Sarah Bowling

All Publications

• Mutation of p53 increases the competitive ability of pluripotent stem cells. Development (Cambridge, England) Perez Montero, S., Paul, P. K., di Gregorio, A., Bowling, S., Shepherd, S., Fernandes, N. J., Lima, A., Pérez-Carrasco, R., Rodriguez, T. A. 2024; 151 (2)

  Abstract

  During development, the rate of tissue growth is determined by the relative balance of cell division and cell death. Cell competition is a fitness quality-control mechanism that contributes to this balance by eliminating viable cells that are less fit than their neighbours. The mutations that confer cells with a competitive advantage and the dynamics of the interactions between winner and loser cells are not well understood. Here, we show that embryonic cells lacking the tumour suppressor p53 are 'super-competitors' that eliminate their wild-type neighbours through the direct induction of apoptosis. This elimination is context dependent, as it does not occur when cells are pluripotent and it is triggered by the onset of differentiation. Furthermore, by combining mathematical modelling and cell-based assays we show that the elimination of wild-type cells is not through competition for space or nutrients, but instead is mediated by short-range interactions that are dependent on the local cell neighbourhood. This highlights the importance of the local cell neighbourhood and the competitive interactions within this neighbourhood for the regulation of proliferation during early embryonic development.

  View details for DOI 10.1242/dev.202503

  View details for PubMedID 38131530

  View details for PubMedCentralID PMC10820806

• A mouse model with high clonal barcode diversity for joint lineage, transcriptomic, and epigenomic profiling in single cells. Cell Li, L., Bowling, S., McGeary, S. E., Yu, Q., Lemke, B., Alcedo, K., Jia, Y., Liu, X., Ferreira, M., Klein, A. M., Wang, S. W., Camargo, F. D. 2023; 186 (23): 5183-5199.e22

  Abstract

  Cellular lineage histories and their molecular states encode fundamental principles of tissue development and homeostasis. Current lineage-recording mouse models have insufficient barcode diversity and single-cell lineage coverage for profiling tissues composed of millions of cells. Here, we developed DARLIN, an inducible Cas9 barcoding mouse line that utilizes terminal deoxynucleotidyl transferase (TdT) and 30 CRISPR target sites. DARLIN is inducible, generates massive lineage barcodes across tissues, and enables the detection of edited barcodes in ∼70% of profiled single cells. Using DARLIN, we examined fate bias within developing hematopoietic stem cells (HSCs) and revealed unique features of HSC migration. Additionally, we established a protocol for joint transcriptomic and epigenomic single-cell measurements with DARLIN and found that cellular clonal memory is associated with genome-wide DNA methylation rather than gene expression or chromatin accessibility. DARLIN will enable the high-resolution study of lineage relationships and their molecular signatures in diverse tissues and physiological contexts.

  View details for DOI 10.1016/j.cell.2023.09.019

  View details for PubMedID 37852258

• DRP1 levels determine the apoptotic threshold during embryonic differentiation through a mitophagy-dependent mechanism. Developmental cell Pernaute, B., Pérez-Montero, S., Sánchez Nieto, J. M., Di Gregorio, A., Lima, A., Lawlor, K., Bowling, S., Liccardi, G., Tomás, A., Meier, P., Sesaki, H., Rutter, G. A., Barbaric, I., Rodríguez, T. A. 2022; 57 (11): 1316-1330.e7

  Abstract

  The changes that drive differentiation facilitate the emergence of abnormal cells that need to be removed before they contribute to further development or the germline. Consequently, in mice in the lead-up to gastrulation, ∼35% of embryonic cells are eliminated. This elimination is caused by hypersensitivity to apoptosis, but how it is regulated is poorly understood. Here, we show that upon exit of naive pluripotency, mouse embryonic stem cells lower their mitochondrial apoptotic threshold, and this increases their sensitivity to cell death. We demonstrate that this enhanced apoptotic response is induced by a decrease in mitochondrial fission due to a reduction in the activity of dynamin-related protein 1 (DRP1). Furthermore, we show that in naive pluripotent cells, DRP1 prevents apoptosis by promoting mitophagy. In contrast, during differentiation, reduced mitophagy levels facilitate apoptosis. Together, these results indicate that during early mammalian development, DRP1 regulation of mitophagy determines the apoptotic response.

