Figure 4: TEM regulates GA biosynthetic genes.

(a,b) Expression analysis of GA metabolism genes in wild-type and 35S::TEM1 seedlings, and in wild-type, tem1-1, tem2-2 and tem1-1 tem2-2 mutants. After 1 week under SD, samples were collected at ZT8 and subjected to RT–qPCR. (c,d) GUS staining of pGA3OX1-GUS-TL and pGA3OX1-GUS-TL 35S::TEM1 (F3) grown for 10 days under SD (SAM close-up in the inset) and for 3 weeks under LD. Bar represents 1 cm. (e) ChIP analysis of TEM1 binding to GA3OX1 regulatory regions. Precipitated chromatin was used as template in qPCR with primer sets AB, amplifying a region 2.4 kb upstream of the ATG, and CD, amplifying the region of the first exon containing the RAV-binding site. (f) ChIP analysis of TEM1 binding to GA3OX2 regulatory regions. Precipitated chromatin was used as template in qPCR with primer sets GH, amplifying a region 2.7 kb upstream of the ATG, and CD, amplifying the region of the first exon containing a non-canonical RAV-binding site. For expression and ChIP analyses, three biological replicates were performed with similar results. One representative is shown with error bars of three qPCR replicates; other biological replicates are shown in Supplementary Fig. S6b. Wt, wild type.