The transcription factors SIX3 and VAX1 are required for suprachiasmatic nucleus circadian output and fertility in female mice

The homeodomain transcription factors sine oculis homeobox 3 (Six3) and ventral anterior homeobox 1 (Vax1) are required for brain development. Their expression in specific brain areas is maintained in adulthood, where their functions are poorly understood. To identify the roles of Six3 and Vax1 in neurons, we conditionally deleted each gene using Synapsin(cre), a promoter targeting maturing neurons, and generated Six3(syn) and Vax1(syn) mice. Six3(syn) and Vax1(syn) females, but not males, had reduced fertility, due to impairment of the luteinizing hormone (LH) surge driving ovulation. In nocturnal rodents, the LH surge requires a precise timing signal from the brain's circadian pacemaker, the suprachiasmatic nucleus (SCN), near the time of activity onset. Indeed, both Six3(syn) and Vax1(syn) females had impaired rhythmic SCN output, which was associated with weakened Period 2 molecular clock function in both Six3(syn) and Vax1(syn) mice. These impairments were associated with a reduction of the SCN neuropeptide vasoactive intestinal peptide in Vax1(syn) mice and a modest weakening of SCN timekeeping function in both Six3(syn) and Vax1(syn) mice. Changes in SCN function were associated with mistimed peak PER2::LUC expression in the SCN and pituitary in both Six3(syn) and Vax1(syn) females. Interestingly, Six3(syn) ovaries presented reduced sensitivity to LH, causing reduced ovulation during superovulation. In conclusion, we have identified novel roles of the homeodomain transcription factors SIX3 and VAX1 in neurons, where they are required for proper molecular circadian clock function, SCN rhythmic output, and female fertility.