An EHBP-1-SID-3-DYN-1 axis promotes membranous tubule fission during endocytic recycling

Autoři: Jinghu Gao aff001;  Linyue Zhao aff001;  Qian Luo aff001;  Shuyao Liu aff001;  Ziyang Lin aff001;  Peixiang Wang aff001;  Xin Fu aff001;  Juan Chen aff001;  Hongjie Zhang aff002;  Long Lin aff001;  Anbing Shi aff001
Působiště autorů: Department of Biochemistry and Molecular Biology, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China aff001;  Faculty of Health Sciences, University of Macau, Avenida da Universidade, Taipa, Macau SAR, China aff002;  Institute for Brain Research, Huazhong University of Science and Technology, Wuhan, Hubei, China aff003;  Key Laboratory of Neurological Disease of National Education Ministry, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China aff004
Vyšlo v časopise: An EHBP-1-SID-3-DYN-1 axis promotes membranous tubule fission during endocytic recycling. PLoS Genet 16(5): e32767. doi:10.1371/journal.pgen.1008763
Kategorie: Research Article
doi: 10.1371/journal.pgen.1008763


The ACK family tyrosine kinase SID-3 is involved in the endocytic uptake of double-stranded RNA. Here we identified SID-3 as a previously unappreciated recycling regulator in the Caenorhabditis elegans intestine. The RAB-10 effector EHBP-1 is required for the endosomal localization of SID-3. Accordingly, animals with loss of SID-3 phenocopied the recycling defects observed in ehbp-1 and rab-10 single mutants. Moreover, we detected sequential protein interactions between EHBP-1, SID-3, NCK-1, and DYN-1. In the absence of SID-3, DYN-1 failed to localize at tubular recycling endosomes, and membrane tubules breaking away from endosomes were mostly absent, suggesting that SID-3 acts synergistically with the downstream DYN-1 to promote endosomal tubule fission. In agreement with these observations, overexpression of DYN-1 significantly increased recycling transport in SID-3-deficient cells. Finally, we noticed that loss of RAB-10 or EHBP-1 compromised feeding RNAi efficiency in multiple tissues, implicating basolateral recycling in the transport of RNA silencing signals. Taken together, our study demonstrated that in C. elegans intestinal epithelia, SID-3 acts downstream of EHBP-1 to direct fission of recycling endosomal tubules in concert with NCK-1 and DYN-1.

Klíčová slova:

Caenorhabditis elegans – Cell membranes – Endosomes – Fluorescence imaging – Gastrointestinal tract – RNA interference – Tyrosine kinases – Vacuoles


1. Maxfield FR, McGraw TE. Endocytic recycling. Nature reviews. 2004;5(2):121–32.

2. Grant BD, Donaldson JG. Pathways and mechanisms of endocytic recycling. Nature reviews. 2009;10(9):597–608. doi: 10.1038/nrm2755 19696797

3. Folsch H, Mattila PE, Weisz OA. Taking the scenic route: biosynthetic traffic to the plasma membrane in polarized epithelial cells. Traffic (Copenhagen, Denmark). 2009;10(8):972–81. Epub 2009/05/21.

4. Brown PS, Wang E, Aroeti B, Chapin SJ, Mostov KE, Dunn KW. Definition of distinct compartments in polarized Madin-Darby canine kidney (MDCK) cells for membrane-volume sorting, polarized sorting and apical recycling. Traffic (Copenhagen, Denmark). 2000;1(2):124–40. Epub 2001/02/24.

5. Wang P, Liu H, Wang Y, Liu O, Zhang J, Gleason A, et al. RAB-10 Promotes EHBP-1 Bridging of Filamentous Actin and Tubular Recycling Endosomes. PLoS genetics. 2016;12(6):e1006093. Epub 2016/06/09. doi: 10.1371/journal.pgen.1006093 27272733

6. Delevoye C, Heiligenstein X, Ripoll L, Gilles-Marsens F, Dennis MK, Linares RA, et al. BLOC-1 Brings Together the Actin and Microtubule Cytoskeletons to Generate Recycling Endosomes. Curr Biol. 2016;26(1):1–13. Epub 2016/01/05. doi: 10.1016/j.cub.2015.11.020 26725201

