Marcksb plays a key role in the secretory pathway of zebrafish Bmp2b

English version

Autoři: Ding Ye ^aff001; Xiaosi Wang ^aff001; Changyong Wei ^aff001; Mudan He ^aff001; Houpeng Wang ^aff001; Yanwu Wang ^aff001; Zuoyan Zhu ^aff001; Yonghua Sun ^aff001
Působiště autorů: State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Innovative Academy for Seed Design, Chinese Academy of Sciences, Wuhan, China ^aff001; College of Advanced Agricultural Sciences, University of Chinese Academy of Sciences, Beijing, China ^aff002; School of Basic Medical Sciences, Wuhan University, Wuhan, China ^aff003
Vyšlo v časopise: Marcksb plays a key role in the secretory pathway of zebrafish Bmp2b. PLoS Genet 15(9): e32767. doi:10.1371/journal.pgen.1008306
Kategorie: Research Article
doi: https://doi.org/10.1371/journal.pgen.1008306

Souhrn

During vertebrate early embryogenesis, the ventral development is directed by the ventral-to-dorsal activity gradient of the bone morphogenetic protein (BMP) signaling. As secreted ligands, the extracellular traffic of BMP has been extensively studied. However, it remains poorly understood that how BMP ligands are secreted from BMP-producing cells. In this work, we show the dominant role of Marcksb controlling the secretory process of Bmp2b via interaction with Hsp70 in vivo. We firstly carefully characterized the role of Marcksb in promoting BMP signaling during dorsoventral axis formation through knockdown approach. We then showed that Marcksb cell autonomously regulates the trafficking of Bmp2b from producing cell to the extracellular space and both the total and the extracellular Bmp2b was decreased in Marcksb-deficient embryos. However, neither the zygotic mutant of marcksb (Zmarcksb) nor the maternal zygotic mutant of marcksb (MZmarcksb) showed any defects of dorsalization. In contrast, the MZmarcksb embryos even showed increased BMP signaling activity as measured by expression of BMP targets, phosphorylated Smad1/5/9 levels and imaging of Bmp2b, suggesting that a phenomenon of “genetic over-compensation” arose. Finally, we revealed that the over-compensation effects of BMP signaling in MZmarcksb was achieved through a sequential up-regulation of MARCKS-family members Marcksa, Marcksl1a and Marcksl1b, and MARCKS-interacting protein Hsp70.3. We concluded that the Marcksb modulates BMP signaling through regulating the secretory pathway of Bmp2b.

Klíčová slova:

Embryos – Immunoblotting – Phosphorylation – Secretion – Secretory pathway – Zebrafish – BMP signaling – Guide RNA

Zdroje

1. Bier E, De Robertis EM. BMP gradients: A paradigm for morphogen-mediated developmental patterning. Science. 2015;348(6242):aaa5838. doi: 10.1126/science.aaa5838 26113727

2. Zinski J, Bu Y, Wang X, Dou W, Umulis D, Mullins MC. Systems biology derived source-sink mechanism of BMP gradient formation. eLife. 2017;6:e22199. doi: 10.7554/eLife.22199 28826472

3. Wei CY, Wang HP, Zhu ZY, Sun YH. Transcriptional factors smad1 and smad9 act redundantly to mediate zebrafish ventral specification downstream of smad5. The Journal of biological chemistry. 2014;289(10):6604–18. doi: 10.1074/jbc.M114.549758 24488494; PubMed Central PMCID: PMC3945323.

4. Little SC, Mullins MC. Bone morphogenetic protein heterodimers assemble heteromeric type I receptor complexes to pattern the dorsoventral axis. Nature cell biology. 2009;11(5):637–43. Epub 2009/04/21. doi: 10.1038/ncb1870 19377468; PubMed Central PMCID: PMC2757091.

