Fig. 3.

Transient expression of PU.1 in neural stem cells is sufficient for reprogramming. (A) Over time, an increasing number of neural stem cell–derived cells transduce with lenti-PU.1-GFP lack GFP expression, but are still expressing CD45 (CD45⁺/GFP⁻) at 8 days after transduction (Fig. 1). This population of cells could have arisen in at least two ways; either transduced cells silenced the lentiviral expression of PU.1-HA-GFP but the initial expression of PU.1 was sufficient to activate an endogenous genetic monocyte program, or, alternatively, nontransduced cells were induced to express CD45 by paracrine signals from transduced cells. (B) We analyzed genomic DNA for the presence of integrated viral genetic material. The presence of the viral 5′ LTR sequence was found at similar levels in flow cytometry–isolated CD45⁺/GFP⁻ cells as in CD45⁺/GFP⁺ and CD45⁻/GFP⁺ cells. In contrast, 5′ LTR sequences were detected at levels only marginally above background in CD45⁻/GFP⁻ cells. This indicates that the CD45⁺/GFP⁻ cells were initially transduced and the new monocytic marker profile in these cells was established and stabilized, even though the viral expression of PU.1 was later silenced.