guanosine-triphosphate and Lipomatosis

Removal of the assembly factor eukaryotic initiation factor 6 (eIF6) is critical for late cytoplasmic maturation of 60S ribosomal subunits. In mammalian cells, the current model posits that eIF6 release is triggered following phosphorylation of Ser 235 by activated protein kinase C. In contrast, genetic studies in yeast indicate a requirement for the ortholog of the SBDS (Shwachman-Bodian-Diamond syndrome) gene that is mutated in the inherited leukemia predisposition disorder Shwachman-Diamond syndrome (SDS). Here, by isolating late cytoplasmic 60S ribosomal subunits from Sbds-deleted mice, we show that SBDS and the GTPase elongation factor-like 1 (EFL1) directly catalyze eIF6 removal in mammalian cells by a mechanism that requires GTP binding and hydrolysis by EFL1 but not phosphorylation of eIF6 Ser 235. Functional analysis of disease-associated missense variants reveals that the essential role of SBDS is to tightly couple GTP hydrolysis by EFL1 on the ribosome to eIF6 release. Furthermore, complementary NMR spectroscopic studies suggest unanticipated mechanistic parallels between this late step in 60S maturation and aspects of bacterial ribosome disassembly. Our findings establish a direct role for SBDS and EFL1 in catalyzing the translational activation of ribosomes in all eukaryotes, and define SDS as a ribosomopathy caused by uncoupling GTP hydrolysis from eIF6 release.

Topics: Animals; Bone Marrow Diseases; Catalysis; Cells, Cultured; Disease Models, Animal; Eukaryotic Initiation Factors; Exocrine Pancreatic Insufficiency; Guanosine Triphosphate; Humans; Hydrolysis; Lipomatosis; Liver; Mice; Mice, Inbred C57BL; Models, Molecular; Mutation; Peptide Initiation Factors; Phosphorylation; Protein Binding; Protein Structure, Tertiary; Proteins; Ribosome Subunits, Large, Eukaryotic; Ribosomes; Shwachman-Diamond Syndrome

Most beta-adrenergic effects are mediated by activation of the enzyme adenylate cyclase. Hormone binds to the receptor leading to an accelarated binding of GTP to the coupling protein, the N-protein, which is activated. This causes an activation of the adenylate cyclase and an increased formation of cAMP, the intracellular second messenger. The same principles hold good for other hormones coupled to adenylate cyclase. The sensitivity of the adenylate cyclase may be altered in different clinical and experimental conditions. An increased sensitivity is seen in hyperthyroidism in man and in the rat, and during starvation in rats. A decreased sensitivity is seen in hypothyroidism, in patients with pheochromocytoma, pseudohypoparathyroidism type I or multiple symmetric lipomatosis. Several reasons for the altered sensitivity have been suggested. The number of hormone receptors, the coupling between receptor and N-protein, the amount or function of the N-protein or the PDE activity may all vary in different conditions.

Topics: Adenylyl Cyclases; Adrenal Gland Neoplasms; Animals; Catecholamines; Cell Membrane; Cyclic AMP; Enzyme Activation; GTP-Binding Proteins; Guanosine Triphosphate; Humans; Lipolysis; Lipomatosis; Pheochromocytoma; Pituitary Hormones, Anterior; Pseudohypoparathyroidism; Rats; Receptors, Adrenergic, beta; Receptors, Cell Surface; Thyroid Diseases; Thyroid Hormones

Multiple symmetric lipomatosis has been proposed to be associated with impaired catecholamine-responsiveness of hypertrophic adipose tissue at the level of beta-adrenergic receptors or adenylate cyclase respectively. We have studied the regulation of the adenylate cyclase by guanine nucleotides and adrenaline in 5 subjects suffering from multiple encapsulated lipomata. In the presence of GTP (0.1 mmol/l) basal adenylate cyclase activity averaged 0.5 +/- 0.3 nmol cAMP/mg protein/10 minutes in normal adipose tissue and 1.0 +/- 0.4 nmol cAMP/mg protein/10 minutes in hypertrophic adipose tissue respectively. The synthetic GTP-analogue GMP(PNP) (0.1 mmol/l) increased non-stimulated activity by about 100% in both tissues. Adrenaline (1 mumol/l-1 mmol/l) caused a dose-dependent increase of enzymic activity in both tissues which had a maximum of 130% above basal levels in the presence of GTP and of 300% in the presence of GMP(PNP) respectively. In one of the six subjects suffering from gluteal lipomata normal adipose tissue was obtained from the gluteal as well as the abdominal region on two occasions. Maximally effective concentrations of adrenaline (1 mmol/l) induced a 3-fold increase of enzymic activity in abdominal membranes compared with about a 1.7- and 1.75-fold increase in normal and lipomatous tissue from the gluteal region. The results show that encapsulated lipomata contain a normally reactive adenylate cyclase system.

Topics: Adenylyl Cyclases; Adipose Tissue; Adult; Cyclic AMP; Epinephrine; Guanosine Triphosphate; Humans; In Vitro Techniques; Kinetics; Lipomatosis; Male; Middle Aged