Adipocyte phosphatases and the antilipolytic action of insulin

Detta är en avhandling från Svante Resjö, BMC, C11, SE-221 84 LUND, SWEDEN

Sammanfattning: Adipose tissue is the main site of energy storage of the body and an important endocrine organ. Knowledge of the regulation of fat metabolism and the endocrine factors secreted by the adipocyte is crucial for the understanding of diseases such as obesity and diabetes. Insulin is the main antilipolytic hormone. Protein kinase B (PKB) and phosphodiesterase 3B (PDE3B) are both phosphorylated and activated in adipocytes in response to insulin. PDE3B is known to be a key intermediate in insulin's antilipolytic signal transduction pathway and PKB is believed to be the kinase responsible for phosphorylating and activating PDE3B in response to insulin. This thesis deals with the reversible phosphorylation of PKB and PDE3B in adipocytes and with the purification of membrane associated protein phosphatase 2A (PP2A) from adipose tissue. Using treatment of rat adipocytes with phosphatase inhibitors and in vitro dephosphorylations of PKB and PDE3B with partially purified adipocyte phosphatases, we were able to demonstrate that PP2A has an important role in the dephosphorylation of both PKB and PDE3B. In addition we were able to show that in adipocytes PKB is mainly phosphorylated on a single site, Ser 473 (PKB-alpha sequence), in contrast to previously published reports using cultured cells overexpressing PKB. Finally, since both PKB and PDE3B are membrane associated enzymes we managed to purify a membrane associated form of PP2A from bovine adipose tissue. The PP2A form was identified as being a trimer composed of PP2Ac-beta, PR65-alpha and PR55-alpha. These results provide information that will make it possible to study the dephosphorylation of PKB and PDE3B in greater detail with the possible goal of finding ways to specifically modify the phosphorylation states of these enzymes.

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