Banaja P Dash1, Tina M Schnöder1,2, Carolin Kathner1,2, Juliane Mohr3,4, Sönke Weinert5, Carolin Herzog4, Parimala Sonika Godavarthy6, Costanza Zanetti6, Florian Perner1,2, Rüdiger Braun-Dullaeus5, Björn Hartleben7, Tobias B Huber8,9,10, Gerd Walz9, Michael Naumann11, Sarah Ellis12, Valera Vasioukhin13, Thilo Kähne11, Daniela S Krause6, Florian H Heidel14,15. 1. Innere Medizin II, Hämatologie und Onkologie, Universitätsklinikum Jena, Am Klinikum 1, 07747, Jena, Germany. 2. Leibniz Institute on Aging, Fritz-Lipmann Institute, Jena, Germany. 3. Institute for Molecular and Clinical Immunology, Otto-von-Guericke University Medical Center, Magdeburg, Germany. 4. Department of Hematology and Oncology, Otto-von-Guericke University Medical Center, Magdeburg, Germany. 5. Department of Cardiology and Angiology, Otto-von-Guericke University Medical Center, Magdeburg, Germany. 6. Institute for Tumor Biology and Experimental Therapy, Georg-Speyer-Haus, Frankfurt am Main, Germany. 7. Institute of Pathology, Medizinische Hochschule Hannover, Hannover, Germany. 8. III Department of Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany. 9. Department of Medicine IV, Medical Center, University of Freiburg, Freiburg, Germany. 10. BIOSS Center for Biological Signaling Studies, Albert-Ludwigs-University, Freiburg, Germany. 11. Institute for Experimental Medicine, Otto-von-Guericke University Medical Center, Magdeburg, Germany. 12. Cancer Research Division, Peter MacCallum Cancer Centre, Melbourne, Australia. 13. Division of Human Biology, Fred Hutchinson Cancer Research Center, Seattle, WA, USA. 14. Innere Medizin II, Hämatologie und Onkologie, Universitätsklinikum Jena, Am Klinikum 1, 07747, Jena, Germany. Florian.Heidel@med.uni-jena.de. 15. Leibniz Institute on Aging, Fritz-Lipmann Institute, Jena, Germany. Florian.Heidel@med.uni-jena.de.
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.
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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