7. Puthenveedu MA, Lauffer B, Temkin P, Vistein R, Carlton P, Thorn K, et al. Sequence-dependent sorting of recycling proteins by actin-stabilized endosomal microdomains. Cell. 2010;143(5):761–73. Epub 2010/11/30. doi: 10.1016/j.cell.2010.10.003 21111236

8. Chen S, Li L, Li J, Liu B, Zhu X, Zheng L, et al. SEC-10 and RAB-10 coordinate basolateral recycling of clathrin-independent cargo through endosomal tubules in Caenorhabditis elegans. Proceedings of the National Academy of Sciences of the United States of America. 2014;111(43):15432–7. Epub 2014/10/11. doi: 10.1073/pnas.1408327111 25301900

9. Cullen PJ, Steinberg F. To degrade or not to degrade: mechanisms and significance of endocytic recycling. Nature reviews Molecular cell biology. 2018.

10. Chen CC, Schweinsberg PJ, Vashist S, Mareiniss DP, Lambie EJ, Grant BD. RAB-10 is required for endocytic recycling in the Caenorhabditis elegans intestine. Molecular biology of the cell. 2006;17(3):1286–97. doi: 10.1091/mbc.E05-08-0787 16394106

11. Pant S, Sharma M, Patel K, Caplan S, Carr CM, Grant BD. AMPH-1/Amphiphysin/Bin1 functions with RME-1/Ehd1 in endocytic recycling. Nat Cell Biol. 2009;11(12):1399–410. doi: 10.1038/ncb1986 19915558

12. Shi A, Chen CC, Banerjee R, Glodowski D, Audhya A, Rongo C, et al. EHBP-1 functions with RAB-10 during endocytic recycling in Caenorhabditis elegans. Molecular biology of the cell. 2010;21(16):2930–43. Epub 2010/06/25. doi: 10.1091/mbc.E10-02-0149 20573983

13. Guilherme A, Soriano NA, Bose S, Holik J, Bose A, Pomerleau DP, et al. EHD2 and the novel EH domain binding protein EHBP1 couple endocytosis to the actin cytoskeleton. The Journal of biological chemistry. 2004;279(11):10593–605. doi: 10.1074/jbc.M307702200 14676205

14. Guilherme A, Soriano NA, Furcinitti PS, Czech MP. Role of EHD1 and EHBP1 in perinuclear sorting and insulin-regulated GLUT4 recycling in 3T3-L1 adipocytes. The Journal of biological chemistry. 2004;279(38):40062–75. doi: 10.1074/jbc.M401918200 15247266

15. Grant B, Zhang Y, Paupard MC, Lin SX, Hall DH, Hirsh D. Evidence that RME-1, a conserved C. elegans EH-domain protein, functions in endocytic recycling. Nat Cell Biol. 2001;3(6):573–9. doi: 10.1038/35078549 11389442

16. Manser E, Leung T, Salihuddin H, Tan L, Lim L. A non-receptor tyrosine kinase that inhibits the GTPase activity of p21cdc42. Nature. 1993;363(6427):364–7. Epub 1993/05/27. doi: 10.1038/363364a0 8497321

17. Prieto-Echague V, Miller WT. Regulation of ack-family nonreceptor tyrosine kinases. J Signal Transduct. 2011;2011:742372. Epub 2011/06/04. doi: 10.1155/2011/742372 21637378

18. Teo M, Tan L, Lim L, Manser E. The tyrosine kinase ACK1 associates with clathrin-coated vesicles through a binding motif shared by arrestin and other adaptors. The Journal of biological chemistry. 2001;276(21):18392–8. Epub 2001/03/30. doi: 10.1074/jbc.M008795200 11278436

19. Shen H, Ferguson SM, Dephoure N, Park R, Yang Y, Volpicelli-Daley L, et al. Constitutive activated Cdc42-associated kinase (Ack) phosphorylation at arrested endocytic clathrin-coated pits of cells that lack dynamin. Molecular biology of the cell. 2011;22(4):493–502. Epub 2010/12/21. doi: 10.1091/mbc.E10-07-0637 21169560

20. Yeow-Fong L, Lim L, Manser E. SNX9 as an adaptor for linking synaptojanin-1 to the Cdc42 effector ACK1. FEBS letters. 2005;579(22):5040–8. Epub 2005/09/03. doi: 10.1016/j.febslet.2005.07.093 16137687

