Research

Asymmetric stem cell division (ACD) produces one stem cell and one differentiating cell. Fate-determining factors (intrinsic or extrinsic) are partitioned into the two daughter cells asymmetrically through programmed orientation of the mitotic spindle. Because each stem cell undergoes ACD produces one stem cell and one differentiating cell at the same time, it is essential for maintaining multiple stem cell clones. Defective ACD accelerates the clonal expansion of stem cells, which is observed in various pathogenic conditions including clonal hematopoiesis and clonal evolution (1-2). Our research goal is to elucidate the molecular and cellular mechanisms of ACD, explaining how two daughter cells of different fates are made after only one cell division. We primarily uses the Drosophila testicular niche, in which we can monitor asymmetric division of germline stem cells in vivo. In particular, we use a combination of fly genetics with various imaging techniques, including immunofluorescence, single molecule RNA fluorescent in situ hybridization (smFISH), OligoPaint DNA FISH, and whole-mount tissue live-imaging by superresolution microscopy. Owing to the simple anatomy and abundant genetically encoded imaging tools, this system allows us to discover previously unrecognized regulatory mechanisms.

 

How extrinsic factor regulates asymmetry?

In the Drosophila testis, germline stem cells (GSCs) are continuously producing sperm throughout the fly’s lifetime. When a GSC divides in niche, one daughter cell remains a GSC, while the other daughter cell starts differentiation. This mechanism, called asymmetric division, is essential for maintaining GSCs in the niche for a long time.

Niche derived BMP ligand only activate GSCs but not other daughter cells despite of physical proximity of these cells. MT (microtubule based)-nanotubes provide the platform of receptor-ligand interaction contributing to the highly selective niche-stem cell interaction. BMP receptor present on MT-nanotubes is constantly transferred to hub cells and subsequently degraded. This mechanism limits the localization of receptor only on MT-nanotubes.

 

How intrinsic factor regulates asymmetry?

The pairing of homologous chromosomes is a fundamental process for meiotic recombination to exchange maternal and paternal genomic information. Homolog pairing also occurs in non-meiotic cells in a broad range of organisms including mammals (3, 4). However, the regulatory mechanism and significance of non-meiotic homolog pairing is less understood. Recent studies using 3D genomics and FISH analyses have indicated that non-meiotic homolog pairing is an actively regulated process and provides an opportunity for interchromosomal interaction (4-6). The local pairing status of a particular gene locus differs in a cell type-specific manner (7) and also correlates with local chromatin status (6), suggesting the possibility that non-meiotic homolog pairing may take part in gene regulation. Currently, it is still unclear whether pairing is a cause or a consequence of local chromatin status.

We recently found a change in the physical interaction of a homologous gene locus of a stem-cell specific protein, Signal transducer and activator of transcription 92E (Stat92E), during asymmetric division of GSCs. Stat92E is a highly conserved transcription factor required for stem cell identity, and is exclusively expressed in GSCs (8). We found that the stat92E locus on two homologous chromosomes interacts closely (paired) in GSCs but becomes separated (unpaired) in the differentiating daughter cells, gonialblasts (GBs). OligoPaint DNA FISH (9) analyses revealed that the observed change in pairing is locus-specific (i.e. other gene loci are always paired). These data suggest that the pairing of the stat92E locus is uniquely regulated during the cell-fate switch and is correlated with its transcriptional activity (10). We are currently investigating the roles of local homologous chromosome pairing for cell fate determination. Failure of proper gene regulation will affect the asymmetric outcome of stem cell division, which is implicated in multiple pathogenic conditions. Therefore, knowledge obtained through this project has the potential to contribute to the development of new therapeutic approaches for multiple diseases.

Pairing of the stat92E locus is regulated during differentiation.

