Figure 6.

PRLs are cysteine phosphorylated in cells and tissues. A, Western blots of HeLa cell extracts show changes in PRL levels and phosphorylation in response to removal and re-addition of Mg²⁺ in the culture medium. Cells were cultured for the times shown without Mg²⁺ and after its re-addition. Total cell extracts were analyzed on SDS-PAGE gels with and without the Phos-tag reagent and blots were developed with an antibody that detects PRL1 and PRL2. Boiling prior to electrophoresis releases the phosphate from phosphocysteine. B, Western blots of mouse tissues show cysteine phosphorylation in all tissues. PRL1 and PRL2 from colon were roughly 50% phosphorylated whereas skeletal muscle PRLs were essentially 100% phosphorylated.