5. Schwank G, Dalessi S, Yang SF, Yagi R, de Lachapelle AM, Affolter M, et al. Formation of the long range Dpp morphogen gradient. PLoS biology. 2011;9(7):e1001111. doi: 10.1371/journal.pbio.100111 21814489; PubMed Central PMCID: PMC3144185.

6. Belenkaya TY, Han C, Yan D, Opoka RJ, Khodoun M, Liu H, et al. Drosophila Dpp morphogen movement is independent of dynamin-mediated endocytosis but regulated by the glypican members of heparan sulfate proteoglycans. Cell. 2004;119(2):231–44. doi: 10.1016/j.cell.2004.09.031 15479640.

7. Ramel MC, Hill CS. The ventral to dorsal BMP activity gradient in the early zebrafish embryo is determined by graded expression of BMP ligands. Developmental biology. 2013;378(2):170–82. doi: 10.1016/j.ydbio.2013.03.003 23499658; PubMed Central PMCID: PMC3899928.

8. Takada S, Fujimori S, Shinozuka T, Takada R, Mii Y. Differences in the secretion and transport of Wnt proteins. Journal of biochemistry. 2017;161(1):1–7. doi: 10.1093/jb/mvw071 28053142.

9. Burke R, Nellen D, Bellotto M, Hafen E, Senti KA, Dickson BJ, et al. Dispatched, a novel sterol-sensing domain protein dedicated to the release of cholesterol-modified hedgehog from signaling cells. Cell. 1999;99(7):803–15. doi: 10.1016/s0092-8674(00)81677-3 10619433.

10. Callejo A, Bilioni A, Mollica E, Gorfinkiel N, Andres G, Ibanez C, et al. Dispatched mediates Hedgehog basolateral release to form the long-range morphogenetic gradient in the Drosophila wing disk epithelium. Proceedings of the National Academy of Sciences of the United States of America. 2011;108(31):12591–8. doi: 10.1073/pnas.1106881108 21690386; PubMed Central PMCID: PMC3150953.

11. Zehe C, Engling A, Wegehingel S, Schafer T, Nickel W. Cell-surface heparan sulfate proteoglycans are essential components of the unconventional export machinery of FGF-2. Proceedings of the National Academy of Sciences of the United States of America. 2006;103(42):15479–84. doi: 10.1073/pnas.0605997103 17030799; PubMed Central PMCID: PMC1622848.

12. Dahal GR, Pradhan SJ, Bates EA. Inwardly rectifying potassium channels influence Drosophila wing morphogenesis by regulating Dpp release. Development. 2017;144(15):2771–83. Epub 2017/07/08. doi: 10.1242/dev.146647 28684627; PubMed Central PMCID: PMC5560040.

13. Nakaoka T, Kojima N, Ogita T, Tsuji S. Characterization of the phosphatidylserine-binding region of rat MARCKS (myristoylated, alanine-rich protein kinase C substrate). Its regulation through phosphorylation of serine 152. The Journal of biological chemistry. 1995;270(20):12147–51. doi: 10.1074/jbc.270.20.12147 7744864.

14. McIlroy BK, Walters JD, Blackshear PJ, Johnson JD. Phosphorylation-dependent binding of a synthetic MARCKS peptide to calmodulin. The Journal of biological chemistry. 1991;266(8):4959–64. 2002042.

15. Graff JM, Rajan RR, Randall RR, Nairn AC, Blackshear PJ. Protein kinase C substrate and inhibitor characteristics of peptides derived from the myristoylated alanine-rich C kinase substrate (MARCKS) protein phosphorylation site domain. The Journal of biological chemistry. 1991;266(22):14390–8. 1650359.

16. Bubb MR, Lenox RH, Edison AS. Phosphorylation-dependent conformational changes induce a switch in the actin-binding function of MARCKS. The Journal of biological chemistry. 1999;274(51):36472–8. doi: 10.1074/jbc.274.51.36472 10593944.

