Fig. 2: LAT-1 modulates Notch signaling in the distal gonad.

a In the distal gonad, the Notch pathway controls cell proliferation and the switch between germ cell division and differentiation. It is initiated by the Notch ligand LAG-2 on the distal tip cell (DTC) binding to the Notch receptor GLP-1 present on germ cell membranes. This causes the Notch intracellular domain (NICD) to be cleaved off to translocate to the nucleus, initiating together with LAG-1 and LAG-3 the transcription of downstream effectors such as LST-1 and SYGL-1. In turn, these effectors repress expression of e.g., gld-1-3. b Expression of gld-1 occurs more distally in lat-1 mutant (bottom) than in wild-type (top) L4 + 8 h gonads. Asterisks: DTC, dashed lines: end of the progenitor zone. c Quantification of gld-1::GFP expression/germ cell row in lat-1 and wild-type gonads by measuring fluorescence intensity with increasing distance from the distal tip cell based on images from (b). Loss of lat-1 leads to a reduced repression of gld-1. Wild-type: n = 17, lat-1: n = 22 in four independent experiments. d, e Comparison of progenitor zones of Notch pathway component single mutants (lag-2(q420), glp-1(bn18)) and respective double mutants with lat-1(ok1465). Analyses are based on DAPI-/PH3-stained dissected gonads to visualize all and specifically M phase nuclei (for images see Supplementary Fig. 2a). The progenitor zone sizes in the double mutants differ from those in lat-1 single mutants, but are similar to the respective Notch component single mutants (d). Overall, no differences in PH3-positive germ cell number were observed in single compared to double mutants (e). Replicate values: wild-type: 44, lat-1: 38, lag-2: 42, lat-1; lag-2: 36, glp-1: 41, lat-1; glp-1: 46 gonads in 6 independent experiments. f Notch activation was visualized using the GLP-1 NICD::V5 reporter glp-1(q1000[glp-1::4xV5]. In lat-1 germline nuclei of L4 + 8 h-old hermaphrodites, less NICD is present than in the wild-type (red arrowheads) which is ameliorated by the presence of the LNT. The lag-2(q420) mutant (positive control) shows severely reduced NICD in cell nuclei. asterisks: DTC. g Quantification of the NICD fraction located in germ cell nuclei based on images in (f) confirms the reduced activity of Notch in lat-1 mutants. Replicate values (independent experiments): wild-type: 33 (5), lat-1: 33 (3), lag-2: 21 (4), LNT: 16 (3). h DIC images showing anus morphology defects in L1 nematodes, which occurs both in lat-1 and lag-2 mutants. i Quantification of the images from (h). Wild-type: 127, lag-2: 122, lat-1: 167, lat-1; lag-2: 86 in three independent experiments. j lat-1 mutant nematodes exhibit a similar delayed reversal upon exposure to octanol as lag-2 mutants. Replicate values (independent experiments): wild-type: 75 (5), osm-11: 45 (3), lat-1: 75 (6), lag-2: 45 (3), lat-1, lag-2: 45 (3). Graph raw data are provided in the Source Data. Graph details and statistics are: (c): Mean gld-1::GFP expression/germ cell row ± SEM (replicates and exact p values in the Source Data). Two-sided unpaired t-test without multiple comparison correction. (d), (e), (g), (i), (j): Box plots with median (center), interquartal range, 5th (lower whisker) and 95th (upper whisker) percentiles. One-way ANOVA with Bonferroni post-hoc test. (i): Mean ± SEM. One-way ANOVA with Bonferroni post-hoc test.