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The Role of Ubiquitination and the E3 Ligase Nedd4 in Regulating Corneal Epithelial Wound Healing.
- Source :
-
Investigative ophthalmology & visual science [Invest Ophthalmol Vis Sci] 2024 Jun 03; Vol. 65 (6), pp. 29. - Publication Year :
- 2024
-
Abstract
- Purpose: Ubiquitination serves as a fundamental post-translational modification in numerous cellular events. Yet, its role in regulating corneal epithelial wound healing (CEWH) remains elusive. This study endeavored to determine the function and mechanism of ubiquitination in CEWH.<br />Methods: Western blot and immunoprecipitation were used to discern ubiquitination alterations during CEWH in mice. Interventions, including neuronally expressed developmentally downregulated 4 (Nedd4) siRNA and proteasome/lysosome inhibitor, assessed their impact on CEWH. In vitro analyses, such as the scratch wound assay, MTS assay, and EdU staining, were conducted to gauge cell migration and proliferation in human corneal epithelial cells (HCECs). Moreover, transfection of miR-30/200 coupled with a luciferase activity assay ascertained their regulatory mechanism on Nedd4.<br />Results: Global ubiquitination levels were markedly increased during the mouse CEWH. Importantly, the application of either proteasomal or lysosomal inhibitors notably impeded the healing process both in vivo and in vitro. Furthermore, Nedd4 was identified as an essential E3 ligase for CEWH. Nedd4 expression was significantly upregulated during CEWH. In vivo studies revealed that downregulation of Nedd4 substantially delayed CEWH, whereas further investigations underscored its role in regulating cell proliferation and migration, through the Stat3 pathway by targeting phosphatase and tensin homolog (PTEN). Notably, our findings pinpointed miR-30/200 family members as direct regulators of Nedd4.<br />Conclusions: Ubiquitination holds pivotal significance in orchestrating CEWH. The critical E3 ligase Nedd4, under the regulatory purview of miR-30 and miR-200, facilitates CEWH through PTEN-mediated Stat3 signaling. This revelation sheds light on a prospective therapeutic target within the realm of CEWH.
- Subjects :
- Animals
Mice
Humans
Mice, Inbred C57BL
Endosomal Sorting Complexes Required for Transport metabolism
Blotting, Western
STAT3 Transcription Factor metabolism
Cells, Cultured
Disease Models, Animal
MicroRNAs genetics
Immunoprecipitation
Male
Gene Expression Regulation physiology
Nedd4 Ubiquitin Protein Ligases metabolism
Nedd4 Ubiquitin Protein Ligases genetics
Ubiquitination
Cell Movement physiology
Cell Proliferation physiology
Wound Healing physiology
PTEN Phosphohydrolase metabolism
PTEN Phosphohydrolase genetics
Epithelium, Corneal metabolism
Ubiquitin-Protein Ligases metabolism
Ubiquitin-Protein Ligases genetics
Subjects
Details
- Language :
- English
- ISSN :
- 1552-5783
- Volume :
- 65
- Issue :
- 6
- Database :
- MEDLINE
- Journal :
- Investigative ophthalmology & visual science
- Publication Type :
- Academic Journal
- Accession number :
- 38888282
- Full Text :
- https://doi.org/10.1167/iovs.65.6.29