17. Swierczynski SL, Blackshear PJ. Myristoylation-dependent and electrostatic interactions exert independent effects on the membrane association of the myristoylated alanine-rich protein kinase C substrate protein in intact cells. The Journal of biological chemistry. 1996;271(38):23424–30. doi: 10.1074/jbc.271.38.23424 8798548.

18. El Amri M, Fitzgerald U, Schlosser G. MARCKS and MARCKS-like proteins in development and regeneration. J Biomed Sci. 2018;25(1):43. doi: 10.1186/s12929-018-0445-1 29788979; PubMed Central PMCID: PMC5964646.

19. Green TD, Crews AL, Park J, Fang S, Adler KB. Regulation of mucin secretion and inflammation in asthma: a role for MARCKS protein? Biochimica et biophysica acta. 2011;1810(11):1110–3. doi: 10.1016/j.bbagen.2011.01.009 21281703; PubMed Central PMCID: PMC3097255.

20. Hartwig JH, Thelen M, Rosen A, Janmey PA, Nairn AC, Aderem A. MARCKS is an actin filament crosslinking protein regulated by protein kinase C and calcium-calmodulin. Nature. 1992;356(6370):618–22. doi: 10.1038/356618a0 1560845.

21. Taniguchi H, Manenti S. Interaction of myristoylated alanine-rich protein kinase C substrate (MARCKS) with membrane phospholipids. The Journal of biological chemistry. 1993;268(14):9960–3. 8486722.

22. Park J, Fang S, Crews AL, Lin KW, Adler KB. MARCKS regulation of mucin secretion by airway epithelium in vitro: interaction with chaperones. American journal of respiratory cell and molecular biology. 2008;39(1):68–76. doi: 10.1165/rcmb.2007-0139OC 18314541; PubMed Central PMCID: PMC2438449.

23. Adler KB, Tuvim MJ, Dickey BF. Regulated mucin secretion from airway epithelial cells. Frontiers in endocrinology. 2013;4:129. Epub 2013/09/26. doi: 10.3389/fendo.2013.00129 24065956; PubMed Central PMCID: PMC3776272.

24. Iioka H, Ueno N, Kinoshita N. Essential role of MARCKS in cortical actin dynamics during gastrulation movements. The Journal of cell biology. 2004;164(2):169–74. Epub 2004/01/14. doi: 10.1083/jcb.200310027 [pii]. 14718521.

25. Wang YW, Wei CY, Dai HP, Zhu ZY, Sun YH. Subtractive phage display technology identifies zebrafish marcksb that is required for gastrulation. Gene. 2013;521(1):69–77. doi: 10.1016/j.gene.2013.03.028 23537994.

26. Stumpo DJ, Bock CB, Tuttle JS, Blackshear PJ. MARCKS deficiency in mice leads to abnormal brain development and perinatal death. Proceedings of the National Academy of Sciences of the United States of America. 1995;92(4):944–8. doi: 10.1073/pnas.92.4.944 7862670; PubMed Central PMCID: PMC42613.

27. Zolessi FR, Arruti C. Apical accumulation of MARCKS in neural plate cells during neurulation in the chick embryo. BMC Dev Biol. 2001;1:7. doi: 10.1186/1471-213X-1-7 11329360; PubMed Central PMCID: PMC31341.

28. Agathon A, Thisse C, Thisse B. The molecular nature of the zebrafish tail organizer. Nature. 2003;424(6947):448–52. Epub 2003/07/25. doi: 10.1038/nature01822 12879074

29. Xu XH, Deng CY, Liu Y, He M, Peng J, Wang T, et al. MARCKS regulates membrane targeting of Rab10 vesicles to promote axon development. Cell Res. 2014;24(5):576–94. doi: 10.1038/cr.2014.33 24662485; PubMed Central PMCID: PMC4011341.

30. Teleman AA, Cohen SM. Dpp gradient formation in the Drosophila wing imaginal disc. Cell. 2000;103(6):971–80. doi: 10.1016/s0092-8674(00)00199-9 11136981.

