Expression of the Rho-GEF Pbl/ECT2 is regulated by the UBE3A E3 ubiquitin ligase.
We applied genetic tools available in Drosophila to identify candidate substrates of the UBE3A ubiquitin ligase, the gene responsible for Angelman syndrome (AS). Human UBE3A was expressed in Drosophila heads to identify proteins differentially regulated in UBE3A-expressing versus wild-type extracts. Using two-dimensional gel and MALDI-TOF analysis, we detected 20 proteins that were differentially regulated by over-expression of human UBE3A in Drosophila heads. One protein responsive to UBE3A was the Rho-GEF pebble (pbl). Here, we present three lines of evidence suggesting that UBE3A regulates Pbl. First, we show genetic evidence that UBE3A and the Drosophila de-ubiquitinase fat facets (faf) exert opposing effects on Pbl function. Secondly, we find that both Pbl and ECT2, the mammalian orthologue of Pbl called epithelial cell transforming sequence 2 oncogene, physically interact with their respective ubiquitin E3 ligases. Finally, we show that Ect2 expression is regulated by Ube3a in mouse neurons as the pattern of Ect2 expression is dramatically altered in the hippocampus and cerebellum of Ube3a null mice. These results suggest that an orthologous UBE3A post-translational regulatory pathway regulates neuronal outgrowth in the mammalian brain and that dysregulation of this pathway may result in neurological phenotypes including AS and possibly other autism spectrum disorders.
Reiter
Lawrence T
LT
Section of Cell and Developmental Biology, University of California, San Diego, La Jolla 92093, USA.
Seagroves
Tiffany N
TN
Bowers
Megan
M
Bier
Ethan
E
D000595:Amino Acid Sequence; D017204:Angelman Syndrome / Q000235:genetics / Q000378:metabolism; D000818:Animals; D030801:Animals, Genetically Modified; D002531:Cerebellum / Q000378:metabolism; D004330:Drosophila / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D029721:Drosophila Proteins / Q000235:genetics / Q000378:metabolism; D005123:Eye / Q000254:growth & development; D005786:Gene Expression Regulation; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics / Q000378:metabolism; D006624:Hippocampus / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D051379:Mice; D018345:Mice, Knockout; D008969:Molecular Sequence Data; D010641:Phenotype; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D013379:Substrate Specificity; D044767:Ubiquitin-Protein Ligases / Q000172:deficiency / Q000235:genetics* / Q000378:metabolism*
D029721:Drosophila Proteins; C489850:ECT2 protein, human; C080509:Ect2 protein, mouse; D020662:Guanine Nucleotide Exchange Factors; C121865:Pbl protein, Drosophila; D011518:Proto-Oncogene Proteins; D011994:Recombinant Proteins; C471639:UBE3A protein, human; C502429:Ube3a protein, mouse; D044767:Ubiquitin-Protein Ligases