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Principal Investigator: MICHAEL S BOYCE
Organization: DUKE UNIVERSITY
Fiscal Year: 2024
Award: $329,660
Funding agency: National Institute of General Medical Sciences
Project Summary/Abstract
One third of eukaryotic proteins transit the secretory pathway for sorting to specific locations, including
the endoplasmic reticulum (ER), Golgi, plasma membrane or extracellular milieu. Since misdirected proteins
cannot function, the secretory pathway is critical for establishing and maintaining normal cell and tissue
physiology. The COPII coat protein complex, which mediates anterograde trafficking from the ER, is a key control
point for protein targeting. Indeed, mutations in COPII genes cause a range of human diseases, including cranio-
lenticulo-sutural dysplasia (CLSD), a subtype of osteogenesis imperfecta (OI), hematologic disorders and myriad
neurological defects. Detailed knowledge of COPII trafficking is required to understand its role in cell physiology
and to treat disorders in which it is disrupted. However, while the core COPII machinery is well defined, little is
known about how vertebrate cells regulate COPII activity in response to normal or pathological signals or stress.
We and others have found that several COPII proteins are modified by O-linked b-N-acetylglucosamine
(O-GlcNAc), a dynamic form of intracellular protein glycosylation. At the start of the prior project period, the
effects of O-GlcNAcylation on COPII remained almost entirely unknown. Since then, we have defined the scope
of O-GlcNAcylation in the core COPII system, identified functional effects of O-GlcNAc cycling in vesicle
trafficking, devised new quantitative glycoproteomics methods to profile O-GlcNAc changes in response to
secretory pathway stress and other stimuli, and demonstrated that particular Sec23A O-GlcNAc sites are
required for endogenous collagen trafficking in cultured human cells and in the chondrocytes of developing
zebrafish. Together, these results demonstrate that site-specific O-GlcNAcylation of COPII proteins governs
cargo trafficking in vertebrate cells and tissues. However, major unanswered questions remain, including the
mechanistic effects that O-GlcNAc exerts on COPII proteins, the upstream stimuli that modulate COPII O-
GlcNAcylation and the global landscape of O-GlcNAc signaling in the early secretory pathway. Here, we propose
to address these important questions in the next award period.
In Aim 1, we will dissect the molecular mechanisms by which O-GlcNAc cycling influences COPII vesicle
trafficking. In Aim 2, we will identify upstream stimuli that control COPII protein O-GlcNAcylation and determine
the downstream effects of this signaling in protein secretion and cell cycle progression. In Aim 3, we will use our
glycoproteomics methods to canvass proteome-wide O-GlcNAc signaling in response to COPII cargo trafficking,
and provide an integrated picture of crosstalk between O-GlcNAcylation and phosphorylation in protein secretion.
Our work will shed new light on how O-GlcNAc regulates trafficking in cells and tissues, and may reveal new
opportunities to treat diseases of COPII dysfunction by manipulating protein glycosylation.
Terms: <Address><Ascotoxin><Autoregulation><Award><Biochemical><Biological><Biological Function><Biological Process><Blood Diseases><Body Tissues><Brachydanio rerio><Brefeldin A><Brittle bone disorder><Capsid Proteins><Cell Body><Cell Communication and Signaling><Cell Cycle Progression><Cell Function><Cell Growth in Number><Cell Multiplication><Cell Physiology><Cell Process><Cell Proliferation><Cell Signaling><Cell membrane><Cells><Cellular Function><Cellular Physiology><Cellular Process><Cellular Proliferation><Cellular biology><Chemicals><Chondrocytes><Coat Proteins><Collagen><Complex><Cues><Cyanein><Cytoplasmic Membrane><D-Glucose><Danio rerio><Data><Decumbin><Defect><Development><Dextrose><Disease><Disorder><Dysfunction><Dysplasia><ETS domain protein pointed-P2><Endoplasmic Reticulum><Ergastoplasm><FIRST Award><First Independent Research Support and Transition Awards><Fragilitas Ossium><Functional disorder><Genes><Genetic Alteration><Genetic Change><Genetic defect><Glucose><Goals><Golgi><Golgi Apparatus><Golgi Complex><Hematologic Diseases><Hematological Disease><Hematological Disorder><Hematology><Hereditary><Homeostasis><Human><Human Pathology><Individual><Inherited><Intermediary Metabolism><Intracellular Communication and Signaling><Kinetics><Knowledge><Link><Location><Mammalian Cell><Mass Photometry/Spectrum Analysis><Mass Spectrometry><Mass Spectroscopy><Mass Spectrum><Mass Spectrum Analyses><Mass Spectrum Analysis><Mediating><Membrane><Metabolic><Metabolic Glycosylation><Metabolic Processes><Metabolism><Methods><Modern Man><Modification><Molecular><Mutation><Nervous System Diseases><Nervous System Disorder><Neurologic><Neurologic Disorders><Neurological><Neurological Disorders><Normal Cell><Normal Tissue><Normal tissue morphology><Nutrient><O-GlcNAc Transferase gene><O-GlcNAc transferase><O-GlcNAcase><Osteogenesis Imperfect><Osteogenesis Imperfecta><Pathologic><Pathway interactions><Phosphorylation><Physiologic><Physiological><Physiological Homeostasis><Physiology><Physiopathology><Plasma Membrane><Prevalence Study><Protein Glycosylation><Protein Phosphorylation><Protein Secretion><Protein Sortings><Protein Trafficking><Proteins><Proteome><Proteomics><Rapamune><Rapamycin><Regulation><Role><Signal Transduction><Signal Transduction Systems><Signaling><Sirolimus><Site><Skeleton><Sorting><Starvation><Stimulus><Stress><Subcellular Process><Surgical sutures><Sutures><Synergisidin><System><Tissues><Vertebrate Animals><Vertebrates><Vesicle><Viral Coat Proteins><Viral Outer Coat Protein><Work><Zebra Danio><Zebra Fish><Zebrafish><biologic><biological signal transduction><blood disorder><brittle bone disease><cell biology><developmental><dyscrasia><experiment><experimental research><experimental study><experiments><extracellular><genetic approach><genetic strategy><genome mutation><glycoproteomics><glycosylation><human disease><insight><interdisciplinary approach><membrane structure><multidisciplinary approach><neurological disease><novel><paralog><paralogous gene><pathophysiology><pathway><peptide O-GlcNAc-beta-N-acetylglucosaminidase><peptide O-linked N-acetylglucosamine-beta-N-acetylglucosaminidase><plasmalemma><pnt gene product><pointed gene product><pointed protein><protein complex><protein protein interaction><protein transport><response><skeletons><social role><sugar><trafficking><vertebrata>