21. Yokoyama N, Lougheed J, Miller WT. Phosphorylation of WASP by the Cdc42-associated kinase ACK1: dual hydroxyamino acid specificity in a tyrosine kinase. The Journal of biological chemistry. 2005;280(51):42219–26. Epub 2005/11/01. doi: 10.1074/jbc.M506996200 16257963

22. Abdallah AM, Zhou X, Kim C, Shah KK, Hogden C, Schoenherr JA, et al. Activated Cdc42 kinase regulates Dock localization in male germ cells during Drosophila spermatogenesis. Developmental biology. 2013;378(2):141–53. Epub 2013/04/09. doi: 10.1016/j.ydbio.2013.02.025 23562806

23. Meisenhelder J, Hunter T. The SH2/SH3 domain-containing protein Nck is recognized by certain anti-phospholipase C-gamma 1 monoclonal antibodies, and its phosphorylation on tyrosine is stimulated by platelet-derived growth factor and epidermal growth factor treatment. Molecular and cellular biology. 1992;12(12):5843–56. Epub 1992/12/01. doi: 10.1128/mcb.12.12.5843 1448108

24. Galisteo ML, Yang Y, Urena J, Schlessinger J. Activation of the nonreceptor protein tyrosine kinase Ack by multiple extracellular stimuli. Proceedings of the National Academy of Sciences of the United States of America. 2006;103(26):9796–801. Epub 2006/06/17. doi: 10.1073/pnas.0603714103 16777958

25. Wunderlich L, Farago A, Buday L. Characterization of interactions of Nck with Sos and dynamin. Cell Signal. 1999;11(1):25–9. Epub 1999/04/17. doi: 10.1016/s0898-6568(98)00027-8 10206341

26. Jose AM, Kim YA, Leal-Ekman S, Hunter CP. Conserved tyrosine kinase promotes the import of silencing RNA into Caenorhabditis elegans cells. Proceedings of the National Academy of Sciences of the United States of America. 2012;109(36):14520–5. Epub 2012/08/23. doi: 10.1073/pnas.1201153109 22912399

27. Glodowski DR, Chen CC, Schaefer H, Grant BD, Rongo C. RAB-10 regulates glutamate receptor recycling in a cholesterol-dependent endocytosis pathway. Molecular biology of the cell. 2007;18(11):4387–96. doi: 10.1091/mbc.E07-05-0486 17761527

28. Shi A, Liu O, Koenig S, Banerjee R, Chen CC, Eimer S, et al. RAB-10-GTPase-mediated regulation of endosomal phosphatidylinositol-4,5-bisphosphate. Proceedings of the National Academy of Sciences of the United States of America. 2012;109(35):E2306–15. Epub 2012/08/08. doi: 10.1073/pnas.1205278109 22869721

29. Chen D, Yang C, Liu S, Hang W, Wang X, Chen J, et al. SAC-1 ensures epithelial endocytic recycling by restricting ARF-6 activity. The Journal of cell biology. 2018;217(6):2121–39. Epub 2018/03/23. doi: 10.1083/jcb.201711065 29563216

30. Gong T, Yan Y, Zhang J, Liu S, Liu H, Gao J, et al. PTRN-1/CAMSAP promotes CYK-1/formin-dependent actin polymerization during endocytic recycling. The EMBO journal. 2018;37(9). Epub 2018/03/24.

31. Shi A, Pant S, Balklava Z, Chen CC, Figueroa V, Grant BD. A novel requirement for C. elegans Alix/ALX-1 in RME-1-mediated membrane transport. Curr Biol. 2007;17(22):1913–24. doi: 10.1016/j.cub.2007.10.045 17997305

32. Shi A, Sun L, Banerjee R, Tobin M, Zhang Y, Grant BD. Regulation of endosomal clathrin and retromer-mediated endosome to Golgi retrograde transport by the J-domain protein RME-8. The EMBO journal. 2009;28(21):3290–302. doi: 10.1038/emboj.2009.272 19763082

33. Liu H, Wang S, Hang W, Gao J, Zhang W, Cheng Z, et al. LET-413/Erbin acts as a RAB-5 effector to promote RAB-10 activation during endocytic recycling. The Journal of cell biology. 2018;217(1):299–314. Epub 2017/10/29. doi: 10.1083/jcb.201705136 29079669

34. Neukomm LJ, Zeng S, Frei AP, Huegli PA, Hengartner MO. Small GTPase CDC-42 promotes apoptotic cell corpse clearance in response to PAT-2 and CED-1 in C. elegans. Cell death and differentiation. 2014;21(6):845–53. Epub 2014/03/19. doi: 10.1038/cdd.2014.23 24632947

35. Kamath RS, Ahringer J. Genome-wide RNAi screening in Caenorhabditis elegans. Methods (San Diego, Calif. 2003;30(4):313–21.

