遗传 ›› 2026, Vol. 48 ›› Issue (6): 601-613.doi: 10.16288/j.yczz.26-004
吉长皓(
), 王暖, 王欣阳, 张美珠, 孙锦()
收稿日期:2026-01-14
修回日期:2026-02-16
出版日期:2026-04-24
发布日期:2026-04-24
通讯作者:
孙锦，副教授，研究方向：果蝇肠道干细胞调控。E-mail: sunjin@sdfmu.edu.cn作者简介:吉长皓，硕士研究生，专业方向：动物学。E-mail: 1051482846@qq.com
基金资助:
Changhao Ji(), Nuan Wang, Xinyang Wang, Meizhu Zhang, Jin Sun()
Received:2026-01-14
Revised:2026-02-16
Published:2026-04-24
Online:2026-04-24
Supported by:摘要：
蜕皮激素信号通路(ecdysone signaling)通过促进干细胞增殖与分化，在成年黑腹果蝇(Drosophila melanogaster)肠道稳态维持中具有重要作用。然而，其下游靶基因broad(br)在该过程中的功能尚不明确。本研究证明，br基因对肠道干细胞(intestinal stem cells，ISCs)的维持及肠道损伤修复过程不可或缺。克隆分析结果显示，br同时调控ISCs的增殖与分化。敲低br表达会严重阻碍干细胞向肠上皮细胞(enterocytes，ECs)的分化，而过表达br则会促进其分化。进一步分析发现，敲低br表达会导致ISCs中Jak/STAT信号通路活性降低，而上调Jak/STAT或EGFR信号通路的活性都可以显著挽救br敲低导致的表型。这些结果表明，在成年果蝇中，br可能通过Jak/STAT和EGFR信号通路调控ISCs的增殖与分化。本研究揭示了br基因在果蝇成虫ISCs中的作用，并初步探讨了其机制。
吉长皓, 王暖, 王欣阳, 张美珠, 孙锦. broad在果蝇成虫肠道干细胞中的作用及其机制研究[J]. 遗传, 2026, 48(6): 601-613.
Changhao Ji, Nuan Wang, Xinyang Wang, Meizhu Zhang, Jin Sun. Role of broad in intestinal stem cells of adult Drosophila[J]. Hereditas(Beijing), 2026, 48(6): 601-613.
表1
引物序列"
| 引物名称 | 序列(5′→3′) | 用途 |
|---|---|---|
| br-F | ATCCCATATTCAGCCGCTAGCATGGACGACACACAGCACTT | br-OE果蝇构建 |
| br-R1 | TAGATCTTCTCGAGTGGATCCGCCCTATGAAGAGGAGTGGT | |
| br-R2 | TAGATCTTCTCGAGTGGATCCTAGTTGCTCGTGGTGGCAA | |
| br-R3 | TAGATCTTCTCGAGTGGATCCTCCTATTGGATGCCCGGTGG | |
| br-R4 | TAGATCTTCTCGAGTGGATCCGCCTTGCTTATTGGTCTAGT | |
| nub-F1 | AAGATCTGGAATTCAGCTAGTTCGGTGAAATCGGCCACAG | nub-HA果蝇构建 |
| nub-R1 | GATCTCGGATCCTTAGCTAGCGTGCATCATATAGGAGGACTCG | |
| nub-F2 | ATGCACGCTAGCTAAGGATCCGAGATCAAGGATCGTCTATGGAC | |
| nub-R2 | CCAATTGGTCTAGACGGATCTTCCCTCGGTCGAATTCCGT | |
| HA-F | CTAGCTACCCATACGATGTTCCAGATTACGCTTAAG | |
| HA-R | GATCCTTAAGCGTAATCTGGAACATCGTATGGGTAG | |
| nub-sgRNA-F | TTCGAAGCGCATCAATCCCTCCC | |
| nub-sgRNA-R | AAACGGGAGGGATTGATGCGCTT | |
| dome-F | GGCGGCGACTTTAATCTGAG | 实时荧光定量PCR |
| dome-R | GGTGTTGTTCAGGATTCGGAT | |
| stat92E-F | CAACAATCCACCCACAGTCGAG | |
| stat92E-R | GTATTGCGCGTAACGAAC |
图1
