Abstract
The dynamin family of GTP-binding proteins has been implicated as playing an important role in endocytosis. In Drosophila shibire, mutations of the single dynamin gene cause blockade of endocytosis and neurotransmitter release, manifest as temperature-sensitive neuromuscular paralysis. Mammals express three dynamin genes: the neural specific dynamin I, ubiquitous dynamin II, and predominantly testicular dynamin III. Mutations of dynamin I result in a blockade of synaptic vesicle recycling and receptor-mediated endocytosis. Here, we show that dynamin II plays a key role in controlling constitutive and regulated hormone secretion from mouse pituitary corticotrope (AtT20) cells. Dynamin II is preferentially localized to the Golgi apparatus where it interacts with G-protein betagamma subunit and regulates secretory vesicle release. The presence of dynamin II at the Golgi apparatus and its interaction with the betagamma subunit are mediated by the pleckstrin homology domain of the GTPase. Overexpression of the pleckstrin homology domain, or a dynamin II mutant lacking the C-terminal SH3-binding domain, induces translocation of endogenous dynamin II from the Golgi apparatus to the plasma membrane and transformation of dynamin II from activity in the secretory pathway to receptor-mediated endocytosis. Thus, dynamin II regulates secretory vesicle formation from the Golgi apparatus and hormone release from mammalian neuroendocrine cells.
Highlights
Dynamin is a polypeptide with a modular structure comprising a GTP-binding domain in the N-terminal third, a middle domain of unknown function, a pleckstrin homology (PH)1 domain and a C-terminal proline-rich or Src homology 3- (SH3-) binding domain
To determine whether the effects of dynamin II on hormone secretion might have resulted from nonspecific changes in cellular proliferation or signaling, basal DNA synthesis and mitogenactivated protein (MAP) kinase activity in response to epidermal growth factor and the phorbol ester, phorbol 12-myristate 13acetate were measured in the different transformants
Given that changes in hormone release induced by dynamin II variants are accompanied by no change in cellular content and increased total synthesis of the hormone, it is likely that dynamin II may be involved in regulating secretory vesicle transport with the coupling between hormone secretion and synthesis not being affected; enhanced effects on secretion may lead to enhanced levels of synthesis to replenish the released stores of -endorphin
Summary
Dynamin is a polypeptide with a modular structure comprising a GTP-binding domain in the N-terminal third, a middle domain of unknown function, a pleckstrin homology (PH) domain and a C-terminal proline-rich or Src homology 3- (SH3-) binding domain (for reviews, see Refs. 1– 8). Since dynamin I complexes a large ring-like structure surrounding the necks of clathrin-coated endocytic pits on the cytoplasmic surface of presynaptic plasma membrane during synaptic vesicle recycling [3, 6], it is thought that dynamin II plays a similar role to that of dynamin I in receptor-mediated endocytosis in nonneuronal cells (4 – 8) In support of this hypothesis are the findings that the GTPase activities of both dynamin I and II, and of their SH3-binding domain truncation mutants, are stimulated in vitro by phospholipids, grb, and microtubules [9], suggesting that the modes of interaction of the PH domains from dynamin I and II with phosphatidylinositol [4,5]bisphosphate and of the SH3-binding domain with microtubules and grb are similar in vitro [9]. We found that dynamin II plays a key role in controlling both constitutive and regulated hormone secretions at the Golgi apparatus of these neuroendocrine cells
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