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Lis1 regulates asymmetric division in hematopoietic stem cells and in leukemia

Abstract

Cell fate can be controlled through asymmetric division and segregation of protein determinants, but the regulation of this process in the hematopoietic system is poorly understood. Here we show that the dynein-binding protein Lis1 is critically required for hematopoietic stem cell function and leukemogenesis. Conditional deletion of Lis1 (also known as Pafah1b1) in the hematopoietic system led to a severe bloodless phenotype, depletion of the stem cell pool and embryonic lethality. Further, real-time imaging revealed that loss of Lis1 caused defects in spindle positioning and inheritance of cell fate determinants, triggering accelerated differentiation. Finally, deletion of Lis1 blocked the propagation of myeloid leukemia and led to a marked improvement in survival, suggesting that Lis1 is also required for oncogenic growth. These data identify a key role for Lis1 in hematopoietic stem cells and mark its directed control of asymmetric division as a critical regulator of normal and malignant hematopoietic development.

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Figure 1: Genetic deletion of Lis1 impairs establishment of the hematopoietic system during embryonic development.
Figure 2: Lis1 is required cell autonomously for adult HSC self-renewal.
Figure 3: Lis1 deficiency leads to accelerated differentiation of HSCs.
Figure 4: Loss of Lis1 impairs inheritance of fate determinants in hematopoietic development.
Figure 5: Loss of Lis1 impairs spindle orientation in hematopoietic development.
Figure 6: Loss of Lis1 impairs the development and propagation of myeloid leukemia in mouse models and human leukemia cells.

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Acknowledgements

We are grateful to B. Hogan, J. Chang, A. Desai, J. Gleeson, J.E. Lee, M.F. Wu, M. Sander and J. Koop for experimental advice and reagents. We would also like to thank M. Kritzik for advice and comments on the manuscript; M. Nakamura for experimental help; M. Cook, L. Matinek, B. Harvat, E. O'Conner and K. Marquez for cell sorting; and W. Pear (University of Pennsylvania) and A.M. Pendergast (Duke University) for the BCR-ABL construct, D.G. Gilliland (University of Pennsylvania) for the NUP98-HOXA9 construct, S. Armstrong (Memorial Sloan-Kettering Cancer Center) for the MLL-AF9 construct, C. Counter (Duke University) for NRASG12V, S. Russell (Peter MacCallum Cancer Centre) for the mCherry–α-tubulin construct, G. Wahl (Salk Institute) for the H2B-GFP (pEGFPN1) vector and D. Kioussis (Medical Research Council National Institute for Medical Research) for the Vav1-cre transgenic line. B.Z. and C.S.K. received support from US National Institutes of Health (NIH) Cancer Biology Training Grant (T32 CA 59365-18) and NIH Pharmacological Sciences Training Program (T32 GM007752), respectively. T.I. is a recipient of a California Institute for Regenerative Medicine interdisciplinary stem cell training program fellowship, and T.K. is supported by a postdoctoral fellowship from the Japanese Society for the Promotion of Science. This work was also supported by a Leukemia and Lymphoma Society Scholar Award, the University of California San Diego Moores Cancer Center National Cancer Institute Core Grant, P30CA23100, as well as by NIH grants DK63031, HL097767 and DP1 CA174422 awarded to T.R.

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Contributions

B.Z. and T.I. planned and designed the research, performed the majority of experiments and helped write the manuscript. B.Z. and J.B. developed all real-time imaging methods for visualizing and tracking spindle orientation in primary hematopoietic cells. A.B., J.B., T.K., J.W., C.S.K., H.Y.K. and O.A. provided experimental data and help. D.R., H.E.B., C.C. and V.G.O. provided primary patient samples and experimental advice. R.S. and G.H. carried out all bioinformatics analysis on microarray data. T.R. planned and guided the project, provided experimental advice and wrote the manuscript.

Corresponding author

Correspondence to Tannishtha Reya.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–20, Supplementary Tables 1 and 2. (PDF 32604 kb)

Imaging Cell Division in Real Time

Cells were co-infected with H2B-GFP and mCherry-α-tubulin fusion constructs and imaged over time. Representative movies show A. HeLa cell, B. M1 Cell, and C. Primary Hematopoietic Stem & Progenitor Cells undergoing cell division (H2B-GFP is shown in green and α-tubulin is shown in magenta). (MOV 4040 kb)

Symmetric inheritance of Numb

HSC-enriched cells (KLS) were co-infected with Numb-CFP and mCherry-α-tubulin fusion constructs and imaged over time. Representative movie shows a KLS cell undergoing symmetric division (Numb is shown in green and α-tubulin is shown in red). (MOV 15036 kb)

Asymmetric inheritance of Numb

HSC-enriched cells (KLS) were co-infected with Numb-CFP and mCherry-α-tubulin fusion constructs and imaged over time. Representative movie shows a KLS cell undergoing asymmetric division (Numb is shown in green and α-tubulin is shown in red). (MOV 16403 kb)

Asymmetric inheritance of Numb

HSC-enriched cells (KLS) were co-infected with Numb-YFP and mCherry-α-tubulin fusion constructs and imaged over time. Representative movie shows a KLS cell undergoing asymmetric division (Numb is shown in green and α-tubulin is shown in red). (MOV 12302 kb)

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Zimdahl, B., Ito, T., Blevins, A. et al. Lis1 regulates asymmetric division in hematopoietic stem cells and in leukemia. Nat Genet 46, 245–252 (2014). https://doi.org/10.1038/ng.2889

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