NEMO (NFkappaB essential modulator) is a non-catalytic subunit of the cytokine-dependent IkappaB kinase complex that is involved in activation of the transcription factor NFkappaB. The human NEMO gene maps to Xq28 and is arranged head to head with the proximal G6PD gene. Mutations in NEMO have recently been associated with Incontinentia Pigmenti (Smahi et al., Nature 405 (2000) 466), an X-linked dominant disorder. Three alternative transcripts with different non-coding 5' exons (1a, 1b and 1c) of NEMO have been described. In order to identify regulatory elements that control alternative transcription we have established the complete genomic sequence of the murine orthologs Nemo and G6pdx. Sequence comparison suggests the presence of two alternative promoters for NEMO/Nemo. First, a CpG island is shared by both genes driving expression of the NEMO/Nemo transcripts containing exons 1b and 1c in one direction and the housekeeping gene G6PD/G6pdx in the opposite direction. In contrast to human, an additional variant of exon 1c, named 1c+, was identified in several tissues of the mouse. This larger exon utilizes an alternative donor site located 1594 bp within intron 1c. The putative second promoter for NEMO/Nemo transcripts starting with exon 1a is unidirectional, and not associated with a CpG island. Surprisingly, this promoter is located in the second intron of G6PD/G6pdx. It shows very low basal activity and may be involved in stress/time- and/or tissue-dependent expression of NEMO. To our knowledge, an overlapping gene order similar to the G6PD/NEMO complex has not been described before.