Abstract
We have previously reported that the dynein light chain (DLC) km23‐1 is required for Smad2‐dependent TGFβ signaling. Here we describe another member of the km23/DYNLRB/LC7/robl family of DLCs, termed km23‐2, which is also involved in TGFβ signaling. We show not only that TGFβ stimulates the interaction of km23‐2 (DYNLRB2) with TGFβ receptor II (TβRII) but also that TGFβ regulates the interaction between km23‐2 and endogenous TβRII in vivo. In addition, TGFβ treatment causes km23‐2 phosphorylation, whereas a kinase‐deficient form of TβRII prevents km23‐2 phosphorylation. In contrast to the km23‐1 isoform, blockade of km23‐2 expression using small interfering RNAs (siRNAs) decreased key TGFβ/Smad3‐specific responses, including the induction of both plasminogen activator inhibitor‐1 (PAI‐1) gene expression and p21 protein expression. Blockade of km23‐1 expression had no effect on these two major TGFβ/Smad3 responses under similar conditions. Further, km23‐2 was required for TGFβ stimulation of Smad3‐dependent Smad‐binding element (SBE)2‐Luc transcriptional activity, but not for TGFβ stimulation of Smad2‐dependent activin responsive element (ARE)‐Lux transcriptional activity. In order to assess the mechanisms underlying the preferential stimulation of Smad3‐ versus Smad2‐specific TGFβ responses, immunoprecipitation (IP)/blot analyses were performed, which demonstrate that TGFβ stimulated preferential complex formation of km23‐2 with Smad3, relative to Smad2. Collectively, our findings indicate that km23‐2 is required for Smad3‐dependent TGFβ signaling. More importantly, we demonstrate that km23‐2 has functions in TGFβ signaling that are distinct from those for km23‐1. This is the first report to describe a differential requirement for unique isoforms of a specific DLC family in Smad‐specific TGFβ signaling. J. Cell. Physiol. 221: 707–715, 2009. © 2009 Wiley‐Liss, Inc.