TGF–ß Protects RPE via an Increase of Ser–59 Phosphorylated AlphaB–crystallin

Abstract

Abstract: : PURPOSE: In the retinal pigment epithelium (RPE) of the macular region the expression of alphaB–crystallin is increased compared to the periphery. The chaperon activity of αB–crystallin depends on the phosphorylation of specific serine residues (Ser–19, –45, –59). Especially the phosphorylation of Ser–59 confers to the chaperon activity and cytoprotective effects of this small heat shock protein. In the pathogenesis of early age related macular degeneration (AMD) various stressors like oxidative stress and an increased expression of transforming growth factor–beta 2 (TGF–ß2) are discussed. In the present study the influence of oxidative stress and TGF–ß2 on the expression and phosphorylation of alphaB–crystallin and its specific serine residues was investigated. METHODS: Monolayer cultures of human RPE cells were treated with TGF–ß2, or stressed by oxidant–mediated injury. Induction of alphaB–crystallin and the corresponding mRNA was assessed by Western and Northern blot analyses. The site specific phosphorylation of alphaB–crystallin was determined by antibodies recognizing the specific phosphorylated serine residues (Ser–19, –45, –59) of human alphaB–crystallin. Results: An increase of alphaB–crystallin mRNA and protein expression in RPE cells was observed after exposure to oxidative stress and TGF–ß2. Both treatments significantly increased the phosphorylation of alphaB–crystallin. Site specific phosphorylation of Ser–59 was more intense following TGF–ß treatment. The other investigated Ser residues (Ser–19 and –45) were equally induced by both stimuli. CONCLUSIONS: This is the first time to show that alphaB–crystallin is not only induced by stress factors but also by TGF–ß2 in RPE cell cultures. The increased expression and site specific phosphorylation of Ser–59 of alphaB–crystallin by TGF–ß2 might protect RPE cells against stress in the early stages of AMD.

Keywords: retinal pigment epithelium • chaperones