Fig 2: Evaluation of the relation between the cGMP-PKG system and RAF1 expression. Retinal explants from rd1 were treated with 50 µM Rp-8-Br-PET-cGMPS (PKG inhibitor) for 2 h (n = 3), and their untreated counterparts (n = 3) or the same compound in the same concentration for 4 days (n = 4) and their untreated controls (n = 3). (A,B) represent RAF1 (green) immunostaining in untreated retinal explants and their counterparts with 2 h of PKG inhibition. (C,D) represent immunostaining of RAF1 in untreated retinal explants and their counterparts with 4 days of PKG inhibition. DAPI (blue) was used as nuclear counterstain. (E) The box chart compares RAF fluorescence intensities within the ONL between untreated explants and peers with PKG inhibitors in different treatment lengths. * p < 0.05.

Fig 3: Comparison of RAF1 phosphorylation between rd1 and wt retinas. The PLA punctuations indicate that RAF1 is in proximity with phosphoserine, which reveals the RAF1 phosphorylation profile. (A,B) represent PLA punctuations (RAF1 and phosphorylated serine) in ONL from rd1 and wt, respectively. DAPI (blue) was used as nuclear counterstain (C) The box chart shows the comparison of PLA counts in ONL of retinas from rd1 (n = 5) and wt (n = 5). **** p < 0.0001.

Fig 4: Evaluation of RAF proto-oncogene serine/threonine–protein kinase (RAF1) expression in rd1 and wt retinas. (A): Immunostaining of RAF1 (green) in a P11 rd1 retina. (B): Immunostaining of RAF1 (green) in a P11 wt retina. DAPI (blue) was used as a nuclear counterstain. (C): The box chart shows the comparison of RAF fluorescence intensities within the outer nuclear layer (ONL) between rd1 (n = 5) and wt (n = 5), *** p < 0.001.