Green fluorescent protein transgene driven by Kit regulatory sequences is expressed in hematopoietic stem cells

• Published in: Haematologica (Roma) Vol. 94; no. 3; pp. 318 - 325
• Main Authors: Cerisoli, Francesco, Cassinelli, Letizia, Lamorte, Giuseppe, Citterio, Stefania, Bertolotti, Francesca, Magli, Maria Cristina, Ottolenghi, Sergio
• Format: Journal Article
• Language: English
• Published: Pavia Ferrata Storti Foundation 01.03.2009
• Subjects: Animals; Biological and medical sciences; Bone Marrow - metabolism; Bone Marrow Transplantation; Cell Separation; Colony-Forming Units Assay; Female; Flow Cytometry - methods; Green Fluorescent Proteins - genetics; Green Fluorescent Proteins - metabolism; Hematologic and hematopoietic diseases; Hematopoietic Stem Cells; Hematopoietic Stem Cells - cytology; Hematopoietic Stem Cells - metabolism; Liver - embryology; Liver - metabolism; Male; Medical sciences; Mice; Mice, Transgenic; Proto-Oncogene Proteins c-kit - genetics; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - metabolism; Regulatory Sequences, Nucleic Acid - genetics; hematopoietic progenitor cells; Hematology; Stem cell; Rodentia; Transgenic animal; Hematopoietic cell; Transgene; transgenic mouse; Vertebrata; Mammalia; Mouse; Green fluorescent protein; Kit; Regulatory sequence; hematopoietic stem cells
• ISSN: 0390-6078; 1592-8721
• DOI: 10.3324/haematol.13689

1 Institute of Biomedical Technologies, National Council of Research, Pisa, Italy 2 Department of Biotechnology and Bioscience, University of Milano-Bicocca, Milan, Italy Correspondence: Maria Cristina Magli and Sergio Ottolenghi, MCM, Istituto di Tecnologie Biomediche-CNR, Via Moruzzi 1, 56124 Pisa, Italy/Dipartimento di Biotecnologie e Bioscienze, Università degli Studi Milano-Bicocca, Piazza della Scienza 2, 20126 Milano, Italy. E-mail: mariacristina.magli{at}itb.cnr.it Background: The transcriptional regulation of stem cell genes is still poorly understood. Kit, encoding the stem cell factor receptor, is a pivotal molecule for multiple types of stem/progenitor cells. We previously generated mouse lines expressing transgenic green fluorescent protein under the control of Kit promoter/first intron regulatory elements, and we demonstrated expression in hematopoietic progenitors. Design and Methods: In the present work we investigated whether the transgene is also expressed in hematopoietic stem cells of adult bone marrow and fetal liver. To this purpose, we tested, in long-term repopulating assays, cell fractions expressing different levels of green fluorescent protein within Kit-positive or SLAM-selected populations. Results: The experiments demonstrated transgene expression in both fetal and adult hematopoietic stem cells and indicated that the transgene is transcribed at distinctly lower levels in hematopoietic stem cells than in pluripotent and committed progenitors. Conclusions: These results, together with previous data, show that a limited subset of DNA sequences drives gene expression in number of stem cell types (hematopoietic stem cells, primordial germ cells, cardiac stem cells). Additionally, our results might help to further improve high level purification of hematopoietic stem cells for experimental purposes. Finally, as the Kit/green fluorescent protein transgene is expressed in multiple stem cell types, our transgenic model provides powerful in vivo system to track these cells during development and tissue regeneration. Key words: Kit, hematopoietic stem cells, hematopoietic progenitor cells, transgenic mouse.