Identification of Dominant Negative Erk1/2 Variants in Cancer Cells

The Erk proteins ubiquitously expressed in various types of cells play a central role in regulating cell growth and oncogenesis. Here we report two Erk1/2 splicing variants (tErk1 and tErk2) that lack the highly conserved insertion domain (44 residues) encoded by the exon 6. RT‐PCR revealed that the tErk1/2 transcripts are present in many cancer cell lines, but not or in significantly less amount in normal tissues. Molecular modeling suggests that deletion of the insertion domain does not alter the global conformation but might reposition the signature motif Thr‐Glu‐Tyr in the activation lip that are required for phosphorylation by dual kinase MEK for Erk activation. The variants were cloned in a phCMV vector and expressed in surrogate COS‐1 and HEK293 cells. Consistent to the calculated molecular weight, Western blot detected tErk1/2 expression at 38/36 KDa. Interestingly, tErk1/2 interacted with the activating kinase MEK as detected with the biomolecular fluorescence complementation (BiFC) technology, but failed to be phosphorylated. Overexpression of tErk1/2 in COS‐1 or HEK293 cells inhibited the angiotensin II, EGF, and PMA‐induced phosphorylation of the full‐length Erk1/2 and Erk‐dependent Elk‐1, suggesting a dominant negative role. In conclusion, we have identified novel Erk variants in cancer cells. These results suggest that the truncated tErk1/2 act as dominant negative variants to antagonize the full‐length Erk1/2. This may represent a general paradigm for regulation of a protein function by its splicing variants. The work is supported by NHLBI HL65492 to YHF