br基因在成虫果蝇肠道中的表达 A：果蝇中肠br表达与EEs细胞定位免疫荧光图。GFP指示br的表达，pros(Prospero)指示肠内分泌细胞(EEs)。B：不同性别及DSS处理下果蝇中肠br表达的免疫荧光图。从左到右分别为雌性(♀)果蝇中肠、雄性(♂)果蝇中肠、未经DSS处理(Control)及DSS处理3天的雌性果蝇中肠，pH3(phosphorylation of H10S10)指示处于有丝分裂中期的细胞。基因型均为br-Gal4>UAS-GFP；比例尺均为20 μm。"
图2
敲低br导致ISCs减少 A：基因型分别为esgts (Control)、esgts>br KD1和esgts>br KD2的果蝇中肠免疫荧光图。GFP指示ISCs与EBs，pros(Prospero)指示EEs。B：每条肠道单位面积内ISCs+EBs以及EEs的定量结果。C：热激诱导克隆3周后基因型分别为Flp-out (Control)、Flp-out>br KD1和Flp-out>br KD2的果蝇中肠免疫荧光图。GFP指示诱导产生的克隆。D：热激后1周、2周、3周的果蝇中肠R4~R5区域克隆数量的统计。比例尺均为20 μm；数据均为平均值±SD；***P<0.001，差异极显著。"
图3
敲低br影响ISCs增殖 A：热激7天后基因型分别为Flp-out(Control)、Flp-out>br KD1和Flp-out>br KD2的果蝇中肠免疫荧光图。GFP指示诱导产生的克隆。B：果蝇中肠单克隆内细胞数量统计。C：DSS处理3天后基因型分别为esgts (Control)和esgts>br KD果蝇中肠免疫荧光图。pH3指示处于有丝分裂中期的细胞。D：果蝇中肠pH3阳性细胞的数量统计。E：基因型分别为esgts (Control)和esgts>br KD果蝇中肠的免疫荧光图。cDcp-1(cleaved Dcp-1)指示凋亡的细胞。比例尺均为20 μm；数据均为平均值±SD；***P<0.001，差异极显著。"
图4
br调控ISCs分化 A：热激7天后基因型分别为Flp-out (Control)和Flp-out>br KD的果蝇中肠免疫荧光图。Dl(Delta)抗体特异性标记ISCs，GFP指示诱导产生的克隆。B：ISCs细胞的数量统计。C：热激7天后基因型分别为Flp-out; nub-HA (Control)和Flp-out>br KD; nub-HA的果蝇中肠免疫荧光图。HA指示ECs，GFP指示诱导产生的克隆。D：ECs细胞的占比。E：热激7天后基因型分别为Flp-out(Control)和Flp-out>br KD的果蝇中肠免疫荧光图。pros指示EEs，GFP指示诱导产生的克隆。F：EEs细胞的数量统计。G：热激7天后基因型分别为Flp-out(Control)、Flp-out>UAS-brZ1、Flp-out>UAS-brZ2、Flp-out>UAS-brZ3和Flp-out>UAS-brZ4的果蝇中肠免疫荧光图。GFP指示诱导产生的克隆。比例尺均为20 μm；数据均为平均值± SD；***P<0.001，差异极显著。"
图5
br敲低降低Jak/STAT信号通路活性 A：dome和stat92E基因实时荧光定量PCR结果。B：热激7天后基因型分别为Flp-out; stat92E-lacZ (Control)和Flp-out>br KD; stat92E-lacZ的果蝇中肠免疫荧光图。lacZ指示stat92E蛋白的表达水平，GFP指示诱导产生的克隆。C：热激7天后基因型分别为Flp-out;10xSTAT-mCherry(Control)和Flp-out>br KD; 10xSTAT-mCherry的果蝇中肠免疫荧光图。mCherry指示Jak/STAT信号通路活性，GFP指示诱导产生的克隆。比例尺均为10 μm。"
图6
br通过Jak/STAT和EGFR信号通路调控ISCs增殖与分化 A：热激7天后基因型分别为Flp-out(Control)、Flp-out; hopT42、Flp-out; rasv12、Flp-out>br KD、Flp-out>br KD; hopT42和Flp-out>br KD; rasv12的果蝇中肠免疫荧光图；B：热激7天后基因型分别为Flp-out (Control)、Flp-out>UAS-brz1、Flp-out>UAS-brz2、Flp-out>stat92E KD、Flp-out>UAS-brz1; stat92E KD和Flp-out>UAS-brz2; stat92E KD的果蝇中肠免疫荧光图。GFP标记诱导产生的克隆，pros标记EEs；比例尺均为20 μm。"
| [1] |
Sharpless NE, DePinho RA. How stem cells age and why this makes us grow old. Nat Rev Mol Cell Biol, 2007, 8(9): 703-713.