A The stat92E locus shows paired pattern in GSC and unpaired pattern in GB and 2-4 SG stage in wild-type testis. White arrows indicate red “puncta” that indicate the position of the stat92E locus. B Measured distance between puncta in A. C A control locus (lacO) shows a paired pattern in these stages. D Measured distance between puncta in C. E Measured distance between puncta in stat92E Df/+.

 

 

 

  1. Jaiswal S, Ebert BL. Clonal hematopoiesis in human aging and disease. Science. 2019;366(6465):eaan4673. doi: doi:10.1126/science.aan4673.
  2. Greaves M, Maley CC. Clonal evolution in cancer. Nature. 2012;481(7381):306-13. doi: 10.1038/nature10762.
  3. Apte MS, Meller VH. Homologue Pairing in Flies and Mammals: Gene Regulation When Two Are Involved. Genetics Research International. 2012;2012:430587. doi: 10.1155/2012/430587.
  4. Joyce EF, Erceg J, Wu CT. Pairing and anti-pairing: a balancing act in the diploid genome. Curr Opin Genet Dev. 2016;37:119-28. Epub 2016/04/12. doi: 10.1016/j.gde.2016.03.002. PubMed PMID: 27065367; PMCID: PMC4939289.
  5. Bateman JR, Larschan E, D'Souza R, Marshall LS, Dempsey KE, Johnson JE, Mellone BG, Kuroda MI. A genome-wide screen identifies genes that affect somatic homolog pairing in Drosophila. G3 (Bethesda). 2012;2(7):731-40. Epub 2012/08/08. doi: 10.1534/g3.112.002840. PubMed PMID: 22870396; PMCID: PMC3385979.
  6. Erceg J, AlHaj Abed J, Goloborodko A, Lajoie BR, Fudenberg G, Abdennur N, Imakaev M, McCole RB, Nguyen SC, Saylor W, Joyce EF, Senaratne TN, Hannan MA, Nir G, Dekker J, Mirny LA, Wu CT. The genome-wide multi-layered architecture of chromosome pairing in early Drosophila embryos. Nat Commun. 2019;10(1):4486. Epub 2019/10/05. doi: 10.1038/s41467-019-12211-8. PubMed PMID: 31582744; PMCID: PMC6776651.
  7. Viets K, Sauria MEG, Chernoff C, Rodriguez Viales R, Echterling M, Anderson C, Tran S, Dove A, Goyal R, Voortman L, Gordus A, Furlong EEM, Taylor J, Johnston RJ, Jr. Characterization of Button Loci that Promote Homologous Chromosome Pairing and Cell-Type-Specific Interchromosomal Gene Regulation. Dev Cell. 2019;51(3):341-56 e7. Epub 2019/10/15. doi: 10.1016/j.devcel.2019.09.007. PubMed PMID: 31607649; PMCID: PMC6934266.
  8. Herrera SC, Bach EA. JAK/STAT signaling in stem cells and regeneration: from Drosophila to vertebrates. Development. 2019;146(2). Epub 2019/01/31. doi: 10.1242/dev.167643. PubMed PMID: 30696713; PMCID: PMC6361132.
  9. Beliveau BJ, Boettiger AN, Avendano MS, Jungmann R, McCole RB, Joyce EF, Kim-Kiselak C, Bantignies F, Fonseka CY, Erceg J, Hannan MA, Hoang HG, Colognori D, Lee JT, Shih WM, Yin P, Zhuang X, Wu CT. Single-molecule super-resolution imaging of chromosomes and in situ haplotype visualization using Oligopaint FISH probes. Nat Commun. 2015;6:7147. Epub 2015/05/13. doi: 10.1038/ncomms8147. PubMed PMID: 25962338; PMCID: PMC4430122.
  10. Antel M, Masoud M, Raj R, Pan Z, Li S, Mellone BG, Inaba M. Interchromosomal interaction of homologous Stat92E alleles regulates transcriptional switch during stem-cell differentiation. bioRxiv. 2021:2021.11.08.467622. doi: 10.1101/2021.11.08.467622.