31. Deneke C, Lipowsky R, Valleriani A. Effect of ribosome shielding on mRNA stability. Physical biology. 2013;10(4):046008. doi: 10.1088/1478-3975/10/4/046008 23883670.

32. Lin F, Chen S, Sepich DS, Panizzi JR, Clendenon SG, Marrs JA, et al. Galpha12/13 regulate epiboly by inhibiting E-cadherin activity and modulating the actin cytoskeleton. The Journal of cell biology. 2009;184(6):909–21. Epub 2009/03/25. jcb.200805148 [pii] doi: 10.1083/jcb.200805148 19307601.

33. El-Brolosy MA, Stainier DYR. Genetic compensation: A phenomenon in search of mechanisms. PLoS Genet. 2017;13(7):e1006780. Epub 2017/07/14. doi: 10.1371/journal.pgen.1006780 28704371; PubMed Central PMCID: PMC5509088.

34. Rossi A, Kontarakis Z, Gerri C, Nolte H, Holper S, Kruger M, et al. Genetic compensation induced by deleterious mutations but not gene knockdowns. Nature. 2015;524(7564):230–3. doi: 10.1038/nature14580 26168398.

35. Kozmikova I, Candiani S, Fabian P, Gurska D, Kozmik Z. Essential role of Bmp signaling and its positive feedback loop in the early cell fate evolution of chordates. Developmental biology. 2013;382(2):538–54. doi: 10.1016/j.ydbio.2013.07.021 23933491.

36. Dick A, Hild M, Bauer H, Imai Y, Maifeld H, Schier AF, et al. Essential role of Bmp7 (snailhouse) and its prodomain in dorsoventral patterning of the zebrafish embryo. Development. 2000;127(2):343–54. 10603351.

37. Fang S, Crews AL, Chen W, Park J, Yin Q, Ren XR, et al. MARCKS and HSP70 interactions regulate mucin secretion by human airway epithelial cells in vitro. American journal of physiology Lung cellular and molecular physiology. 2013;304(8):L511–8. doi: 10.1152/ajplung.00337.2012 23377348; PubMed Central PMCID: PMC3625989.

38. Evans TG, Yamamoto Y, Jeffery WR, Krone PH. Zebrafish Hsp70 is required for embryonic lens formation. Cell Stress Chaperon. 2005;10(1):66–78. doi: 10.1379/Csc-79r.1 WOS:000227778100010. 15832949

39. Ott LE, McDowell ZT, Turner PM, Law JM, Adler KB, Yoder JA, et al. Two myristoylated alanine-rich C-kinase substrate (MARCKS) paralogs are required for normal development in zebrafish. Anatomical record. 2011;294(9):1511–24. doi: 10.1002/ar.21453 21809467; PubMed Central PMCID: PMC4103011.

40. Prieto D, Zolessi FR. Functional Diversification of the Four MARCKS Family Members in Zebrafish Neural Development. Journal of experimental zoology Part B, Molecular and developmental evolution. 2017;328(1–2):119–38. doi: 10.1002/jez.b.22691 27554589.

41. Li Y, Martin LD, Spizz G, Adler KB. MARCKS protein is a key molecule regulating mucin secretion by human airway epithelial cells in vitro. The Journal of biological chemistry. 2001;276(44):40982–90. doi: 10.1074/jbc.M105614200 11533058.

42. Park J, Fang S, Adler KB. Regulation of airway mucin secretion by MARCKS protein involves the chaperones heat shock protein 70 and cysteine string protein. Proc Am Thorac Soc. 2006;3(6):493. doi: 10.1513/pats.200603-067MS 16921125.

43. Agrawal A, Rengarajan S, Adler KB, Ram A, Ghosh B, Fahim M, et al. Inhibition of mucin secretion with MARCKS-related peptide improves airway obstruction in a mouse model of asthma. J Appl Physiol (1985). 2007;102(1):399–405. doi: 10.1152/japplphysiol.00630.2006 16946028.