  View details for DOI 10.1016/j.devcel.2022.04.020

  View details for PubMedID 35597240

  View details for PubMedCentralID PMC9297746

• Lifelong multilineage contribution by embryonic-born blood progenitors. Nature Patel, S. H., Christodoulou, C., Weinreb, C., Yu, Q., da Rocha, E. L., Pepe-Mooney, B. J., Bowling, S., Li, L., Osorio, F. G., Daley, G. Q., Camargo, F. D. 2022; 606 (7915): 747-753

  Abstract

  Haematopoietic stem cells (HSCs) arise in the embryo from the arterial endothelium through a process known as the endothelial-to-haematopoietic transition (EHT)1-4. This process generates hundreds of blood progenitors, of which a fraction go on to become definitive HSCs. It is generally thought that most adult blood is derived from those HSCs, but to what extent other progenitors contribute to adult haematopoiesis is not known. Here we use in situ barcoding and classical fate mapping to assess the developmental and clonal origins of adult blood in mice. Our analysis uncovers an early wave of progenitor specification-independent of traditional HSCs-that begins soon after EHT. These embryonic multipotent progenitors (eMPPs) predominantly drive haematopoiesis in the young adult, have a decreasing yet lifelong contribution over time and are the predominant source of lymphoid output. Putative eMPPs are specified within intra-arterial haematopoietic clusters and represent one fate of the earliest haematopoietic progenitors. Altogether, our results reveal functional heterogeneity during the definitive wave that leads to distinct sources of adult blood.

  View details for DOI 10.1038/s41586-022-04804-z

  View details for PubMedID 35705805

• Cell competition acts as a purifying selection to eliminate cells with mitochondrial defects during early mouse development. Nature metabolism Lima, A., Lubatti, G., Burgstaller, J., Hu, D., Green, A. P., Di Gregorio, A., Zawadzki, T., Pernaute, B., Mahammadov, E., Perez-Montero, S., Dore, M., Sanchez, J. M., Bowling, S., Sancho, M., Kolbe, T., Karimi, M. M., Carling, D., Jones, N., Srinivas, S., Scialdone, A., Rodriguez, T. A. 2021; 3 (8): 1091-1108

  Abstract

  Cell competition is emerging as a quality-control mechanism that eliminates unfit cells in a wide range of settings from development to the adult. However, the nature of the cells normally eliminated by cell competition and what triggers their elimination remains poorly understood. In mice, 35% of epiblast cells are eliminated before gastrulation. Here we show that cells with mitochondrial defects are eliminated by cell competition during early mouse development. Using single-cell transcriptional profiling of eliminated mouse epiblast cells, we identify hallmarks of cell competition and mitochondrial defects. We demonstrate that mitochondrial defects are common to a range of different loser cell types and that manipulating mitochondrial function triggers cell competition. Moreover, we show that in the mouse embryo, cell competition eliminates cells with sequence changes in mt-Rnr1 and mt-Rnr2, and that even non-pathological changes in mitochondrial DNA sequences can induce cell competition. Our results suggest that cell competition is a purifying selection that optimizes mitochondrial performance before gastrulation.

  View details for DOI 10.1038/s42255-021-00422-7

  View details for PubMedID 34253906

  View details for PubMedCentralID PMC7611553

• An Engineered CRISPR-Cas9 Mouse Line for Simultaneous Readout of Lineage Histories and Gene Expression Profiles in Single Cells. Cell Bowling, S., Sritharan, D., Osorio, F. G., Nguyen, M., Cheung, P., Rodriguez-Fraticelli, A., Patel, S., Yuan, W. C., Fujiwara, Y., Li, B. E., Orkin, S. H., Hormoz, S., Camargo, F. D. 2020; 181 (6): 1410-1422.e27

  Abstract

  Tracing the lineage history of cells is key to answering diverse and fundamental questions in biology. Coupling of cell ancestry information with other molecular readouts represents an important goal in the field. Here, we describe the CRISPR array repair lineage tracing (CARLIN) mouse line and corresponding analysis tools that can be used to simultaneously interrogate the lineage and transcriptomic information of single cells in vivo. This model exploits CRISPR technology to generate up to 44,000 transcribed barcodes in an inducible fashion at any point during development or adulthood, is compatible with sequential barcoding, and is fully genetically defined. We have used CARLIN to identify intrinsic biases in the activity of fetal liver hematopoietic stem cell (HSC) clones and to uncover a previously unappreciated clonal bottleneck in the response of HSCs to injury. CARLIN also allows the unbiased identification of transcriptional signatures associated with HSC activity without cell sorting.

