Fig. 2: | Nature Communications

Fig. 2

From: A regulatory circuit of two lncRNAs and a master regulator directs cell fate in yeast

Fig. 2

IRT2 transcription interferes with Rme1 binding, and promotes IME1 expression a IRT2, IRT1, and IME1 expression in diploid cells harboring pCUP-IRT2 and RME1-H tagged with the V5 epitope tag (FW2060) by northern blot. Cells were pre-grown in rich medium, pre-sporulation medium, before shifted to SPO and were either not treated (−Cu) or treated with copper sulfate (+Cu) after 45 min in SPO. Samples were taken at the indicated time points. Northern blot membranes were hybridized with a probe that detects both IRT1 and IRT2, and a probe that detects IME1. As a loading control, the ribosomal RNA is displayed. b Similar as a, except that binding of Rme1 to the IME1 promoter was determined by chromatin immunoprecipitation using V5-antibodies coupled to agarose beads in cells that were either not treated (−Cu), or treated with copper sulfate (+Cu). Samples were taken at the indicated time point after treatment, formaldehyde crosslinked, and chromatin extracts were prepared. Rme1-DNA complexes were isolated by immunoprecipitation, and the recovered DNA fragments were quantified by qPCR using a primer pair directed against the Rme1-binding sites in the IME1 promoter. The signals were normalized to the silent mating locus (HMR), where Rme1 does not bind. The means ± SEM of n = 4 experiments are shown. c Similar as a, except that kinetics of meiotic divisions were determined. Means ± SEM of n = 3 experiments are shown. *Indicates the time of treatment with copper sulfate