44. Park JA, Crews AL, Lampe WR, Fang S, Park J, Adler KB. Protein kinase C delta regulates airway mucin secretion via phosphorylation of MARCKS protein. The American journal of pathology. 2007;171(6):1822–30. doi: 10.2353/ajpath.2007.070318 18055557; PubMed Central PMCID: PMC2111106.

45. Lampe WR, Park J, Fang S, Crews AL, Adler KB. Calpain and MARCKS protein regulation of airway mucin secretion. Pulm Pharmacol Ther. 2012;25(6):427–31. doi: 10.1016/j.pupt.2012.06.003 22710197; PubMed Central PMCID: PMC3486950.

46. Muthusamy N, Sommerville LJ, Moeser AJ, Stumpo DJ, Sannes P, Adler K, et al. MARCKS-dependent mucin clearance and lipid metabolism in ependymal cells are required for maintenance of forebrain homeostasis during aging. Aging cell. 2015;14(5):764–73. doi: 10.1111/acel.12354 26010231; PubMed Central PMCID: PMC4568964.

47. Müller P, Alexander. Extracellular Movement of Signaling Molecules. Developmental cell. 2011;21(1):145–58. doi: 10.1016/j.devcel.2011.06.001 21763615

48. ten Dijke P, Arthur HM. Extracellular control of TGFβ signalling in vascular development and disease. Nature Reviews Molecular Cell Biology. 2007;8:857. doi: 10.1038/nrm2262 17895899

49. Neugebauer JM, Kwon S, Kim HS, Donley N, Tilak A, Sopory S, et al. The prodomain of BMP4 is necessary and sufficient to generate stable BMP4/7 heterodimers with enhanced bioactivity in vivo. Proceedings of the National Academy of Sciences of the United States of America. 2015;112(18):E2307–16. Epub 2015/04/23. doi: 10.1073/pnas.1501449112 25902523; PubMed Central PMCID: PMC4426409.

50. Degnin C, Jean F, Thomas G, Christian JL. Cleavages within the prodomain direct intracellular trafficking and degradation of mature bone morphogenetic protein-4. Molecular biology of the cell. 2004;15(11):5012–20. Epub 2004/09/10. doi: 10.1091/mbc.E04-08-0673 15356272; PubMed Central PMCID: PMC524762.

51. von der Hardt S, Bakkers J, Inbal A, Carvalho L, Solnica-Krezel L, Heisenberg CP, et al. The Bmp gradient of the zebrafish gastrula guides migrating lateral cells by regulating cell-cell adhesion. Curr Biol. 2007;17(6):475–87. Epub 2007/03/03. S0960-9822(07)00988-8 [pii] doi: 10.1016/j.cub.2007.02.013 17331724.

52. Pollard TD, Borisy GG. Cellular motility driven by assembly and disassembly of actin filaments. Cell. 2003;112(4):453–65. doi: 10.1016/s0092-8674(03)00120-x 12600310.

53. El-Brolosy MA, Kontarakis Z, Rossi A, Kuenne C, Gunther S, Fukuda N, et al. Genetic compensation triggered by mutant mRNA degradation. Nature. 2019;568(7751):193–7. Epub 2019/04/05. doi: 10.1038/s41586-019-1064-z 30944477.

54. Ma Z, Zhu P, Shi H, Guo L, Zhang Q, Chen Y, et al. PTC-bearing mRNA elicits a genetic compensation response via Upf3a and COMPASS components. Nature. 2019;568(7751):259–63. Epub 2019/04/05. doi: 10.1038/s41586-019-1057-y 30944473.

55. Kimmel CB, Ballard WW, Kimmel SR, Ullmann B, Schilling TF. Stages of embryonic development of the zebrafish. Dev Dyn. 1995;203(3):253–310. Epub 1995/07/01. doi: 10.1002/aja.1002030302 8589427.

