Using a cDNA library from the chemo-resistant Hodgkin lymphoma (HL) cell line L-1236, we identified a new splicing variant of the human transcription factor ONECUT2 (OC2). This new variant (OC2 s) contains a single CUT domain without an associated HOX domain, which characterizes the proteins of the ONECUT family. Expression analysis by quantitative Real Time PCR (qRT-PCR) showed a high expression of OC2 and OC2 s in HL cell lines as well as in normal liver tissue. In contrast, the cell lines Kasumi-1, HL-60 and MCF-7 as well as normal testis showed high expression of OC2 s only. The majority of normal tissues and cell lines expressed only low levels of OC2 and OC2 s. We observed that knockdown of OC2 lead to decreased OC2 s expression whereas transgenic over-expression of OC2 had no impact on OC2 s. Interestingly, over-expressed OC2 was located predominantly in the cytoplasm and not, as expected, in the nucleus. Gene expression analysis of cells with altered expression of OC2 or OC2 s revealed further candidates regulated by these factors.