Sepsis-induced acute lung injury is associated with greatly decreased synthesis of surfactant, a stabilizing material comprised mainly of the lipid, phosphatidylcholine (PC). The production of PC is exquisitely controlled by the key surfactant enzyme, CCT. CCT is prone to phosphorylation and proteolytic degradation leading to enzyme inactivation and reduced surfactant availability. Our preliminary data show that in bacterial sepsis models, intracellular calcium triggers kinase-mediated CCT phosphorylation (Aim 1) and F-Box driven ubiquitination and lysosomal degradation of the surfactant enzyme (Aim 2) that impairs surfactant production. Calmodulin, a calcium-regulated sensor appears to protect CCT from these modifications. Thus, we hypothesize that in sepsis, lung injury is mediated, in part, by post-translational events within the CCT primary structure. We will test our hypothesis using state-of-art approaches to identify putative phosphorylation and ubiquitin acceptor sites within the CCT sequence and molecular signatures that govern interactions between F-Box, calmodulin, ubiquitin, and the CCT phosphorylation state. We will also examine the ability of calmodulin to oppose action of F-Box and kinase activity in vitro and in vivo. Our approaches include adenoviral gene transfer of novel surfactant CCT enzymes that are resistant to these post-translational modifications and calmodulin gene transfer in mice after bacterial sepsis. Our long-term goal is to devise suitable pharmaceutical inhibitors of bacterially regulated F-box proteins and kinases or calmodulin activators that might stimulate surfactant production in florid sepsis.

Public Health Relevance

Sepsis-induced acute lung injury results in decreased production of surfactant, an essential material that stabilizes lung function. We have discovered that in animal models of septic lung injury, there are two mechanisms that greatly reduce function of a key surfactant synthetic enzyme, CCT: i) CCT phosphorylation by stress kinase, and ii) CCT is degraded a F-Box E 3 ligase;these effects are opposed by calmodulin. In this application we will use several tools to confirm that stress kinase and F-Box are critical factors that reduce CCT availability and surfactant production after bacterial infection. Execution of studies outlined in this application will provide a major advance in the conceptual framework for how sepsis causes lung impairment by altering surfactant production.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL098174-04
Application #
8423737
Study Section
Surgery, Anesthesiology and Trauma Study Section (SAT)
Program Officer
Harabin, Andrea L
Project Start
2010-02-01
Project End
2015-01-31
Budget Start
2013-02-01
Budget End
2014-01-31
Support Year
4
Fiscal Year
2013
Total Cost
$321,268
Indirect Cost
$109,210
Name
University of Pittsburgh
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
004514360
City
Pittsburgh
State
PA
Country
United States
Zip Code
15213
Chen, Yan; Li, Jin; Dunn, Sarah et al. (2014) Histone deacetylase 2 (HDAC2) protein-dependent deacetylation of mortality factor 4-like 1 (MORF4L1) protein enhances its homodimerization. J Biol Chem 289:7092-8
Goetzman, Eric S; Alcorn, John F; Bharathi, Sivakama S et al. (2014) Long-chain acyl-CoA dehydrogenase deficiency as a cause of pulmonary surfactant dysfunction. J Biol Chem 289:10668-79
Zou, Chunbin; Mallampalli, Rama K (2014) Regulation of histone modifying enzymes by the ubiquitin-proteasome system. Biochim Biophys Acta 1843:694-702
Weathington, Nathaniel M; Snavely, Courtney A; Chen, Bill B et al. (2014) Glycogen synthase kinase-3? stabilizes the interleukin (IL)-22 receptor from proteasomal degradation in murine lung epithelia. J Biol Chem 289:17610-9
Chen, Bill B; Coon, Tiffany A; Glasser, Jennifer R et al. (2014) E3 ligase subunit Fbxo15 and PINK1 kinase regulate cardiolipin synthase 1 stability and mitochondrial function in pneumonia. Cell Rep 7:476-87
Han, SeungHye; Mallampalli, Rama K (2014) Sizing up surfactant synthesis. Cell Metab 20:195-6
Weathington, Nathaniel M; Mallampalli, Rama K (2014) Emerging therapies targeting the ubiquitin proteasome system in cancer. J Clin Invest 124:6-12
Liu, Yuan; Mallampalli, Rama K (2014) Decoding the growth advantage of hypoxia-sensitive lung cancer. Am J Respir Crit Care Med 190:603-5
Agassandian, Marianna; Mallampalli, Rama K (2013) Surfactant phospholipid metabolism. Biochim Biophys Acta 1831:612-25
Mallampalli, Rama K; Glasser, Jennifer R; Coon, Tiffany A et al. (2013) Calmodulin protects Aurora B on the midbody to regulate the fidelity of cytokinesis. Cell Cycle 12:663-73

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