36. Lin SX, Grant B, Hirsh D, Maxfield FR. Rme-1 regulates the distribution and function of the endocytic recycling compartment in mammalian cells. Nat Cell Biol. 2001;3(6):567–72. doi: 10.1038/35078543 11389441

37. Daumke O, Lundmark R, Vallis Y, Martens S, Butler PJ, McMahon HT. Architectural and mechanistic insights into an EHD ATPase involved in membrane remodelling. Nature. 2007;449(7164):923–7. doi: 10.1038/nature06173 17914359

38. Rolls MM, Hall DH, Victor M, Stelzer EH, Rapoport TA. Targeting of rough endoplasmic reticulum membrane proteins and ribosomes in invertebrate neurons. Molecular biology of the cell. 2002;13(5):1778–91. Epub 2002/05/15. doi: 10.1091/mbc.01-10-0514 12006669

39. Casanova JE, Wang X, Kumar R, Bhartur SG, Navarre J, Woodrum JE, et al. Association of Rab25 and Rab11a with the apical recycling system of polarized Madin-Darby canine kidney cells. Molecular biology of the cell. 1999;10(1):47–61. Epub 1999/01/08. doi: 10.1091/mbc.10.1.47 9880326

40. Brown FD, Rozelle AL, Yin HL, Balla T, Donaldson JG. Phosphatidylinositol 4,5-bisphosphate and Arf6-regulated membrane traffic. The Journal of cell biology. 2001;154(5):1007–17. doi: 10.1083/jcb.200103107 11535619

41. Liu O, Grant BD. Basolateral Endocytic Recycling Requires RAB-10 and AMPH-1 Mediated Recruitment of RAB-5 GAP TBC-2 to Endosomes. PLoS genetics. 2015;11(9):e1005514. Epub 2015/09/24. doi: 10.1371/journal.pgen.1005514 26393361

42. Mahajan NP, Whang YE, Mohler JL, Earp HS. Activated tyrosine kinase Ack1 promotes prostate tumorigenesis: role of Ack1 in polyubiquitination of tumor suppressor Wwox. Cancer Res. 2005;65(22):10514–23. Epub 2005/11/17. doi: 10.1158/0008-5472.CAN-05-1127 16288044

43. Antonny B, Burd C, De Camilli P, Chen E, Daumke O, Faelber K, et al. Membrane fission by dynamin: what we know and what we need to know. The EMBO journal. 2016;35(21):2270–84. Epub 2016/11/04. doi: 10.15252/embj.201694613 27670760

44. van der Bliek AM, Redelmeier TE, Damke H, Tisdale EJ, Meyerowitz EM, Schmid SL. Mutations in human dynamin block an intermediate stage in coated vesicle formation. The Journal of cell biology. 1993;122(3):553–63. Epub 1993/08/01. doi: 10.1083/jcb.122.3.553 8101525

45. Gibbings DJ, Ciaudo C, Erhardt M, Voinnet O. Multivesicular bodies associate with components of miRNA effector complexes and modulate miRNA activity. Nat Cell Biol. 2009;11(9):1143–9. Epub 2009/08/18. doi: 10.1038/ncb1929 19684575

46. Lee YS, Pressman S, Andress AP, Kim K, White JL, Cassidy JJ, et al. Silencing by small RNAs is linked to endosomal trafficking. Nat Cell Biol. 2009;11(9):1150–6. Epub 2009/08/18. doi: 10.1038/ncb1930 19684574

47. Hinas A, Wright AJ, Hunter CP. SID-5 is an endosome-associated protein required for efficient systemic RNAi in C. elegans. Curr Biol. 2012;22(20):1938–43. Epub 2012/09/18. doi: 10.1016/j.cub.2012.08.020 22981770

48. Sato T, Mushiake S, Kato Y, Sato K, Sato M, Takeda N, et al. The Rab8 GTPase regulates apical protein localization in intestinal cells. Nature. 2007;448(7151):366–9. Epub 2007/06/29. doi: 10.1038/nature05929 17597763

49. Feinberg EH, Hunter CP. Transport of dsRNA into cells by the transmembrane protein SID-1. Science (New York, NY. 2003;301(5639):1545–7. Epub 2003/09/13.

