한국발생생물학회지:발생과생식 (Development and Reproduction)

  • 제27권1호
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  • Pages.39-46
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  • 2023
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  • 2465-9525(pISSN)
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  • 2465-9541(eISSN)
한국발생생물학회 (The Korean Society of Developmental Biology)
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A Pumilio Activity Sensor Reveals Bag-of-Marbles Inhibition of Pum Activity in the Drosophila Ovary

  • Wijeong Jang (School of Biological Sciences and Technology, Chonnam National University) ;
  • Changsoo Kim (School of Biological Sciences and Technology, Chonnam National University)
  • 투고 : 2023.01.08
  • 심사 : 2023.03.17
  • 발행 : 2023.03.15
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초록

Pumilio (Pum) is an RNA-binding protein and translational repressor important to diverse biological processes. In the Drosophila ovary, Pum is expressed in female germline stem cells (GSCs), wherein it acts as an intrinsic stem cell maintenance factor via repressing target mRNAs that are as yet mostly unknown. Pum recognizes the Pum binding sequence (PBS) in the mRNA 3'UTR through its C-terminus Puf domain. Translational repression is mediated through its N-terminal domain, but the molecular mechanism remains largely unknown. We previously showed that Bag-of-marbles, a critical differentiation-promoting factor of female GSCs, physically interacts with the N-terminus of Pum. We further showed that this interaction is critical to Bam inhibition of Pum repressive action in cultured cells, but the physiological relevance was not addressed. Here we design an in vivo GFP translational reporter bearing the PBS in its 3'UTR and show that GFP expression is reduced in cells wherein Pum is known to be active. Furthermore, we demonstrate in pum mutant ovary that this GFP repression requires Pum, and also that the sensor faithfully monitors Pum activity. Finally, we show that forced expression of Bam inhibits Pum-mediated repression, validating that Bam inhibits Pum activity in vivo.

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과제정보

This research was supported by grants from the National University Development Project 2021-4065/202233410001 (Changsoo Kim) and NRF-2020R1I1A1A01074292 (Wijeong Jang). We thank Dr. Cohen for the GFP translational reporter vector, which was used to make the Pum activity sensor. We are grateful to Dr. McKearin and Dr. Wharton for the flies.

