Lipopolysaccharide triggers nuclear import of Lpcat1 to regulate inducible gene expression in lung epithelia

AIM:To report that Lpcat1 plays an important role in regulating lipopolysaccharide (LPS) inducible gene tran-scription. METHODS:Gene expression in Murine Lung Epithelial MLE-12 cells with LPS treatment or Haemophilus influenza and Escherichia coli infection was analyzed by employing quantitative Reverse Transcription Polymerase Chain Reaction techniques. Nucleofection was used to deliver Lenti-viral system to express or knock down Lpcat1 in MLE cells. Subcellular protein fractionation and Western blotting were utilized to study Lpcat1 nuclear relocation. RESULTS:Lpcat1 translocates into the nucleus from thecytoplasm in murine lung epithelia (MLE) after LPS treatment. Haemophilus influenza and Escherichia coli , two LPS-containing pathogens that cause pneumonia, triggered Lpcat1 nuclear translocation from the cytoplasm. The LPS inducible gene expression profile was determined by quantitative reverse transcription polymerase chain reaction after silencing Lpcat1 or overexpression of the enzyme in MLE cells. We detected that 17 out of a total 38 screened genes were upregulated, 14 genes were suppressed, and 7 genes remained unchanged in LPS treated cells in comparison to controls. Knockdown of Lpcat1 by shRNA dramatically changed the spectrum of the LPS inducible gene transcription, as 18 genes out of 38 genes were upregulated, of which 20 genes were suppressed or unchanged. Notably, in Lpcat1 overex-pressed cells, 25 genes out of 38 genes were reduced in the setting of LPS treatment.CONCLUSION:These observations suggest that Lpcat1 relocates into the nucleus in response to bacterial infection to differentially regulate gene transcriptional repression.

The role of ubiquitination and deubiquitination in the regulation of cell junctions

Maintenance of cell junctions plays a crucial role in the regulation of cellular functions including cell proliferation, permeability, and cell death. Disruption of cell junctions is implicated in a variety of human disorders, such as inflammatory diseases and cancers. Understanding molecular regulation of cell junctions is important for development of therapeutic strategies for intervention of human diseases. Ubiquitination is an important type of post-translational modification that primarily regulates endogenous protein stability, recep- tor internalization, enzyme activity, and protein-protein interactions. Ubiquitination is tightly regulated by ubiq- uitin E3 ligases and can be reversed by deubiquitinating enzymes. Recent studies have been focusing on inves- tigating the effect of protein stability in the regulation of cell-cell junctions. Ubiquitination and degradation of cadherins, claudins, and their interacting proteins are implicated in epithelial and endothelial barrier disruption. Recent studies have revealed that ubiquitination is involved in regulation of Rho GTPases＇ biological activities. Taken together these studies, ubiquitination plays a critical role in modulating cell junctions and motility. In this review, we will discuss the effects of ubiquitination and deubiquitination on protein stability and expression of key proteins in the cell-cell junctions, including junction proteins, their interacting proteins, and small Rho GTPases. We provide an overview of protein stability in modulation of epithelial and endothelial barrier integrity and introduce potential future search directions to better understand the effects of ubiquitination on human disorders caused by dysfunction of cell junctions.

Regulation of inflammasomes by ubiquitination

Inflammasomes are multi-protein complexes that regulate the innate immune response by facilitating the release of inflammatory cytokines in response to pathogen exposure or cellular damage. Pro-inflammatory inflammasome signaling is vital to host defense and helps initiate the process of tissue repair following an insult to the host, but can be injurious, when excessive or chronic. As such, inflammasome activity is tightly regulated. Here we discuss one critical mechanism of inflammasome regulation, ubiquitination, that functions as a universal modulator of protein stability and trafficking. Recent studies have provided important insights into the regulation of inflammasome activation by protein ubiquitination. We review the molecular regulation of inflammasome function, specifically, as it relates to ubiquitination, and discuss the implications for the development of theraoeutics to soecificallv target aberrant inflammasome signaling.

Phosphorylated E2F1 ts stabilized by nuclear USPll to drive PeglO gene expression and activate lung epithelial cells

Phosphorylation affects ubiquitination, stability, and activity of transcriptional factors, thus regulating various cellular functions. E2F transcriptional factor I （E2F1） regulates paternally expressed imprinted gene 10 （Peg10） expression, thereby promoting cell proliferation. However, the effect of E2FZ stability on PeglO expression and the molecular regulation of E2FZ stability by its phos- phorylation have not been well demonstrated. Here, we describe a new pathway in which phosphorylation of E2F1 by GSK3p increases E2FZ association with the deubiquitinating enzyme, ubiquitin-specific protease 11 （USPll）, which removes K63-1inked ubiquitin chains thereby preventing E2FZ degradation in the nuclei. Downregulation of USPlZ increases E2FZ ubiquitination and reduces E2F1 stability and protein levels, thereby decreasing PeglO mRNA levels. Physiologically, USPll depletion suppresses cell proliferation and wound healing in lung epithelial cells, and these effects are reversed by E2F1 and PEGIO overexpression. Thus, our study reveals a new molecular model that phosphorylation promotes substrate stability through increasing its associ- ation with a deubiquitinating enzyme. The data suggest that GSK3p and USPll act in concert to modulate E2FZ abundance and PEGIO expression in lung epithelial celts to affect cell wound healing. This study provides new therapeutic targets to lessen lung injury by improving lung epithelial cell repair and remodeling after injury.