Retrotransposons as mutagens in the induction of growth autonomy in hematopoietic cells

Factor-independent mutants of hematopoietic cells, especially of multipotent cells, are valuable tools to identify genes that regulate stem cell proliferation and differentiation and thus may be important in leukemogenesis. Factor-independent mutants from both myeloid precursor and hematopoietic stem cell lines were isolated. The frequency of such mutants in a given cell population was one to two orders of magnitude lower for the multipotent cell line FDC-Pmix (3.6 x 10(-9)) than for the myeloid precursors, FDC-P1-M (1.7 x 10(-8)) and D35 (2.2 x 10(-7)). Analysis of these mutants revealed several mechanisms by which growth autonomy was obtained, either with or without direct contribution of growth factor gene activation. The molecular basis of spontaneous activation of the Multi-CSF (Interleukin3) gene was determined and compared to activation of the GM-CSF gene in a previous study. Multi-CSF gene activation in both precursor and stem cells was caused by the insertion of an intracisternal A particle (IAP) provirus. In two independent mutants of the D35 cell line, activation of the Multi-CSF or the GM-CSF gene was caused by almost identical IAPs with a 99% homology in the U3 and R region of the long terminal repeat. This result demonstrates that only one class of IAPs, or perhaps a single provirus, is involved in transposition and gene activation in a particular cell line. A unique example of anti-sense promotion from an IAP provirus in one Multi-CSF mutant underlines the versatility of these elements as natural insertional mutagens.