A-F’) Representative images of Drosophila adult midguts of esg ts ( A-A’), esg ts>yki ( B-B’), esg ts> lola ( C-C’), and esg ts>yki; lola (D-D’) immunostained with Arm and Pros (red).ISCs and EBs are marked with esg-GFP (green). Merged images are shown in A’, B’, C’, and D’ (green and red). ( E) Quantifications of the p-H3 + cell number in adult midguts with the indicated genotypes in A-D (n=6, 6, 7, 9). Note that co-expression of lola and yki does not suppress the increase in the p-H3 + cell number induced by yki overexpression (ns p>0.05). Scale bars: 30 μm. Data are shown as mean±SEM. ns p>0.05, **p<0.01, and ***p<0.001 by Student’s T-test. At least 10 midguts were dissected for each genotype. Confocal images were taken from the basal layer of the midgut where ISCs can be clearly visualized. Single layer image is shown. Ohlstein B, Spradling A. The adult Drosophila posterior midgut is maintained by pluripotent stem cells. Nature. 2006; 439:470–474. doi: 10.1038/nature04333. [ PubMed] [ CrossRef] [ Google Scholar] Antonello ZA, Reiff T, Ballesta-Illan E, Dominguez M. Robust intestinal homeostasis relies on cellular plasticity in enteroblasts mediated by miR-8-Escargot switch. The EMBO Journal. 2015; 34:2025–2041. doi: 10.15252/embj.201591517. [ PMC free article] [ PubMed] [ CrossRef] [ Google Scholar] Our present findings suggest that Lola regulates ISC proliferation and midgut homeostasis. Reduced lola expression in both ISCs and ECs promotes ISC proliferation, indicating that Lola functions both cell-autonomously and non-cell-autonomously. These precursors remain correctly differentiated when Lola is absent, suggesting that Lola only affects ISC proliferation, but not differentiation. Similarly, inactivation of Hippo signaling also causes enhanced ISC proliferation, indicating the possibility that Lola functions downstream of Hippo. Consistent to the characterized phenotype for Hippo signaling inactivation, downregulating lola expression in ECs activates EGFR and JAK-STAT signaling, and Lola regulates Hippo downstream bantam expression levels in ISCs. Taken together, these results suggest that Lola regulates ISC proliferation and homeostasis likely via mechanisms involving Hippo signaling.

Anders S, Pyl PT, Huber W. HTSeq--a Python framework to work with high-throughput sequencing data. Bioinformatics. 2015; 31:166–169. doi: 10.1093/bioinformatics/btu638. [ PMC free article] [ PubMed] [ CrossRef] [ Google Scholar] Probably earlier this year when [I found out] I was going to be working with Make Up For Ever and Vogue. Last year was a pretty difficult year for me. I completed my transition, I had surgery in the beginning of last year, and I had to deal with changing agencies and changing careers, so it was a pretty intense year. I faced quite a lot of rejection, and it made me doubt things. So it felt amazing to come out on top of it all in such a major way."

EGFR ligands and Upd1,2,3 are upregulated upon Lola-KD in ECs (Figure 2—figure supplement 2). These genes have been demonstrated to be regulated by the canonical Hippo/Yki pathway in ECs upon stress conditions. Given this, the authors should clarify if these genes' upregulation is Yki-dependent or not. In addition, the authors show that esg ts>LolaRNAi upregulates Bantam, which is a classical Yki target. So, what is the Yki's function in this context? Please explain.

Giniger E, Tietje K, Jan LY, Jan YN. Lola encodes a putative transcription factor required for axon growth and guidance in Drosophila. Development. 1994; 120:1385–1398. [ PubMed] [ Google Scholar]The role of canonical Hippo signaling in restricting intestinal stem cell (ISC) proliferation is well established, but whether and how the non-canonical Hippo signaling is involved in ISC basal proliferation are less well-defined.

Furukawa M, He YJ, Borchers C, Xiong Y. Targeting of protein ubiquitination by BTB-Cullin 3-Roc1 ubiquitin ligases. Nature Cell Biology. 2003; 5:1001–1007. doi: 10.1038/ncb1056. [ PubMed] [ CrossRef] [ Google Scholar] Children's animation. Join Lola and Charlie, a brother and sister, as they deal with everyday topics. Lola does not want a haircut. MoreFluorescent microscopy was performed on a Leica LAS SP8 confocal microscope; confocal images were obtained using the Leica AF Lite system. Confocal images from the basal layer of the posterior midguts (sub-region R5a, https://flygut.epfl.ch/overview) where ISCs can be clearly visualized were taken under 40×objective. Single layer image is shown. The quantification of p-H3 + cell per guts was undertaken by counting the p-H3 + cell numbers over the whole gut of indicated genotypes. The p-H3 + cell number quantification data were statistically present as average with the standard error of mean (SEM) and p-values of significance is calculated by Student’s T-test (tails=2, Two-sample unequal variance): * is p<0.05, ** is p<0.01, *** is p<0.001, ns is no significance with p>0.05. Pintard L, Willems A, Peter M. Cullin-based ubiquitin ligases: cul3-btb complexes join the family. The EMBO Journal. 2004; 23:1681–1687. doi: 10.1038/sj.emboj.7600186. [ PMC free article] [ PubMed] [ CrossRef] [ Google Scholar]