZ1647 testes contain significantly fewer GSCs than do wild-type (red, 3.8 ± 0.2, n = 12 testes, versus yellow, 9.9 ± 0.4, n = 14 testes, P < 0.0001) or socs36E heterozygotes (green, 9.5 ± 0.4, n = 14 testes). Testes from socs36E^PZ1647 in trans to a deletion of socs36E (Df(2L)M36F-S5) phenocopy socs36E^PZ1647 (blue, 4.0 ± 0.2, n = 12 testes). Leaky expression from a socs36E cDNA transgene rescues the phenotype (gray, 8.9 ± 0.7 GSCs, n = 15 testes). Precise excision of the P-element restores GSC number to wild-type (white, 9.6 ± 0.6, n = 11 testes). (D) In wild-type testes, GSCs surround the hub; nuclei of ZFH-1–positive CPCs (arrowhead) are displaced from the hub. (E) In socs36E^PZ1647, CPCs (arrowhead) occupy positions at the hub that usually contain GSCs. (F and G) Ultrastructural analysis of the testis niche. Hub cells (red), GSCs (yellow), and a gonialblast (blue), identified by differences in morphology and staining density (29), are indicated. (F) In wild-type testes, GSCs make broad contact with the hub; CPCs extend thin projections (arrowheads) between GSCs. (G) In socs36E^PZ1647, CPCs broadly contact the hub, whereas GSCs make small areas of contact with the hub (rectangle). Scale bars, 10 μm.

(A and B) Confocal section through the testis apex immunostained with antibody to Vasa (red), antibody to βPS-integrin (green, and in insets), and DAPI (blue). (A) In wild-type testes, βPS-integrin is expressed strongly in the sheath (open arrowhead) and at moderate levels throughout the apex; within the niche, it is enriched at points of CPC-hub cell contact (arrowheads). (B) In socs36E^PZ1647 testes, expression of βPS-integrin at the CPC-hub interface (arrowheads) is increased relative to wild-type controls. (C) socs36E^PZ1647 GSC loss is partially rescued by introducing one copy of a talin loss-of-function allele (rhea^6-66) (red, 4.2 ± 0.2, n = 24 testes, versus gray, 7.7 ± 0.4, n = 18 testes, P < 0.0001). Testes from control flies (socs36E^PZ1647 or rhea^6-66 heterozygotes and socs36E^PZ1647; rhea^6-66 transheterozygotes) contain wild-type numbers of GSCs (yellow, 9.6 ± 0.3, n = 14 testes; green, 9.3 ± 0.2, n = 22 testes; and blue, 8.8 ± 0.3, n = 24 testes, respectively). CyO and MKRS denote balancer chromosomes containing wild-type alleles of socs36E and rhea. (D) Expression of ectopic βPS-integrin in CPCs and cyst cells significantly reduces the number of GSCs per testis as compared with UAS-GFP controls. Scale bars, 10 μm.

(A to F) Confocal sections through the testis apex. [(A) to (D)] Hubs denoted by asterisks. [(A) and (B)] socs36E^PZ1647 GSC clones (outlined, arrowhead) and their differentiating progeny (outlined, arrow) identified by the absence of green fluorescent protein (GFP) (green) and the presence of Vasa (red) at (A) 2 days and (B) 14 days after clonal induction (ACI). GSCs at the hub and their progeny are maintained. [(C) and (D)] socs36E^PZ1647 CPC clones (outlined, arrowhead) and their differentiating daughters (outlined, arrow) identified by the absence of GFP (green) and the presence of ZFH-1 (red) at (C) 2 days and (D) 14 days ACI. CPCs lacking socs36E accumulate around the hub, competing for niche occupancy and leading to a loss of GSCs over time. [(E) and (F)] Testes immunostained with antibody to Stat92E (green), antibody to Vasa (red), and DAPI (blue); GSCs (arrows), CPCs (arrowheads), and hubs (outlined) are indicated. [(E′) and (F′)] Stat92E channel. [(E) and (E′)] In wild-type testes, Stat92E is detectable at higher levels in GSCs than in CPCs or the hub. [(F) and (F′)] In socs36E^PZ1647 testes, Stat92E levels in hub cells and CPCs are increased relative to wild-type controls. Scale bars, 10 μm.