Differentiated cells can be reprogrammed through the formation of heterokaryons and hybrid cells when fused with
embryonic stem (ES) cells. Here, we provide evidence that conversion of human B-lymphocytes towards a multipotent state
is initiated much more rapidly than previously thought, occurring in transient heterokaryons before nuclear fusion and cell
division. Interestingly, reprogramming of human lymphocytes by mouse ES cells elicits the expression of a human ESspecific gene profile, in which markers of human ES cells are expressed (hSSEA4, hFGF receptors and ligands), but markers
that are specific to mouse ES cells are not (e.g., Bmp4 and LIF receptor). Using genetically engineered mouse ES cells, we
demonstrate that successful reprogramming of human lymphocytes is independent of Sox2, a factor thought to be required
for induced pluripotent stem (iPS) cells. In contrast, there is a distinct requirement for Oct4 in the establishment but not the
maintenance of the reprogrammed state. Experimental heterokaryons, therefore, offer a powerful approach to trace the
contribution of individual factors to the reprogramming of human somatic cells towards a multipotent state.