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Principal Investigator: Y. S. Prakash
Organization: MAYO CLINIC ROCHESTER
Fiscal Year: 2024
Award: $670,051
Funding agency: National Heart Lung and Blood Institute
The impact of acute airway inflammation is mediated by pro-inflammatory cytokines (e.g., TNFα),
and underlies a number of respiratory diseases. A fundamental question is why are some
individuals more susceptible than others to the negative impact of airway inflammation. We will
explore a novel homeostatic mechanism, which protects airway smooth muscle (hASM) cells from
the negative impact of inflammation-induced reactive oxygen species (ROS) formation and
protein unfolding (endoplasmic reticulum (ER) stress). We believe that a failure in this homeostatic
mechanism leads to increased ROS formation thereby exacerbating oxidative and ER stress.
Overall Hypothesis: TNFα-induced ROS formation and protein unfolding activates the
pIRE1α/XBP1s ER stress pathway in hASM, which initiates a homeostatic response directed
towards increasing mitochondrial biogenesis and mitochondrial volume density to reduce O2
consumption and ROS formation by individual mitochondrion, while still meeting the increase in
ATP demand – sharing the energetic load across mitochondria. Furthermore, reduced Mfn2
disrupts mitochondrial tethering to the ER, thereby decreasing mitochondrial Ca2+ influx and
maximum respiratory capacity of mitochondria.
Aim 1: TNFα-induced activation of pIRE1α/XBP1s ER stress pathway increases mitochondrial
volume density and reduces O2 consumption and ROS formation per mitochondrion.
In hASM cells, the downstream impact of TNFα-induced activation of the pIRE1α/XBP1s ER
stress pathway will be explored using transfection of a non-phosphorylatable IRE1α mutant
plasmid (DP-IRE1α) or an unspliceable XBP1 (uXBP1) mRNA. In addition, we will examine the
effects of siRNA knockdown of PGC1α and Mfn2 overexpression on TNFα-induced changes in
mitochondrial biogenesis, mitochondrial volume density, O2 consumption and ROS formation.
Aim 2: TNFα-induced reduction in Mfn2 disrupts mitochondrial tethering to ER, decreases
mitochondrial Ca2+ influx and reduces maximum respiratory capacity of mitochondria.
In hASM cells, we will examine the impact of DP-IRE1α or uXBP1 mRNA transfection and
siRNA Mfn2 knockdown on TNFα-induced disruption of mitochondrial/ ER tethering, decreased
mitochondrial Ca2+ influx and reduced maximum respiratory capacity of mitochondria.
Aim 3: The impact of TNFα on activation of the pIRE1α/XBP1s ER stress pathway and
downstream effects are mitigated by ROS scavenging and chemical chaperone treatment.
In hASM cells, the mitigating effects of ROS scavenging and chemical chaperone treatment
on TNFα-induced activation of the pIRE1α/XBP1s ER stress pathway will be examined.
Terms: <(TNF)-α><Active Oxygen><Acute><Affect><Airway Disease><Asthma><Awareness><Biogenesis><Bronchial Asthma><COVID crisis><COVID epidemic><COVID pandemic><COVID-19 crisis><COVID-19 epidemic><COVID-19 era><COVID-19 global health crisis><COVID-19 global pandemic><COVID-19 health crisis><COVID-19 pandemic><COVID-19 period><COVID-19 public health crisis><COVID-19 years><Cachectin><Cell Communication and Signaling><Cell Signaling><Chaperone><Chemicals><Chiro-Inositol><Consumption><Contractile Proteins><ER stress><Endoplasmic Reticulum><Enzyme Gene><Enzymes><Ergastoplasm><Experimental Designs><Failure><Generations><Goals><Human><Individual><Inflammation><Inflammatory><Inositol><Intracellular Communication and Signaling><Leiomyocyte><Macrophage-Derived TNF><Mediating><Mesoinositol><Messenger RNA><Mitochondria><Modern Man><Molecular Chaperones><Monocyte-Derived TNF><Muscle Mitochondria><Origin of Life><Oxidative Stress><Oxygen Radicals><PARK6><PARK6 gene><PARK6 protein><PINK1><PINK1 gene><PINK1 gene product><PINK1 protein><PTEN induced kinase 1><PTEN induced putative kinase 1><PTEN-induced putative kinase><Parkin><Parkin gene><Parkinson disease 6 gene><Pathway interactions><Persons><Phosphatase and tensin homolog induced kinase 1><Phosphorylation><Plasmids><Predisposition><Pro-Oxidants><Protein Phosphorylation><Proteins><RNA Splicing><Reactive Oxygen Species><Research><Respiratory Disease><Respiratory System Disease><Respiratory System Disorder><SARS-CoV-2 epidemic><SARS-CoV-2 global health crisis><SARS-CoV-2 global pandemic><SARS-CoV-2 pandemic><SARS-coronavirus-2 epidemic><SARS-coronavirus-2 pandemic><Sarcosomes><Severe Acute Respiratory Syndrome CoV 2 epidemic><Severe Acute Respiratory Syndrome CoV 2 pandemic><Severe acute respiratory syndrome coronavirus 2 epidemic><Severe acute respiratory syndrome coronavirus 2 pandemic><Short interfering RNA><Signal Transduction><Signal Transduction Systems><Signaling><Site><Small Interfering RNA><Smooth Muscle Cells><Smooth Muscle Myocytes><Smooth Muscle Tissue Cell><Splicing><Stress><Structure><Susceptibility><TNF><TNF A><TNF Alpha><TNF gene><TNF-α><TNFA><TNFα><Testing><Transfection><Tumor Necrosis Factor><Tumor Necrosis Factor-alpha><Ubiquitilation><Ubiquitination><Ubiquitinoylation><XBP1><XBP1 gene><airway epithelium inflammation><airway inflammation><airway smooth muscle><biological signal transduction><coronavirus disease 2019 crisis><coronavirus disease 2019 epidemic><coronavirus disease 2019 global health crisis><coronavirus disease 2019 global pandemic><coronavirus disease 2019 health crisis><coronavirus disease 2019 pandemic><coronavirus disease 2019 public health crisis><coronavirus disease crisis><coronavirus disease epidemic><coronavirus disease pandemic><coronavirus disease-19 global pandemic><coronavirus disease-19 pandemic><cytokine><density><endoplasmic reticulum stress><knock-down><knockdown><loss of function><mRNA><meeting><meetings><mitochondrial><mutant><novel><overexpress><overexpression><pandemic><pandemic disease><parkin protein><pathway><protein expression><protein kinase BRPK><protein kinase BRPK gene><respiratory><respiratory inflammation><respiratory smooth muscle><respiratory tract inflammation><response><serine/threonine-protein kinase PINK1><severe acute respiratory syndrome coronavirus 2 global health crisis><severe acute respiratory syndrome coronavirus 2 global pandemic><siRNA><ubiquination><ubiquitin conjugation>