Regulation of Kruppel-like factor 8 by the NEDD4 E3 ubiquitin ligase

Kruppel-like factor 8 (KLF8) plays many important roles in various diseases, especially cancer. Previous studies have shown that KLF8 is regulated by ubiquitylation. The molecular mechanism underlying this posttranslational modification of KLF8, however, has not been investigated. Reported here is our identification of the neural precursor cell expressed, developmentally down-regulated 4 (NEDD4) as the E3 ubiquitin ligase for this modification. By co-immunoprecipitation and ubiquitylation assays, we determined that KLF8 interacts with NEDD4 and is ubiquitylated by NEDD4. By site-directed mutagenesis and pharmacological inhibition of MEK, we found that the ubiquitylation of KLF8 by NEDD4 depends upon the phosphorylation of KLF8 at serine 48 by ERK. Cycloheximide chase analysis, target gene promoter reporter assay and fluorescent staining indicated that NEDD4 plays a critical role in promoting the stability and transcriptional activity of KLF8 in the nucleus. Taken together, this work identified NEDD4 as a novel E3 ubiquitin ligase for KLF8 that provides insights into targeting the KLF8-NEDD4 axis to treat various types of cancer associated with overexpression of both proteins.