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Fig. 1. aak-2 larvae are defective in establishing germline quiescence
under compromised insulin-like signalling. (A) Open arrowheads
delineate the distal extremities of the gonad of daf-2(e1370) dauer
larvae. (B) An integrated lag-2p::GFP transgene
(qIs56) is strongly expressed in the DTCs throughout the dauer stage.
A lag-2p::CFP transgene (arEx645) containing the full 6.2 kb
promoter (Chen and Greenwald,
2004) was also strongly expressed in the DTCs of dauer larvae
(data not shown). We similarly found strong GFP expression in the DTCs of
qEx308 dauer larvae (data not shown), which contain a functional
lag-2p::LAG-2::GFP lag-2(q411) rescuing construct (S.
Crittenden and J. Kimble, personal communication). (C) Extruded
daf-2(e1370) dauer gonad stained with DAPI (blue) and anti-GLP-1
antibodies (red). (D) Enlarged gonad of daf-2(e1370);
aak-2(rr48) dauer larvae. (E) aak-2 is required to
progressively suppress germline proliferation during L2d/dauer formation. Grey
zone indicates the period during which >90% of the larvae undergo the dauer
moult. The dauer moult was delayed by 
1 and 3 hours in
aak-2(ok524) and aak-2(rr48) mutants, respectively. Error
bars represent standard deviation, and the sample size was 
25 for each
point. (F) In both daf-2(e1370) and daf-7(e1372)
larvae, germ cells arrest in mitotic interphase during dauer (arrowhead), as
shown in a DAPI-stained, extruded daf-7(e1372) dauer gonad.
(G) In daf-2(e1370) glp-1(e2141) dauer larvae, germ cell
nuclei arrest in late stage meiotic prophase I, the nuclear size and
chromosome morphology being consistent with the pachytene stage (inset).
(H) Germ cells often (26/30 animals) progress further through meiosis
in daf-2(e1370) glp-1(e2141); aak-2(rr48) dauer larvae, and
their chromosomal morphology closely resembles that of spermatids (arrow).
Similar defects were observed in daf-2(e1370) glp-1(e2141);
aak-2(ok524) (19/40 animals) and in daf-2(e1370)
glp-1(e2141); par-4(it57) (25/30 animals) dauer larvae.
(I-L) Extruded daf-2(e1370) glp-1(e2141); aak-2(rr48)
dauer gonad. Asterisks indicate germ cells undergoing spermatogenesis. Cells
were stained with (I) DAPI (blue); (J) anti-P-granule (green), a germ cell
marker that is lost during spermatogenesis
(Gruidl et al., 1996); (K)
anti-membranous organelle (1CB4; red), a sperm marker
(Arduengo et al., 1998). A
merged image is shown in L. Images in B,C,F-H are single focal planes;
condensed z-stacks are shown in I-L. Scale bars: 50 µm in A,B,D;
10 µm in C,F-H,I-L.
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