Figure 1.

Characterization of protein complex-binding to EUM1 within the env1 or gna3-promotor. (A) EMSA analysis with annealed and labeled oligonucleotide derived from the env1 or gna3 promotor (env1EUM1F 5′ GAT CTC TTG TCC CTT TAC TCT GTG CTC TCT CTA CCT GCC T 3′; env1EUM1R 5′ GAT CAG GCA GGT AGA GAG AGC ACA GAG TAA AGG GAC AAG A 3′; gna3mEUM1F 5′ GAT CGA CTC GTT GCT GTG CTG TGC TGT GCT GTG CTG TGC TGT 3′ gna3mEUM1R 5′ GAT CAC AGC ACA GCA CAG CAC AGC ACA GCA CAG CAA CGA GTC 3′; lower case letters indicate bases added for labeling) and 30 µg of H. jecorina cell free extracts.²⁴,²⁵ The wild-type strain QM9414 was pregrown in darkness and harvested after illumination for the time indicated. (B) Competition experiments with cell free extracts prepared as described above after 60 minutes of illumination with 10-fold, 50-fold or 100-fold excess of cold competitor. Arrows point at the light-dependent protein complex found binding to both the env1 and the gna3-promotor. Experiments were carried out with both probes on the same gel and in case of self-competitions with 120-fold excess, uncompeted protein extracts were loaded on the same gel as control and exposed to similar signal strength. All experimental procedures after harvesting the mycelia were performed in complete darkness.