  View details for DOI 10.1016/j.cell.2020.04.048

  View details for PubMedID 32413320

  View details for PubMedCentralID PMC7529102

• Genetic Deletion of Hesx1 Promotes Exit from the Pluripotent State and Impairs Developmental Diapause. Stem cell reports Pozzi, S., Bowling, S., Apps, J., Brickman, J. M., Rodriguez, T. A., Martinez-Barbera, J. P. 2019; 13 (6): 970-979

  Abstract

  The role of the homeobox transcriptional repressor HESX1 in embryonic stem cells (ESCs) remains mostly unknown. Here, we show that Hesx1 is expressed in the preimplantation mouse embryo, where it is required during developmental diapause. Absence of Hesx1 leads to reduced expression of epiblast and primitive endoderm determinants and failure of diapaused embryos to resume embryonic development after implantation. Genetic deletion of Hesx1 impairs self-renewal and promotes differentiation toward epiblast by reducing the expression of pluripotency factors and decreasing the activity of LIF/STAT3 signaling. We reveal that Hesx1-deficient ESCs show elevated ERK pathway activation, resulting in accelerated differentiation toward primitive endoderm, which can be prevented by overexpression of Hesx1. Together, our data provide evidence for a novel role of Hesx1 in the control of self-renewal and maintenance of the undifferentiated state in ESCs and mouse embryos.

  View details for DOI 10.1016/j.stemcr.2019.10.014

  View details for PubMedID 31761678

  View details for PubMedCentralID PMC6915801

• Cell competition: the winners and losers of fitness selection. Development (Cambridge, England) Bowling, S., Lawlor, K., Rodríguez, T. A. 2019; 146 (13)

  Abstract

  The process of cell competition results in the 'elimination of cells that are viable but less fit than surrounding cells'. Given the highly heterogeneous nature of our tissues, it seems increasingly likely that cells are engaged in a 'survival of the fittest' battle throughout life. The process has a myriad of positive roles in the organism: it selects against mutant cells in developing tissues, prevents the propagation of oncogenic cells and eliminates damaged cells during ageing. However, 'super-fit' cancer cells can exploit cell competition mechanisms to expand and spread. Here, we review the regulation, roles and risks of cell competition in organism development, ageing and disease.

  View details for DOI 10.1242/dev.167486

  View details for PubMedID 31278123

• P53 and mTOR signalling determine fitness selection through cell competition during early mouse embryonic development. Nature communications Bowling, S., Di Gregorio, A., Sancho, M., Pozzi, S., Aarts, M., Signore, M., D Schneider, M., Martinez-Barbera, J. P., Gil, J., Rodríguez, T. A. 2018; 9 (1): 1763

  Abstract

  Ensuring the fitness of the pluripotent cells that will contribute to future development is important both for the integrity of the germline and for proper embryogenesis. Consequently, it is becoming increasingly apparent that pluripotent cells can compare their fitness levels and signal the elimination of those cells that are less fit than their neighbours. In mammals the nature of the pathways that communicate fitness remain largely unknown. Here we identify that in the early mouse embryo and upon exit from naive pluripotency, the confrontation of cells with different fitness levels leads to an inhibition of mTOR signalling in the less fit cell type, causing its elimination. We show that during this process, p53 acts upstream of mTOR and is required to repress its activity. Finally, we demonstrate that during normal development around 35% of cells are eliminated by this pathway, highlighting the importance of this mechanism for embryonic development.

  View details for DOI 10.1038/s41467-018-04167-y

  View details for PubMedID 29720666

  View details for PubMedCentralID PMC5932021

• Cell Competition and Its Role in the Regulation of Cell Fitness from Development to Cancer. Developmental cell Di Gregorio, A., Bowling, S., Rodriguez, T. A. 2016; 38 (6): 621-34

  Abstract

  Cell competition is a cell fitness-sensing mechanism conserved from insects to mammals that eliminates those cells that, although viable, are less fit than their neighbors. An important implication of cell competition is that cellular fitness is not only a cell-intrinsic property but is also determined relative to the fitness of neighboring cells: a cell that is of suboptimal fitness in one context may be "super-fit" in the context of a different cell population. Here we discuss the mechanisms by which cell competition measures and communicates cell fitness levels and the implications of this mechanism for development, regeneration, and tumor progression.

  View details for DOI 10.1016/j.devcel.2016.08.012

  View details for PubMedID 27676435