pmid: 17717515 |
| [2] |
Visvader JE, Lindeman GJ. Cancer stem cells in solid tumours: accumulating evidence and unresolved questions. Nat Rev Cancer, 2008, 8(10): 755-768.
pmid: 18784658 |
| [3] | Trivanović D. Adult stem cells in aging. J Pers Med, 2022, 12(5): 795. |
| [4] |
Hsu YC, Li LS, Fuchs E. Emerging interactions between skin stem cells and their niches. Nat Med, 2014, 20(8): 847-856.
pmid: 25100530 |
| [5] |
Gadhave DG, Sugandhi VV, Jha SK, Nangare SN, Gupta G, Singh SK, Dua K, Cho H, Hansbro PM, Paudel KR. Neurodegenerative disorders: mechanisms of degeneration and therapeutic approaches with their clinical relevance. Ageing Res Rev, 2024, 99: 102357.
pmid: 38830548 |
| [6] |
Shao B, Yang W, Cao Q. Landscape and predictions of inflammatory bowel disease in China: China will enter the Compounding Prevalence stage around 2030. Front Public Health, 2022, 10: 1032679.
pmid: 36388296 |
| [7] |
Zheng R, Zhang S, Zeng H, Wang S, Sun K, Chen R, Li L, Wei W, He J. Cancer incidence and mortality in China, 2016. J Natl Cancer Cent, 2022, 2(1): 1-9.
pmid: 39035212 |
| [8] |
Micchelli CA, Perrimon N. Evidence that stem cells reside in the adult Drosophila midgut epithelium. Nature, 2006, 439(7075): 475-479.
pmid: 16340959 |
| [9] |
Ohlstein B, Spradling A. The adult Drosophila posterior midgut is maintained by pluripotent stem cells. Nature, 2006, 439(7075): 470-474.
pmid: 16340960 |
| [10] |
Ohlstein B, Spradling A. Multipotent Drosophila intestinal stem cells specify daughter cell fates by differential notch signaling. Science, 2007, 315(5814): 988-992.
pmid: 17303754 |
| [11] |
Zeng X, Hou SX. Enteroendocrine cells are generated from stem cells through a distinct progenitor in the adult Drosophila posterior midgut. Development, 2015, 142(4): 644-653.
pmid: 25670791 |
| [12] |
Chen J, Xu N, Wang C, Huang P, Huang H, Jin Z, Yu Z, Cai T, Jiao R, Xi R. Transient scute activation via a self-stimulatory loop directs enteroendocrine cell pair specification from self-renewing intestinal stem cells. Nat Cell Biol, 2018, 20(2):152-161.
pmid: 29335529 |
| [13] |
Miguel-Aliaga I, Jasper H, Lemaitre B. Anatomy and physiology of the digestive tract of Drosophila melanogaster. Genetics, 2018, 210(2):357-396.
pmid: 30287514 |
| [14] |
Guo X, Lv J, Xi R. The specification and function of enteroendocrine cells in Drosophila and mammals: a comparative review. FEBS J, 2022, 289(16): 4773-4796.
pmid: 34115929 |
| [15] |
Tang R, Qin P, Liu X, Wu S, Yao R, Cai G, Gao J, Wu Y, Guo Z. Intravital imaging strategy FlyVAB reveals the dependence of Drosophila enteroblast differentiation on the local physiology. Commun Biol, 2021, 4(1): 1223.
pmid: 34697396 |
| [16] |
Amcheslavsky A, Jiang J, Ip YT. Tissue damage-induced intestinal stem cell division in Drosophila. Cell Stem Cell, 2009, 4(1): 49-61.
pmid: 19128792 |
| [17] |
Jiang H, Tian A, Jiang J. Intestinal stem cell response to injury: lessons from Drosophila. Cell Mol Life Sci, 2016, 73(17): 3337-3349.
pmid: 27137186 |
| [18] |
Johansson J, Naszai M, Hodder MC, Pickering KA, Miller BW, Ridgway RA, Yu Y, Peschard P, Brachmann S, Campbell AD, Cordero JB, Sansom OJ. RAL GTPases drive intestinal stem cell function and regeneration through internalization of WNT signalosomes. Cell Stem Cell, 2019, 24(4): 592-607.e7.
pmid: 30853556 |
| [19] |
Jiang H, Grenley MO, Bravo MJ, Blumhagen RZ, Edgar BA. EGFR/Ras/MAPK signaling mediates adult midgut epithelial homeostasis and regeneration in Drosophila. Cell Stem Cell, 2011, 8(1): 84-95.