56. Nasevicius A, Ekker SC. Effective targeted gene 'knockdown' in zebrafish. Nature genetics. 2000;26(2):216–20. doi: 10.1038/79951 11017081.

57. Moreno-Mateos MA, Vejnar CE, Beaudoin JD, Fernandez JP, Mis EK, Khokha MK, et al. CRISPRscan: designing highly efficient sgRNAs for CRISPR-Cas9 targeting in vivo. Nat Methods. 2015;12(10):982–8. doi: 10.1038/nmeth.3543 26322839; PubMed Central PMCID: PMC4589495.

58. Yin L, Jao LE, Chen W. Generation of Targeted Mutations in Zebrafish Using the CRISPR/Cas System. Methods in molecular biology. 2015;1332:205–17. doi: 10.1007/978-1-4939-2917-7_16 26285757.

59. Zhang F, Wang H, Huang S, Xiong F, Zhu Z, Sun Y. [A comparison of the knockout efficiencies of two codon-optimized Cas9 coding sequences in zebrafish embryos]. Yi chuan = Hereditas. 2016;38(2):144–54. Epub 2016/02/26. doi: 10.16288/j.yczz.15-452 26907778.

60. Thisse C, Thisse B. High-resolution in situ hybridization to whole-mount zebrafish embryos. Nat Protoc. 2008;3(1):59–69. Epub 2008/01/15. nprot.2007.514 [pii] doi: 10.1038/nprot.2007.514 18193022.

61. Ye D, Lin F. S1pr2/Galpha13 signaling controls myocardial migration by regulating endoderm convergence. Development. 2013;140(4):789–99. doi: 10.1242/dev.085340 23318642; PubMed Central PMCID: PMC3557776.

62. 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. doi: 10.1038/nmeth.2019 22743772; PubMed Central PMCID: PMC3855844.

63. Weissgerber TL, Milic NM, Winham SJ, Garovic VD. Beyond bar and line graphs: time for a new data presentation paradigm. PLoS biology. 2015;13(4):e1002128. Epub 2015/04/23. doi: 10.1371/journal.pbio.1002128 25901488; PubMed Central PMCID: PMC4406565.

64. Lu FI, Sun YH, Wei CY, Thisse C, Thisse B. Tissue-specific derepression of TCF/LEF controls the activity of the Wnt/beta-catenin pathway. Nature communications. 2014;5:5368. doi: 10.1038/ncomms6368 25371059.

65. Kim D, Langmead B, Salzberg SL. HISAT: a fast spliced aligner with low memory requirements. Nature methods. 2015;12(4):357–60. doi: 10.1038/nmeth.3317 25751142; PubMed Central PMCID: PMC4655817.

66. Trapnell C, Hendrickson DG, Sauvageau M, Goff L, Rinn JL, Pachter L. Differential analysis of gene regulation at transcript resolution with RNA-seq. Nature biotechnology. 2013;31(1):46–53. doi: 10.1038/nbt.2450 23222703; PubMed Central PMCID: PMC3869392.

67. Trapnell C, Williams BA, Pertea G, Mortazavi A, Kwan G, van Baren MJ, et al. Transcript assembly and quantification by RNA-Seq reveals unannotated transcripts and isoform switching during cell differentiation. Nature biotechnology. 2010;28(5):511–5. doi: 10.1038/nbt.1621 20436464; PubMed Central PMCID: PMC3146043.

68. Love MI, Huber W, Anders S. Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome biology. 2014;15(12):550. doi: 10.1186/s13059-014-0550-8 25516281; PubMed Central PMCID: PMC4302049.

69. Pfaffl MW. A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res. 2001;29(9):e45. doi: 10.1093/nar/29.9.e45 11328886; PubMed Central PMCID: PMC55695.