TWH19 (Tandem winged helix protein previously known as STK19) has been implicated in RAS signaling, yet there is intense debate regarding its expression isoforms and proposed role in NRAS S89 phosphorylation. Here, we confirmed the existence of the debated 41-kD TWH19 mRNA, but only detected minimal levels of its corresponding protein across multiple human cell lines. The two TWH19 isoforms were characterized to bind directly to all RAS isoforms in vitro and in cells, establishing TWH19 as a bona fide RAS interactor. Overexpression or depletion of TWH19 modulates extracellular signal-regulated kinase (ERK) phosphorylation and cell proliferation, confirming its functional involvement in Mitogen-Activated Protein Kinase signaling. However, using multiple approaches, we find no evidence of endogenous or TWH19-induced NRAS or KRAS phosphorylation at S89. Our data thus support a role for TWH19 in regulating RAS-ERK signaling, while indicating that such regulation occurs independently of the disputed S89 phosphorylation event.
Li et al. (Thu,) studied this question.