Prolyl hydroxylase 2: a novel regulator of β2 -adrenoceptor internalization.
Bottom Line: However, it remains to be clarified whether or how PHDs are involved in the regulation of β(2) -adrenoceptor (β(2) -AR) signalling.Here we show that PHD2 can modulate the rate of β(2) -AR internalization through interactions with β-arrestin 2.PHD2 hydroxylates β-arrestin 2 at the proline (Pro)(176), Pro(179) and Pro(181) sites, which retards the recruitment of β-arrestin 2 to the plasma membrane and inhibits subsequent co-internalization with β(2) -AR into the cytosol. β(2) -AR internalization is critical to control the temporal and spatial aspects of β(2) -AR signalling.
Affiliation: Key Laboratory of Arrhythmias, Ministry of Education, China (East Hospital, Tongji University School of Medicine), Shanghai, China.Show MeSH
Mentions: PHD2 is an important oxygen sensor that modifies the corresponding target proteins through its hydroxylase activity . Hypoxia condition or dimethyloxalyglycine (DMOG) treatment resulted in the inhibition of PHD activity, and in turn obviously accelerated β2-AR internalization compared with the cells cultured under nomoxic conditions, suggesting that PHD activity plays a negative role in the regulation of β2-AR internalization (Fig. 4A). However, hypoxia or DMOG is a broad-spectrum PHD inhibitor, and these treatments failed to distinguish which PHD isoform activity participated in the regulation of β2-AR internalization. Next, we adopted double gene knockdown technique combined with DMOG treatment to solve this problem. As shown in Fig 4B, compared with the untreated cells (Ctrl), PHD1/PHD3 double knockdown (PHD1−/PHD3−) did not affect the rate of β2-AR internalization upon β2-AR activation, whereas subsequent DMOG treatment could inhibit PHD2 hydroxylase activity, and accelerate β2-AR internalization in response to agonist stimulation. In contrast, compared with the untreated cells, PHD1/PHD2 (PHD1−/PHD2−) or PHD2/PHD3 (PHD2−/PHD3−) double knockdown obviously accelerated β2-AR internalization, but the effect was mainly mediated by PHD2 depletion. Although subsequent DMOG treatment resulted in the inhibition of PHD1 or PHD3 hydroxylase activity, the rate of β2-AR internalization did not further change. Collectively, these results suggest that the hydroxylase activity of PHD2, but not PHD1 or PHD3, is involved in the regulation of β2-AR internalization.
Affiliation: Key Laboratory of Arrhythmias, Ministry of Education, China (East Hospital, Tongji University School of Medicine), Shanghai, China.