pmid: 21167805 |
| [20] |
Wu Y, Yu L, Yu Y, Wu S, Yuan Q, Duan W, Cai S, Xiong B, Lin R, Guo Z. Drosophila and mouse intestinal stem cells are spatiotemporally specified by Notch suppression and Wnt activation. Sci Adv, 2025, 11(49): eady7272.
pmid: 41337595 |
| [21] |
Guo Z, Ohlstein B. Stem cell regulation. Bidirectional Notch signaling regulates Drosophila intestinal stem cell multipotency. Science, 2015, 350(6263): aab0988.
pmid: 26586765 |
| [22] |
Tian A, Benchabane H, Ahmed Y. Wingless/Wnt signaling in intestinal development, homeostasis, regeneration and tumorigenesis: a Drosophila perspective. J Dev Biol, 2018, 6(2): 8.
pmid: 29615557 |
| [23] |
Jiang H, Patel PH, Kohlmaier A, Grenley MO, McEwen DG, Edgar BA. Cytokine/Jak/Stat signaling mediates regeneration and homeostasis in the Drosophila midgut. Cell, 2009, 137(7): 1343-1355.
pmid: 19563763 |
| [24] |
Xu N, Wang SQ, Tan D, Gao YW, Lin G, Xi R. EGFR, Wingless and JAK/STAT signaling cooperatively maintain Drosophila intestinal stem cells. Dev Biol, 2011, 354(1): 31-43.
pmid: 21440535 |
| [25] |
Okamoto N, Watanabe A. Interorgan communication through peripherally derived peptide hormones in Drosophila. Fly (Austin), 2022, 16(1): 152-176.
pmid: 35499154 |
| [26] |
Tian L, Guo E, Wang S, Liu S, Jiang RJ, Cao Y, Ling E, Li S. Developmental regulation of glycolysis by 20-hydroxyecdysone and juvenile hormone in fat body tissues of the silkworm, Bombyx mori. J Mol Cell Biol, 2010, 2(5): 255-263.
pmid: 20729248 |
| [27] |
Ahmed SMH, Maldera JA, Krunic D, Paiva-Silva GO, Pénalva C, Teleman AA, Edgar BA. Fitness trade-offs incurred by ovary-to-gut steroid signalling in Drosophila. Nature, 2020, 584(7821): 415-419.
pmid: 32641829 |
| [28] |
Zipper L, Jassmann D, Burgmer S, Görlich B, Reiff T. Ecdysone steroid hormone remote controls intestinal stem cell fate decisions via the PPARγ-homolog Eip75B in Drosophila. eLife, 2020, 9: e55795.
pmid: 32773037 |
| [29] |
DiBello PR, Withers DA, Bayer CA, Fristrom JW, Guild GM. The Drosophila broad-complex encodes a family of related proteins containing zinc fingers. Genetics, 1991, 129(2): 385-397.
pmid: 1743483 |
| [30] |
Zeng X, Hou SX. Broad relays hormone signals to regulate stem cell differentiation in Drosophila midgut during metamorphosis. Development, 2012, 139(21): 3917-3925.
pmid: 23048182 |
| [31] |
Zhang S, Wu S, Yao R, Wei X, Ohlstein B, Guo Z. Eclosion muscles secrete ecdysteroids to initiate asymmetric intestinal stem cell division in Drosophila. Dev Cell, 2024, 59(1): 125-140.e12.
pmid: 38096823 |
| [32] |
Wang F, Qiao HH, Xu RG, Sun J, Zhu R, Mao D, Ni JQ. pNP transgenic RNAi system manual in Drosophila. Bio Protoc, 2019, 9(3): e3158.
pmid: 33654966 |
| [33] |
Ren X, Yang Z, Xu J, Sun J, Mao C, Hu Y, Yang SJ, Qiao HH, Wang X, Hu Q, Deng P, Liu LP, Ji JY, Li JB, Ni JQ. Enhanced specificity and efficiency of the CRISPR/Cas 9 system with optimized sgRNA parameters in Drosophila. Cell Rep, 2014, 9(3): 1151-1162.
pmid: 25437567 |
| [34] | Hu JL, Xi RW. A method for damage-induced regeneration of the intestinal epithelium in Drosophila. Bio-101, 2019, e1010303. |
| [35] |
Lin G, Xu N, Xi R. Paracrine Wingless signalling controls self-renewal of Drosophila intestinal stem cells. Nature, 2008, 455(7216): 1119-1123.
