Ribosomal DNA and the rDNA-binding protein Indra mediate non-random sister chromatid segregation in Drosophila male germline stem cells

George Watase, Yukiko Yamashita

Preprint posted on December 16, 2018

Sisters are not always identical: Chromosomal immortality in germline stem cells rests on rDNA loci and their binding partner Indra

Selected by Maiko Kitaoka

Categories: cell biology, genetics


In a canonical cell cycle, replicated sister chromatids are assumed to be identical and segregated randomly into daughter cells. However, sisters can be epigenetically distinct and can undergo non-random segregation. This has thought to contribute to asymmetric cell divisions, particularly in stem cells where one daughter maintains a stem cell state and the other can differentiate.

Here, Watase and Yamashita use the Drosophila male germline stem cells to investigate the non-random sister segregation of X and Y chromosomes. The group has previously shown that autosomes segregate randomly, while X and Y chromosomes do not, but how and why this phenomenon occurs is still unknown.

Key findings

The authors first identified ribosomal DNA (rDNA) to be a key locus on the X and Y chromosomes. D. melanogaster autosomes do not contain any rDNA loci and also segregate randomly, suggesting that this feature on the sex chromosomes is an important aspect. Deletion strains with no rDNA loci on the X or Y chromosomes showed random sister chromatid segregation, demonstrating the requirement of rDNA loci for non-random segregation.

Sex chromosomes segregate non-randomly, as the red and blue strands are inherited with a bias in germ cell divisions. The red strand ends up in the GSC (closest to the asterisk hub) in 80% of divisions, rather than an even 50/50 split (left images). However, when rDNA is perturbed on the X (bb158) or Y (Ybb) chromosomes, segregation becomes random (right graph). From Figure 1.


Since rDNA loci are very large and contain several individual and distinct elements, the authors compared D. melanogaster to D. simulans, whose Y chromosome still segregates non-randomly despite lacking several rDNA elements. Interestingly, both species had intergenic spacer sequence (IGS) repeats, so they attempted to identify any potential IGS-binding proteins by mass spectrometry. This allowed them to uncover 18 proteins enriched for IGS sequence binding, including an uncharacterized zinc finger protein. They named the gene indra after a Hindu god who lost immortality. Consistent with its proposed role with rDNA, Indra localizes to the nucleolus and to the rDNA loci on metaphase chromosomes.

In characterizing indra’s function, Watase and Yamashita discovered that RNAi knockdown of indra led to random sister chromatid segregation. A stronger loss of indra in the germline revealed severe fertility defects due to the rapid loss of germ cells (and loss of cellular immortality). Using DNA FISH, they found that X and Y chromosomes frequently have inter-homolog exchange at the rDNA loci when indra is depleted, indicating indra’s role in preventing recombination between the sex chromosomes. This could be accomplished by preventing DNA double stranded breaks at rDNA loci or by encouraging sister chromatid DNA repair, rather than homolog repair.

In indra knockdown mutants, the X and Y chromosomes recombine at the IGS region of the rDNA loci, likely leading to catastrophic mitoses and cell death. From Figure 4.


Interestingly, when indra is depleted from male germline, progeny exhibited a bobbed phenotype, where the stripes on the adult fly’s back are not continuous and straight across, which has been shown to be a hallmark of rDNA insufficiency. This further implicates the role of rDNA, so how does Indra help to maintain rDNA copy number? Unequal sister chromatid exchange is a proposed mechanism for rDNA copy number expansion, where one chromatid gains copy number at the expense of the other. The authors propose that non-random sister chromatid segregation may reflect non-random segregation of higher vs. lower rDNA copy number after unequal sister chromatid exchange. While they could not measure the absolute copy number at rDNA loci, the authors observed multiple sister chromatid exchanges at rDNA loci in indra-depleted GSCs, potentially equalizing the rDNA copy number between sister chromatids. This indicates that indra limits the number of sister chromatid exchanges, pointing to a mechanism where indra acts to ensure that unequal exchange is productive and allows for unequal expansion of rDNA in the stem cell.

Model describing how indra may mediate unequal sister chromatid exchange at rDNA loci to allow for differences in rDNA copy number between sisters. This inequality leads to the non-random segregation of sister chromatids in germ cells. From Figure 5.