50. Shih JD, Hunter CP. SID-1 is a dsRNA-selective dsRNA-gated channel. RNA. 2011;17(6):1057–65. Epub 2011/04/09. doi: 10.1261/rna.2596511 21474576

51. McEwan DL, Weisman AS, Hunter CP. Uptake of extracellular double-stranded RNA by SID-2. Molecular cell. 2012;47(5):746–54. Epub 2012/08/21. doi: 10.1016/j.molcel.2012.07.014 22902558

52. Winston WM, Sutherlin M, Wright AJ, Feinberg EH, Hunter CP. Caenorhabditis elegans SID-2 is required for environmental RNA interference. Proceedings of the National Academy of Sciences of the United States of America. 2007;104(25):10565–70. Epub 2007/06/15. doi: 10.1073/pnas.0611282104 17563372

53. Jose AM, Smith JJ, Hunter CP. Export of RNA silencing from C. elegans tissues does not require the RNA channel SID-1. Proceedings of the National Academy of Sciences of the United States of America. 2009;106(7):2283–8. Epub 2009/01/27. doi: 10.1073/pnas.0809760106 19168628

54. Rocheleau CE. RNA interference: Systemic RNAi SIDes with endosomes. Curr Biol. 2012;22(20):R873–5. Epub 2012/10/27. doi: 10.1016/j.cub.2012.08.039 23098595

55. Wang E, Hunter CP. SID-1 Functions in Multiple Roles To Support Parental RNAi in Caenorhabditis elegans. Genetics. 2017;207(2):547–57. Epub 2017/07/29. doi: 10.1534/genetics.117.300067 28751423

56. Nguyen TA, Smith BRC, Tate MD, Belz GT, Barrios MH, Elgass KD, et al. SIDT2 Transports Extracellular dsRNA into the Cytoplasm for Innate Immune Recognition. Immunity. 2017;47(3):498–509 e6. Epub 2017/09/17. doi: 10.1016/j.immuni.2017.08.007 28916264

57. Nakajo A, Yoshimura S, Togawa H, Kunii M, Iwano T, Izumi A, et al. EHBP1L1 coordinates Rab8 and Bin1 to regulate apical-directed transport in polarized epithelial cells. The Journal of cell biology. 2016;212(3):297–306. Epub 2016/02/03. doi: 10.1083/jcb.201508086 26833786

58. Whangbo JS, Weisman AS, Chae J, Hunter CP. SID-1 Domains Important for dsRNA Import in Caenorhabditis elegans. G3 (Bethesda). 2017;7(12):3887–99. Epub 2017/10/14.

59. Timmons L, Fire A. Specific interference by ingested dsRNA. Nature. 1998;395(6705):854. doi: 10.1038/27579 9804418

60. Chen Y, Wang Y, Zhang J, Deng Y, Jiang L, Song E, et al. Rab10 and myosin-Va mediate insulin-stimulated GLUT4 storage vesicle translocation in adipocytes. The Journal of cell biology. 2012;198(4):545–60. Epub 2012/08/22. doi: 10.1083/jcb.201111091 22908308

61. Duleh SN, Welch MD. WASH and the Arp2/3 complex regulate endosome shape and trafficking. Cytoskeleton (Hoboken). 2010;67(3):193–206. Epub 2010/02/23.

62. Gutkowska M, Swiezewska E. Structure, regulation and cellular functions of Rab geranylgeranyl transferase and its cellular partner Rab Escort Protein. Mol Membr Biol. 2012;29(7):243–56. Epub 2012/06/15. doi: 10.3109/09687688.2012.693211 22694141

63. Praitis V, Casey E, Collar D, Austin J. Creation of low-copy integrated transgenic lines in Caenorhabditis elegans. Genetics. 2001;157(3):1217–26. 11238406

64. Schindelin J, Arganda-Carreras I, Frise E, Kaynig V, Longair M, Pietzsch T, et al. Fiji: an open-source platform for biological-image analysis. Nat Methods. 2012;9(7):676–82. Epub 2012/06/30. doi: 10.1038/nmeth.2019 22743772

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