Abstract
Purpose
Cell fate determinants Scrib and Llgl1 influence self-renewal capacity of hematopoietic stem cells (HSCs). Scrib-deficient HSCs are functionally impaired and lack sufficient repopulation capacity during serial transplantation and stress. In contrast, loss of Llgl1 leads to increased HSC fitness, gain of self-renewal capacity and expansion of the stem cell pool. Here, we sought to assess for shared and unique molecular functions of Llgl1 and Scrib by analyzing their interactome in hematopoietic cells.
Methods
Interactome analysis was performed by affinity purification followed by mass spectrometry. Motility, migration and adhesion were assessed on primary murine HSCs, which were isolated by FACS sorting following conditional deletion of Scrib or Llgl1, respectively. Imaging of Scrib-deficient HSCs was performed by intravital 2-photon microscopy.
Results
Comparison of Scrib and Llgl1 interactome analyses revealed involvement in common and unique cellular functions. Migration and adhesion were among the cellular functions connected to Scrib but not to Llgl1. Functional validation of these findings confirmed alterations in cell adhesion and migration of Scrib-deficient HSCs in vitro and in vivo. In contrast, genetic inactivation of Llgl1 did not affect adhesion or migratory capacity of hematopoietic stem cells.
Conclusion
Our data provide first evidence for an evolutionarily conserved role of the cell fate determinant Scrib in HSC adhesion and migration in vitro and in vivo, a unique function that is not shared with its putative complex partner Llgl1.
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Acknowledgements
We thank Ms. St. Frey and Mr. V.R. Thangapandi for technical assistance, A. Fenske DVM (Magdeburg) for his support with animal care and Dr. R. Hartig (Magdeburg) and Katrin Schubert (Core Facility Flow Cytometry, FLI, Jena) for their support with cell sorting. Moreover, we thank the Center for Molecular Medicine of the Austrian Academy of Sciences (CeMM, Vienna) for providing the pCeMM-NTAP vector.
Funding
This work was supported by a grant of the German Research Council (DFG HE6233/2-1) and in part by a grant of the Else-Kröner-Fresenius Stiftung (2012-A152) to F.H.H. Moreover, the project was supported by the Thuringian state program ProExzellenz (RegenerAging—FSU-I-03/14) of the Thuringian Ministry for Research (TMWWDG) to F.H.H. TBH was supported by the DFG (CRC1140, CRC 992), by the BMBF (01GM1518C), by the European Research Council-ERC Grant 616891 and by the H2020-IMI2 consortium BEAt-DKD.
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BPD, JM, CK, TMS, CH, SG, CZ, FP and TK performed experiments and analyzed data. MN, BH, TH and GW provided material. TH, TK, DSK and FHH analyzed data and provided material. TK and BPD contributed to the writing of the manuscript. FHH supervised the research and wrote the paper.
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This study does not involve any human samples or material. All animal experiments were conducted after approval by the Landesverwaltungsamt Saxony-Anhalt.
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Dash, B.P., Schnöder, T.M., Kathner, C. et al. Diverging impact of cell fate determinants Scrib and Llgl1 on adhesion and migration of hematopoietic stem cells. J Cancer Res Clin Oncol 144, 1933–1944 (2018). https://doi.org/10.1007/s00432-018-2724-3
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DOI: https://doi.org/10.1007/s00432-018-2724-3