The regulatory and basal phosphorylation sites of hormone‐sensitive lipase are dephosphorylated by protein phosphatase‐1, 2A and 2C but not by protein phosphatase‐2B

Abstract

The activity of hormone‐sensitive lipase, the rate‐limiting enzyme in adipose tissue lipolysis, is controlled by cAMP‐mediated phosphorylation at a specific regulatory phosphorylation site. The lipase is also phosphorylated at a site, termed basal, without any effects on its activity [Strålfors et al. (1984) Proc. Natl Acad. Sci. USA 81, 3317–3321]. The capacity of protein phosphatase‐1, 2A, 2B and 2C to dephosphorylate the lipase, selectively phosphorylated by glycogen synthase kinase‐4 and cAMP‐dependent protein kinase at the basal and regulatory phosphorylation sites, was compared with that towards glycogen phosphorylase and phosphorylase kinase (α subunit). Protein phosphatase‐1, 2A and 2C were found to dephosphorylate both phosphorylation sites of hormone‐sensitive lipase, while protein phosphatase‐2B had no measureable activity towards any of the sites. When the activities of protein phosphatase‐1, 2A and 2C were normalized with respect to the reference substrates, they were found to dephosphorylate the lipase regulatory site in the approximate relations of 1:4:3 and the basal site in the approximate relations of 1:6:4. Protein phosphatase‐1 showed 20% higher and protein phosphatase‐2A and 2C 80% higher activity towards the basal site compared to the regulatory site. The two phosphorylation sites of the lipase were comparable to good substrates for protein phosphatase‐2A and 2C, but relatively poor substrates for protein phosphatase‐1. Protein phosphatase‐2C activity towards the lipase was completely dependent on Mg²⁺ with a half‐maximal effect at 3 mM. Protamine increased the lipase dephosphorylation by protein phosphatase‐1 3–5‐fold with half‐maximal effect at 0.6 μg/ml, and by protein phosphatase‐2A about 2‐fold with half‐maximal effect at 3–5 μg/ml, thus illustrating the potential for control of these lipase phosphatase activities.