pmid: 18806781 |
| [36] |
Luo H, Rose P, Barber D, Hanratty WP, Lee S, Roberts TM, D'Andrea AD, Dearolf CR. Mutation in the Jak kinase JH2 domain hyperactivates Drosophila and mammalian Jak-Stat pathways. Mol Cell Biol, 1997, 17(3): 1562-1571.
pmid: 9032284 |
| [37] |
Reiff T, Jacobson J, Cognigni P, Antonello Z, Ballesta E, Tan KJ, Yew JY, Dominguez M, Miguel-Aliaga I. Endocrine remodelling of the adult intestine sustains reproduction in Drosophila. eLife, 2015, 4: e06930.
pmid: 26216039 |
| [38] |
Viragova S, Li D, Klein OD. Activation of fetal-like molecular programs during regeneration in the intestine and beyond. Cell Stem Cell, 2024, 31(7): 949-960.
pmid: 38971147 |
| [39] |
Zhang P, Edgar BA. Insect gut regeneration. Cold Spring Harb Perspect Biol, 2022, 14(2): a040915.
pmid: 34312250 |
| [40] |
Spokony RF, Restifo LL. Broad Complex isoforms have unique distributions during central nervous system metamorphosis in Drosophila melanogaster. J Comp Neurol, 2009, 517(1): 15-36.
pmid: 19711379 |
| [41] |
Uyehara CM, McKay DJ. Direct and widespread role for the nuclear receptor EcR in mediating the response to ecdysone in Drosophila. Proc Natl Acad Sci USA, 2019, 116(20): 9893-9902.
pmid: 31019084 |
| [1] | 杨剑, 石国娟, 彭昂惠, 徐清波, 王睿琪, 薛雷, 喻昕阳, 孙艺昊. Tip60-FOXO调节果蝇JNK信号通路介导的细胞凋亡【已撤稿】[J]. 遗传, 2024, 46(6): 490-501. |
| [2] | 韦湘怡, 胡冬春, 高祖鹏, 冯从经. JAK/STAT信号通路及其对昆虫免疫的调控[J]. 遗传, 2023, 45(3): 229-236. |
| [3] | 范志露, 杨荔艳, 张娜, 冯丹, 郭静, 常晨, 苑青, 蔡岩, 张钰, 魏文强, 王明荣, 郝佳洁. NEFL通过激活EGFR/AKT/S6信号通路促进食管鳞癌细胞的侵袭迁移[J]. 遗传, 2022, 44(4): 322-334. |
| [4] | 孙帅, 邓宇亮. 肺癌循环肿瘤细胞的单细胞EGFR基因突变检测[J]. 遗传, 2015, 37(12): 1251-1257. |
| [5] | 张柿平，薛雷. 黑腹果蝇细胞谱系分析方法进展[J]. 遗传, 2012, 34(7): 819-828. |
| [6] | 李海静，刘岩，郝海生，杜卫华，赵学明，王栋，秦彤，马友记，朱化彬. 表皮生长因子受体与肺脏发育的关系[J]. 遗传, 2012, 34(1): 27-32. |
| [7] | 张文燕，张菁，钱远槐，曾庆韬. 黑腹果蝇种组五种核型的报道[J]. 遗传, 2006, 28(5): 545-550. |
| [8] | 李红伟，连林生，赵春江，白丽华，邓学梅，吴常信，. 黑腹果蝇中一个Cy染色体上的缺失[J]. 遗传, 2006, 28(10): 1254-1259. |
| [9] | 王慧超，朱勇，夏庆友. 黑腹果蝇的性别控制[J]. 遗传, 2003, 25(1): 97-101. |
| [10] | 周玉萍，田长恩，周伯春，王正询. 黑腹果蝇杂交实验的教学改革[J]. 遗传, 2002, 24(3): 345-348. |
| [11] | 汪旭，合正基，刘素清. 昆明山海棠诱发果蝇生殖细胞非整倍体的研究[J]. 遗传, 1995, 17(5): 34-36. |
| [12] | 许珊，姚珧，沈红，吴学军，庚镇城，朱定良. 黑腹果蝇单胚培养技术的建立[J]. 遗传, 1994, 16(2): 25-28. |
| 阅读次数 | ||||||
|
全文 |
|
|||||
|
摘要 |
|
|||||
www.chinagene.cn
备案号：京ICP备09063187号-4
总访问:,今日访问:,当前在线:
