Leaf angle (LA) is a critical agronomic trait which affects grain yield through planting density in maize. Much research has been conducted in recent years to investigate the genes responsible for LA variation and a few genes were identified through map-based cloning. Here we cloned the ZmIBH1-1 gene, which is a bHLH transcription factor with both a basic binding region and a Helix-Loop-Helix domain; and qRT-PCR results showed that ZmIBH1-1 is a negative regulator of LA in maize. Histological analysis showed that the change in LA was mainly caused by differential cell wall lignification and cell elongation in the ligular region. To reveal the regulatory framework of ZmIBH1-1, we conducted RNA-Seq and DAP-Seq analysis. Overlay of the RNA-Seq and DAP-Seq results revealed 59 ZmIBH1-1 modulated target genes with annotation, and they were mainly cell wall related, cell development or hormone related genes. We have built a new regulatory model of ZmIBH1-1 gene controlling plant architecture in maize.

Supplementary figure1. Sequence alignment of Yu82 and Yu87-1. Polymorphisms are highlighted in cyan and the predicted translation start of ZmIBH1-1 is boxed in yellow and the predicted termination codon of ZmIBH1-1 is boxed in red.

Supplementary figure 2. Protein sequence alignment of ZmIBH1-1 and SvIBH1-1. Red squares indicate the conserved bHLH domain. '*' indicates positions which have a single, fully conserved residue. ':' indicates that one of the following 'strong' groups is fully conserved: STA, NEQK, NHQK, NDEQ, QHRK, MILV, MILF, HY, and FYW. '.' indicates that one of the following 'weaker' groups is fully conserved: CSA, ATV, SAG, STNK, STPA, SGND, SNDEQK, NDEQHK, NEQHRK, FVLIM, and HFY.

Supplementary figure 3. Results of the dual-luciferase transient transcriptional activity assay. (A) The 35S:REN-Pro zein:LUC reporter constructs were transiently expressed in onion epidermal cells together with control vector or 35S:ZmIBH1-1effector, respectively. (B) The expression level of Renilla (REN) was used as an internal control. The LUC/REN ratio represents the relative activity of candidate gene promoters. Data are values of three independent experiments. Significant differences from the corresponding control values (using Student’s t test [n = 3]): *P < 0.05, **P < 0.01, and ***P < 0.001.