Arid5a facilitates protein synthesis in RTECs. (a) HK-2 cells were treated ± IL-17 for 5 h, labeled with puromycin, and immunoblotted with anti-puromycin or β-actin Abs, representative of three independent experiments. (b) Primary mouse RTECs were treated ± IL-17 for 5 h and labeled with puromycin. Lysates were immunoblotted with anti-puromycin or β-actin Abs, representative of three independent experiments. Med, media alone. (c) Cells were transfected with Flag-Arid5a and treated ± IL-17 for 3 h. Cytoplasmic lysates were separated by sucrose gradient fractionation. Y-axis shows A₂₆₀ absorbance. Expression of Flag (Arid5a), eIF4G, eIF4E, and RPL7A was assessed in three pooled fractions per sample. Data representative of three independent experiments. (d) HK-2 cells were treated ± IL-17 for 3 h, stained with DAPI or Abs against Flag, RPL7A, or calnexin, and imaged by confocal microscopy. Size bar = 10 µm. Representative images are shown. Quantitation of nuclear Arid5a is shown below. Each value represents the percentage of colocalized volume from an individual cell (n = 218–225). Arrow denotes region of co-association of Arid5a with RPL7A. Analyzed by Student’s t test. (e) HK-2 cells were treated ± IL-17 for 1 h, and CEBPD in two pooled fractions per sample was assessed by qPCR, presented as a percent of total CEBPD from all fractions. Shading: pink denotes supernatant (Supt), green denotes 40S, 60S, and 80S fractions, and blue denotes polysomes. (f) Proposed mechanism of Arid5a function in AGN: Arid5a binds to 18S rRNA (40S ribosome subunit). Arid5a also binds to the 5S rRNA (60S), though interactions are likely weak or transient. Arid5a enhances translational efficiency of CEBPD and IL6 among other RNAs, cumulatively promoting pathology in AGN. *P < 0.05. Source data are available for this figure: SourceData F6.