THESIS
2012
xv, 228 p. : ill. (some col.) ; 30 cm
Abstract
Regulated secretion is highlighted in secretory cells such as neurons, neuroendocrine
and endocrine cells, in which dense-core granule (DCG) is responsible for the trafficking of
secretory proteins from the Golgi to plasma membrane for regulated exocytosis. Current
understanding on the biogenesis of DCG from the Golgi remains unclear; especially there is no cytoplasmic machinery reported to regulate the vesicle budding of DCG that is a process
common to different secretory tissues. Protein interacting with C kinase 1 (PICK1) and Islet
Cell Autoantigen of 69 kDa (ICA69) are BAR domain proteins which are able to bind to
and bend membrane lipids. They are primarily expressed in tissues performing secretory or
endocrine function. In brain tissues, ~75% of PICK1 forms a heterodimeric...[
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Regulated secretion is highlighted in secretory cells such as neurons, neuroendocrine
and endocrine cells, in which dense-core granule (DCG) is responsible for the trafficking of
secretory proteins from the Golgi to plasma membrane for regulated exocytosis. Current
understanding on the biogenesis of DCG from the Golgi remains unclear; especially there is no cytoplasmic machinery reported to regulate the vesicle budding of DCG that is a process
common to different secretory tissues. Protein interacting with C kinase 1 (PICK1) and Islet
Cell Autoantigen of 69 kDa (ICA69) are BAR domain proteins which are able to bind to
and bend membrane lipids. They are primarily expressed in tissues performing secretory or
endocrine function. In brain tissues, ~75% of PICK1 forms a heterodimeric complex with
ICA69. Their role in other secretory tissues remains unknown.
In the present study, I report the PICK1-ICA69 complex was an effector of small
GTPase Rab2, which is a novel machinery involved in the biogenesis of DCG from the
Golgi. Through biochemical approaches including overexpression, shRNA-mediated
knockdown or rescue experiments, the Golgi localization of the PICK1-ICA69 complex was
found to be dependent on the active GTP-bound Rab2. Using a newly established
pulse-chase assay, it was observed for the first time that endogenous Rab2 and the
PICK1-ICA69 complex were associated with the newly budding DCG from the Golgi.
Conversely, depletion or using lipid-binding defective PICK1-ICA69 complex failed to
induce DCG biogenesis. It was also found that the presence of Rab2 and the PICK1-ICA69
complex in the Golgi could prevent secretory proteins from undergoing lysosomal
degradation. In line with this, mice deficient in PICKI and ICA69 showed markedly
reduction of secretory proteins. Overall, this study provides a new mechanistic insight to a
general scenario of DCG biogenesis mediated by Rab2 and its effector the PICK1-ICA69
complex in the Golgi.
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