참고문헌

  1. Arvola RM, Chang CT, Buytendorp JP, Levdansky Y, Valkov E, Freddolino PL, Goldstrohm AC (2020) Unique repression domains of Pumilio utilize deadenylation and decapping factors to accelerate destruction of target mRNAs. Nucleic Acids Res 48:1843-1871. https://doi.org/10.1093/nar/gkz1187
  2. Brennecke J, Cohen SM (2003) Towards a complete description of the microRNA complement of animal genomes. Genome Biol 4:228. https://doi.org/10.1186/gb-2003-4-9-228
  3. Deng W, Lin H (1997) Spectrosomes and fusomes anchor mitotic spindles during asymmetric germ cell divisions and facilitate the formation of a polarized microtubule array for oocyte specification in Drosophila. Dev Biol 189:79-94. https://doi.org/10.1006/dbio.1997.8669
  4. Edwards TA, Pyle SE, Wharton RP, Aggarwal AK (2001) Structure of Pumilio reveals similarity between RNA and peptide binding motifs. Cell 105:281-289. https://doi.org/10.1016/S0092-8674(01)00318-X
  5. Harris RE, Pargett M, Sutcliffe C, Umulis D, Ashe HL (2011) Brat promotes stem cell differentiation via control of a bistable switch that restricts BMP signaling. Dev Cell 20:72-83. https://doi.org/10.1016/j.devcel.2010.11.019
  6. Kim JY, Lee YC, Kim C (2010) Direct inhibition of Pumilo activity by Bam and Bgcn in Drosophila germ line stem cell differentiation. J Biol Chem 285:4741-4746. https://doi.org/10.1074/jbc.M109.002014
  7. Lavoie CA, Ohlstein B, McKearin DM (1999) Localization and function of Bam protein require the benign gonial cell neoplasm gene product. Dev Biol 212:405-413. https://doi.org/10.1006/dbio.1999.9346
  8. Lin H, Spradling AC (1993) Germline stem cell division and egg chamber development in transplanted Drosophila germaria. Dev Biol 159:140-152. https://doi.org/10.1006/dbio.1993.1228
  9. Lin H, Spradling AC (1995) Fusome asymmetry and oocyte determination in Drosophila. Dev Genet 16:6-12. https://doi.org/10.1002/dvg.1020160104
  10. Lin H, Spradling AC (1997) A novel group of pumilio mutations affects the asymmetric division of germline stem cells in the Drosophila ovary. Development 124:2463-2476. https://doi.org/10.1242/dev.124.12.2463
  11. McKearin DM, Spradling AC (1990) Bag-of-marbles: A Drosophila gene required to initiate both male and female gametogenesis. Genes Dev 4:2242-2251. https://doi.org/10.1101/gad.4.12b.2242
  12. Murata Y, Wharton RP (1995) Binding of pumilio to maternal hunchback mRNA is required for posterior patterning in Drosophila embryos. Cell 80:747-756. https://doi.org/10.1016/0092-8674(95)90353-4
  13. Ohlstein B, Lavoie CA, Vef O, Gateff E, McKearin DM (2000) The Drosophila cystoblast differentiation factor, benign gonial cell neoplasm, is related to DExH-box proteins and interacts genetically with bag-of-marbles. Genetics 155:1809-1819. https://doi.org/10.1093/genetics/155.4.1809
  14. Ohlstein B, McKearin D (1997) Ectopic expression of the Drosophila Bam protein eliminates oogenic germline stem cells. Development 124:3651-3662. https://doi.org/10.1242/dev.124.18.3651
  15. Parisi M, Lin H (1999) The Drosophila pumilio gene encodes two functional protein isoforms that play multiple roles in germline development, gonadogenesis, oogenesis and embryogenesis. Genetics 153:235-250. https://doi.org/10.1093/genetics/153.1.235
  16. Qiu C, Dutcher RC, Porter DF, Arava Y, Wickens M, Hall TMT (2019) Distinct RNA-binding modules in a single PUF protein cooperate to determine RNA specificity. Nucleic Acids Res 47:8770-8784. https://doi.org/10.1093/nar/gkz583
  17. Szakmary A, Cox DN, Wang Z, Lin H (2005) Regulatory relationship among piwi, pumilio, and bag-of-marbles in Drosophila germline stem cell self-renewal and differentiation. Curr Biol 15:171-178. https://doi.org/10.1016/j.cub.2005.01.005
  18. Wang Z, Lin H (2004) Nanos maintains germline stem cell self-renewal by preventing differentiation. Science 303:2016-2019. https://doi.org/10.1126/science.1093983
  19. Weidmann CA, Goldstrohm AC (2012) Drosophila Pumilio protein contains multiple autonomous repression domains that regulate mRNAs independently of Nanos and brain tumor. Mol Cell Biol 32:527-540.
  20. Weidmann CA, Qiu C, Arvola RM, Lou TF, Killingsworth J, Campbell ZT, Tanaka Hall TM, Goldstrohm AC (2016) Drosophila Nanos acts as a molecular clamp that modulates the RNAbinding and repression activities of Pumilio. eLife 5:e17096. https://doi.org/10.7554/eLife.17096
  21. Wharton RP, Sonoda J, Lee T, Patterson M, Murata Y (1998) The Pumilio RNA-binding domain is also a translational regulator. Mol Cell 1:863-872. https://doi.org/10.1016/S1097-2765(00)80085-4
  22. Wharton RP, Struhl G (1991) RNA regulatory elements mediate control of Drosophila body pattern by the posterior morphogen nanos. Cell 67:955-967. https://doi.org/10.1016/0092-8674(91)90368-9
  23. Xie T, Spradling AC (1998) Decapentaplegic is essential for the maintenance and division of germline stem cells in the Drosophila ovary. Cell 94:251-260. https://doi.org/10.1016/S0092-8674(00)81424-5
  24. Xie T, Spradling AC (2000) A niche maintaining germ line stem cells in the Drosophila ovary. Science 290:328-330. https://doi.org/10.1126/science.290.5490.328
  25. Zamore PD, Williamson JR, Lehmann R (1997) The Pumilio protein binds RNA through a conserved domain that defines a new class of RNA-binding proteins. RNA 3:1421-1433.