iTRAQ proteomic identification of pVHL-dependent and -independent targets of Egln1 prolyl hydroxylase knockdown in renal carcinoma cells.

The large subunit of RNA Polymerase II, Rpb1, undergoes hydroxylation on proline 1465, which in turn triggers Ser5 hydroxylation. While Egln2 prolyl hydroxylase appears to mediate P1465 hydroxylation, Egln1 has an inhibitory activity and its knockdown stimulates constitutive hydroxylation and Ser5 phosphorylation of Rpb1, but only in cells that are VHL(+). In this study we have analyzed protein factors affected by the knockdown of Egln1 in VHL(+) and VHL(−) cells. We found that, in VHL(+) cells, several proteins were inhibited but none were induced by Egln2 knockdown. The function of several of those proteins was related to calcium metabolism and the cytoskeleton. In contrast, in VHL(−) cells Egln1 knockdown caused upregulation of several mitochondrial proteins including subunits of ATP synthase. Several of the proteins repressed in VHL(−) cells by Egln1 knockdown were involved in the function of RNA polymerase II during transcription from chromatin templates. These data suggest that the effects of Egln1 knockdown depend on the status of pVHL and can be correlated with effects on Rpb1.