Mammary Cell Cyclic AMP: Regulation of Breakdown and Influence on Protein Phosphorylation

[摘要] This thesis addresses the question of whether certain control mechanisms responsible for metabolic regulation in mammalian tissues such as liver, adipose tissue and muscle, are also functional in mammary tissue. Cyclic AMP-dependent phosphorylation is considered ubiquitous in animal tissues and has been demonstrated to be capable of regulating numerous major metabolic pathways (see section 1.). In mammary tissue, cellular metabolism is dominated overwhelmingly by lactogenesis: the biosynthesis of lipid is a principal pathway in the collection of activities comprising this function. The cyclic AMP-dependent regulation of this and other metabolic pathways of particular relevance to mammary cell function is discussed at length in section 1., highlighting the apparent necessity for control mechanisms analogous to those found in other cell types. A review of these mechanisms demonstrates the potential importance of cyclic AMP as a regulator of mammary cell metabolism and raises many interesting questions. One of the most intriguing of these is whether or not mammary cell cyclic AMP levels are subject to the same regulatory influences as have been described for tissues such as liver and adipose tissue. An obvious extension of this is whether modulation of intracellular cyclic AMP levels provokes the same metabolic response, in mammary tissue, as has been observed in these other tissues. Both of these questions have been addressed and the results are presented here. The discovery that the activity of a high affinity form of cyclic AMP-phosphodiesterase found in adipose tissue and liver can be modulated by hormones such as insulin has stimulated a great deal of interest since it provides a possible mechanism for at least some of the metabolic effects of this hormone. Mammary tissue also contains "high affinity" cyclic AMP-phosphodiesterase and its regulatory properties, particularly with respect to insulin-sensitivity, have been investigated. The results show that rat mammary high affinity cyclic AMP phosphodiesterase activity is stimulated by treatment of isolated acinus preparations with insulin but that the effect cannot be reproduced in a broken-cell system. These results are discussed with reference to similar studies in other tissues. Although intracellular cyclic AMP concentrations can be raised many-fold by treatment of mammary cells with agents such as forskolin and B-adrenergic agonists (in the presence of a phosphodiesterase inhibitor), no discernable effect on the activity of key enzymes such as acetyl-CoA carboxylase (known to be phosphorylated and inactivated by cyclic AMP-dependent protein kinase) has been observed. Consequently, the existence, in mammary tissue, of a competent combination of cyclic AMP-dependent protein kinase and endogenous substrate has been investigated. Complementary to this, a preliminary survey of the existence in mammary tissue, of three other effector-dependent phosphorylation systems (governed by calcium/calmodulin-dependent protein kinase, protein kinase C and polyamine-dependent protein kinase) has been conducted. The major finding was that rat mammary tissue does indeed contain competent kinase/endogenous substrate combinations for at least two known effectors of metabolic regulation. One of these is Ca2+/calmodulin but of greater significance to the present study, the other is cyclic AMP, for which phosphorylation was shown to display a dose-dependent relationship (in the physiological range) with at least two endogenous substrates. The molecular weights of these and other endogenous substrates for effector-dependent phosphorylation in mammary tissue are compared with substrates for similar phosphorylations already identified (in the literature) in other tissues.

[发布机构] University:University of Glasgow

[关键词] Physiology, Molecular biology