FIGURE 3.

CMT3 binding to NIA2 decreased in response to ammonium. (A) Wild-type Arabidopsis was infiltrated by Agrobacteria transformed with 35S-CMT3-FLAG₄, and then treated with or without 5 mM (NH₄)₂SO₄. CMT3-FLAG₄ expression was examined by western blotting with anti-FLAG antibody. Tubulin was used as the loading control. (B) CMT3-FLAG₄ was pulled down with an anti-FLAG antibody, and DNA was extracted. The purified DNA was analyzed by PCR using gene-specific primers, and NIA1 and NIA2 levels were examined by agarose gel electrophoresis. Numbers under lanes indicate relative intensities. DNA levels were normalized to a value of 1.00 for DNA level in the “-” (NH₄)₂SO₄ lane in each panel. (C) Genomic DNA was isolated from wild-type Arabidopsis treated with or without 5 mM (NH₄)₂SO₄ and then digested with McrBC. The remaining DNA was used for PCR amplification with specific primers for NIA1, NIA2, and CMT3. Numbers under lanes indicate relative intensities. DNA levels were normalized to a value of 1.00 for DNA level in the “-” McrBC and “-” (NH₄)₂SO₄ lane in each panel. Tubulin was used as the loading control.