Document text
Principal Investigator: Phong Thanh Nguyen
Organization: UT SOUTHWESTERN MEDICAL CENTER
Fiscal Year: 2024
Award: $125,000
Funding agency: National Institute of General Medical Sciences
Project Summary
Cells use post-translational modifications (PTM) to modulate protein function by conjugating and deconjugating modifiers
to protein targets. Imbalance in these reactions leads to human disease. Conjugation of ubiquitin-fold modifier 1 (UFM1)
to protein targets (UFMylation) has been linked to many cellular processes at the endoplasmic reticulum (ER). In contrast,
much less is known about the roles of UFM1 deconjugation (de-UFMylation), the process of removing UFM1 from
UFMylated proteins. Defects in UFSP2, a de-UFMylation enzyme, were identified in patients with skeletal and
neurodevelopmental disorders. Notably, patient-derived fibroblasts harboring UFSP2 deficiency show excessive amounts
of UFMylation (hyper-UFMylation) and defects in mitochondrial respiration and nucleotide metabolism, indicating as-yet
undescribed roles of UFMylation in cellular metabolism. These findings suggest UFMylation as a novel regulator of
mitochondrial function and nucleotide metabolism. The long-term goal is to understand how UFMylation regulates cellular
metabolism. The overall objective is to understand the molecular mechanism by which UFMylation regulates mitochondrial
respiration and nucleotide metabolism, and the molecular mechanism of action of UFSP2-mediated de-UFMylation. The
central hypothesis is that UFSP2 deficiency causes (i) hyper-UFMylation of mitochondrial ribosomes (mitoribosomes) and
the electron transport chain (ETC) complex I which change protein localization and/or function, leading to decreased protein
abundance and/or loss-of-function of ETC complexes; and (ii) hyper-UFMylation of enzymes in nucleotide metabolic
pathways, leading to changes in enzyme activity and perturbation of nucleotide metabolism. The rationale is that probing
localization and measuring activity of the hyper-UFMylated mitoribosomes, the ETC Complex I and serine
hydroxymethyltransferase-2 (SHMT2), will reveal how UFMylation regulates mitochondrial respiration and nucleotide
metabolism. In addition, the structure of UFSP2 in complex with UFMylated targets will reveal the structural basis for
substrate recognition and de-UFMylation reaction. The central hypothesis will be tested by pursuing three specific aims: 1)
Determine how hyper-UFMylation of mitoribosomes and Complex I impairs mitochondrial respiration; 2) Investigate the
effect of hyper-UFMylation of SHMT2 on nucleotide metabolism; and 3) Determine the structural basis for substrate
recognition and deconjugation reaction of human UFSP2. For the first aim, protein abundance, localization and enzyme
activity of mitochondrial ribosomes and the ETC Complex I will be measured in patient-derived UFSP2-depleted versus
WT UFSP2 cells. For the second aim, isotope tracing will be used to evaluate metabolic rates of serine and nucleotide
synthesis, while the localization and enzyme activity of SHMT2 will be assessed in patient-derived UFSP2-depleted versus
WT UFSP2 cells. For the third aim, the UFSP2 in complex with its UFM1-conjugated substrate will be trapped, purified
and structure-determined. The research proposed is innovative because it will elucidate the roles of de-UFMylation and the
effects of UFMylation on cellular metabolism for the first time. The proposed research is significant because it (1)
contributes to the identity of the UFMylome, (2) reveals UFMylation as a novel regulator of mitochondrial respiration and
nucleotide metabolism, and (3) reveals the structural basis for substrate recognition and deconjugation reaction of UFSP2.
Terms: <0-11 years old><APF-1><ATP-Dependent Proteolysis Factor 1><Allothreonine Aldolase><Autoregulation><Biological><Biological Function><Biological Process><C-terminal><Cell Body><Cell Function><Cell Physiology><Cell Process><Cell Respiration><Cells><Cellular Function><Cellular Physiology><Cellular Process><Cellular Respiration><Child><Child Youth><Children (0-21)><Complex><D-Glucose><Defect><Dextrose><Disease><Disorder><Electron Transport><Endoplasmic Reticulum><Enzyme Gene><Enzymes><Ergastoplasm><Fibroblasts><Foundations><Genetic><Germ-Line Mutation><Germline Mutation><Glucose><Glycine Hydroxymethyltransferase><Goals><HMG-20><Hereditary Mutation><High Mobility Protein 20><Homeostasis><Human><Impairment><Intermediary Metabolism><Investigators><Isotope Labeling><Isotopes><L-Serine><Link><Measures><Mediating><Metabolic Diseases><Metabolic Disorder><Metabolic Pathway><Metabolic Processes><Metabolism><Mission><Mitochondria><Mitochondrial Diseases><Mitochondrial Disorders><Mitochondrial Proteins><Modern Man><Modification><Molecular><Molecular Mechanisms of Action><N-terminal><NH2-terminal><NIGMS><National Institute of General Medical Sciences><Neurodevelopmental Disability><Neurodevelopmental Disorder><Neurological Development Disorder><Nucleotide Biosynthesis><Nucleotide Synthesis><Nucleotides><Oxidative Phosphorylation><Oxidative Phosphorylation Deficiency><Oxidative Phosphorylation Pathway><Pathway interactions><Patients><Physiological Homeostasis><Post-Translational Modification Protein/Amino Acid Biochemistry><Post-Translational Modifications><Post-Translational Protein Modification><Post-Translational Protein Processing><Posttranslational Modifications><Posttranslational Protein Processing><Process><Protein Biosynthesis><Protein Modification><Proteins><Public Health><Purines/Pyrimidines/Nucleotides/Nucleic Acids Metabolism><Reaction><Reporting><Research><Research Personnel><Researchers><Respiration><Ribosomal Peptide Biosynthesis><Ribosomal Protein Biosynthesis><Ribosomal Protein Synthesis><Ribosomes><Role><Serine><Serine Aldolase><Serine Hydroxymethylase><Serine Hydroxymethyltransferase><Serine Transhydroxymethylase><Structure><Subcellular Process><Testing><Thesaurismosis><Threonine Aldolase><Time><Ubiquitilation><Ubiquitin><Ubiquitination><Ubiquitinoylation><Variant><Variation><Work><aerobic metabolism><aerobic respiration><biologic><career><electron transfer><enzyme activity><germ-line defect><germline variant><human disease><innovate><innovation><innovative><kids><loss of function><metabolic rate><metabolism disorder><metabolism measurement><metabolomics><metabonomics><mitochondrial><mitochondrial dysfunction><neurodevelopmental disease><novel><nucleotide metabolism><oxidative metabolism><pathway><protein complex><protein function><protein synthesis><respiratory><respiratory mechanism><skeletal disease><skeletal disorder><skills><social role><ubiquination><ubiquitin conjugation><youngster>