To conclude, Watase and Yamashita have discovered a mechanistic explanation to non-random sister chromatid segregation in asymmetric cell division, proving not only the importance of rDNA loci and the newly-named IGS binding protein Indra but also shedding light on the unique identities of sister chromatids. This exciting work opens many more questions for future investigation. Most immediately, it would be interesting to see whether an Indra binding site/IGS repeats are sufficient to induce non-random segregation on the D. melanogaster autosomes. Although, given that the rDNA locus is already very large, artificially adding it to autosomes may prove to be a technical challenge. Second, and more physiologically, what advantage do GSCs maintain by inheriting the sister chromatid with an expanded rDNA copy number? Perhaps the ability of GSCs to undergo repeated rounds of asymmetric cell division may contribute to this, since they would add rDNA copies gradually over time instead of all at once. Finally, the mass spectrometry pull-down experiment identified several other proteins as well that bound to IGS repeats. While some candidates are probably essential for cell survival, it will be interesting to investigate these further and see how they also interact with the rDNA loci and/or Indra.

Questions for the authors

The D. simulans Y chromosome has the Indra binding site and IGS repeats, but no other rDNA loci elements, so do they still show rDNA expansion? If not, how might the sister chromatids segregate non-randomly without the unequal rDNA expansion?

Can you speculate on Indra’s mechanism of action after binding to IGS repeats? How does the binding prevent inter-homolog exchange and/or multiple sister chromatid exchanges?

In somatic stem cells, do the sex chromosomes segregate non-randomly and the autosomes randomly as well?

Tags: asymmetric cell division, cell biology, drosophila, genetics, rdna, ribosomal dna, stem cells

Posted on: 20th December 2018


Read preprint (No Ratings Yet)

  • Have your say

    Your email address will not be published. Required fields are marked *

    This site uses Akismet to reduce spam. Learn how your comment data is processed.

    Sign up to customise the site to your preferences and to receive alerts

    Register here

    preLists in the cell biology category:

    Planar Cell Polarity – PCP

    This preList contains preprints about the latest findings on Planar Cell Polarity (PCP) in various model organisms at the molecular, cellular and tissue levels.


    List by Ana Dorrego-Rivas

    BioMalPar XVI: Biology and Pathology of the Malaria Parasite

    [under construction] Preprints presented at the (fully virtual) EMBL BioMalPar XVI, 17-18 May 2020 #emblmalaria


    List by Gautam Dey, Samantha Seah


    Cell Polarity

    Recent research from the field of cell polarity is summarized in this list of preprints. It comprises of studies focusing on various forms of cell polarity ranging from epithelial polarity, planar cell polarity to front-to-rear polarity.


    List by Yamini Ravichandran

    TAGC 2020

    Preprints recently presented at the virtual Allied Genetics Conference, April 22-26, 2020. #TAGC20


    List by Maiko Kitaoka, Madhuja Samaddar, Miguel V. Almeida, Sejal Davla, Jennifer Ann Black, Gautam Dey

    3D Gastruloids

    A curated list of preprints related to Gastruloids (in vitro models of early development obtained by 3D aggregation of embryonic cells)


    List by Paul Gerald L. Sanchez and Stefano Vianello

    ECFG15 – Fungal biology

    Preprints presented at 15th European Conference on Fungal Genetics 17-20 February 2020 Rome


    List by Hiral Shah

    ASCB EMBO Annual Meeting 2019

    A collection of preprints presented at the 2019 ASCB EMBO Meeting in Washington, DC (December 7-11)


    List by Madhuja Samaddar, Ramona Jühlen, Amanda Haage, Laura McCormick, Maiko Kitaoka

    EMBL Seeing is Believing – Imaging the Molecular Processes of Life

    Preprints discussed at the 2019 edition of Seeing is Believing, at EMBL Heidelberg from the 9th-12th October 2019


    List by Gautam Dey


    Preprints on autophagy and lysosomal degradation and its role in neurodegeneration and disease. Includes molecular mechanisms, upstream signalling and regulation as well as studies on pharmaceutical interventions to upregulate the process.


    List by Sandra Malmgren Hill

    Lung Disease and Regeneration

    This preprint list compiles highlights from the field of lung biology.


    List by Rob Hynds

    Cellular metabolism

    A curated list of preprints related to cellular metabolism at Biorxiv by Pablo Ranea Robles from the Prelights community. Special interest on lipid metabolism, peroxisomes and mitochondria.


    List by Pablo Ranea Robles

    BSCB/BSDB Annual Meeting 2019

    Preprints presented at the BSCB/BSDB Annual Meeting 2019


    List by Gautam Dey


    This list of preprints is focused on work expanding our knowledge on mitochondria in any organism, tissue or cell type, from the normal biology to the pathology.


    List by Sandra Franco Iborra

    Biophysical Society Annual Meeting 2019

    Few of the preprints that were discussed in the recent BPS annual meeting at Baltimore, USA


    List by Joseph Jose Thottacherry

    ASCB/EMBO Annual Meeting 2018

    This list relates to preprints that were discussed at the recent ASCB conference.


    List by Gautam Dey, Amanda Haage