Genetic, Immunofluorescence Labeling, and in Situ Hybridization Techniques in Identification of Stem Cells in Male and Female Germline Niches

Overview

Journal Methods Mol Biol

Specialty Molecular Biology

Date 2013 Aug 21

PMID 23959978

Citations 1

Authors

Shree Ram Singh

Ying Liu

Madhuri Kango-Singh

Eviatar Nevo

Affiliations

Soon will be listed here.

Abstract

Stem cells have an enormous capacity of self-renewal, as well as the ability to differentiate into specialized cell types. Proper control of these two properties of stem cells is crucial for animal development, growth control, and reproduction. Germline stem cells (GSCs) are a self-renewing population of germ cells, which generate haploid gametes (sperms or oocyte) that transmit genetic information from generation to generation. In Drosophila testis and ovary, GSCs are anchored around the niche cells. The cap cells cluster in females and hub cells in males act as a niche to control GSC behavior. With highly sophisticated genetic techniques in Drosophila, tremendous progress has been made in understanding the interactions between stem cells and niches at cellular and molecular levels. Here, we provide details of genetic, immunofluorescence labeling, and in situ hybridization techniques in identification and characterization of stem cells in Drosophila male and female germline niches.

Citing Articles

RNAi-Based Techniques for the Analysis of Gene Function in Drosophila Germline Stem Cells.

Blake A, Finger D, Hardy V, Ables E Methods Mol Biol. 2017; 1622:161-184.

PMID: 28674809 PMC: 5798201. DOI: 10.1007/978-1-4939-7108-4_13.

References

Xing Y, Kurtz I, Thuparani M, Legard J, Ruohola-Baker H . Loss-of-Function Screen Reveals Novel Regulators Required for Drosophila Germline Stem Cell Self-Renewal. G3 (Bethesda). 2012; 2(3):343-51. PMC: 3291504. DOI: 10.1534/g3.111.001651. View

Song X, Xie T . DE-cadherin-mediated cell adhesion is essential for maintaining somatic stem cells in the Drosophila ovary. Proc Natl Acad Sci U S A. 2002; 99(23):14813-8. PMC: 137501. DOI: 10.1073/pnas.232389399. View

Casper A, Baxter K, Van Doren M . no child left behind encodes a novel chromatin factor required for germline stem cell maintenance in males but not females. Development. 2011; 138(16):3357-66. PMC: 3143560. DOI: 10.1242/dev.067942. View

Toledano H, DAlterio C, Czech B, Levine E, Jones D . The let-7-Imp axis regulates ageing of the Drosophila testis stem-cell niche. Nature. 2012; 485(7400):605-10. PMC: 4829122. DOI: 10.1038/nature11061. View

Reya T, Morrison S, Clarke M, Weissman I . Stem cells, cancer, and cancer stem cells. Nature. 2001; 414(6859):105-11. DOI: 10.1038/35102167. View

Singh S, Chen X, Hou S . JAK/STAT signaling regulates tissue outgrowth and male germline stem cell fate in Drosophila. Cell Res. 2005; 15(1):1-5. DOI: 10.1038/sj.cr.7290255. View

Shivdasani A, Ingham P . Regulation of stem cell maintenance and transit amplifying cell proliferation by tgf-beta signaling in Drosophila spermatogenesis. Curr Biol. 2003; 13(23):2065-72. DOI: 10.1016/j.cub.2003.10.063. View

Wang H, Singh S, Zheng Z, Oh S, Chen X, Edwards K . Rap-GEF signaling controls stem cell anchoring to their niche through regulating DE-cadherin-mediated cell adhesion in the Drosophila testis. Dev Cell. 2006; 10(1):117-26. DOI: 10.1016/j.devcel.2005.11.004. View

Yamashita Y, Mahowald A, Perlin J, Fuller M . Asymmetric inheritance of mother versus daughter centrosome in stem cell division. Science. 2007; 315(5811):518-21. PMC: 2563045. DOI: 10.1126/science.1134910. View

10.

Sarkar A, Parikh N, Hearn S, Fuller M, Tazuke S, Schulz C . Antagonistic roles of Rac and Rho in organizing the germ cell microenvironment. Curr Biol. 2007; 17(14):1253-8. DOI: 10.1016/j.cub.2007.06.048. View

11.

Wang X, Pan L, Wang S, Zhou J, McDowell W, Park J . Histone H3K9 trimethylase Eggless controls germline stem cell maintenance and differentiation. PLoS Genet. 2012; 7(12):e1002426. PMC: 3245301. DOI: 10.1371/journal.pgen.1002426. View

12.

DiNardo S, Okegbe T, Wingert L, Freilich S, Terry N . lines and bowl affect the specification of cyst stem cells and niche cells in the Drosophila testis. Development. 2011; 138(9):1687-96. PMC: 3074445. DOI: 10.1242/dev.057364. View

13.

Schulz C, Wood C, Jones D, Tazuke S, Fuller M . Signaling from germ cells mediated by the rhomboid homolog stet organizes encapsulation by somatic support cells. Development. 2002; 129(19):4523-34. DOI: 10.1242/dev.129.19.4523. View

14.

Morrison S, Spradling A . Stem cells and niches: mechanisms that promote stem cell maintenance throughout life. Cell. 2008; 132(4):598-611. PMC: 4505728. DOI: 10.1016/j.cell.2008.01.038. View

15.

Rojas-Rios P, Guerrero I, Gonzalez-Reyes A . Cytoneme-mediated delivery of hedgehog regulates the expression of bone morphogenetic proteins to maintain germline stem cells in Drosophila. PLoS Biol. 2012; 10(4):e1001298. PMC: 3317903. DOI: 10.1371/journal.pbio.1001298. View

16.

Singh S, Zheng Z, Wang H, Oh S, Chen X, Hou S . Competitiveness for the niche and mutual dependence of the germline and somatic stem cells in the Drosophila testis are regulated by the JAK/STAT signaling. J Cell Physiol. 2010; 223(2):500-10. PMC: 2894562. DOI: 10.1002/jcp.22073. View

17.

Singh S, Liu W, Hou S . The adult Drosophila malpighian tubules are maintained by multipotent stem cells. Cell Stem Cell. 2008; 1(2):191-203. PMC: 2040309. DOI: 10.1016/j.stem.2007.07.003. View

18.

Bunt S, Hime G . Ectopic activation of Dpp signalling in the male Drosophila germline inhibits germ cell differentiation. Genesis. 2004; 39(2):84-93. DOI: 10.1002/gene.20030. View

19.

Lu W, Casanueva M, Mahowald A, Kato M, Lauterbach D, Ferguson E . Niche-associated activation of rac promotes the asymmetric division of Drosophila female germline stem cells. PLoS Biol. 2012; 10(7):e1001357. PMC: 3389017. DOI: 10.1371/journal.pbio.1001357. View

20.

Xie T, Spradling A . decapentaplegic is essential for the maintenance and division of germline stem cells in the Drosophila ovary. Cell. 1998; 94(2):251-60. DOI: 10.1016/s0092-8674(00)81424-5. View