Requirement of a Dynein Light Chain in TGFβ/Smad3 Signaling

We have previously reported that the dynein light chain (DLC) km23-1 is required for Smad2-dependent TGF beta signaling. Here we describe another member of the km23/DYNLRB/LC7/robl family of DLCs, termed km23-2, which is also involved in TGF beta signaling. We show not only that TGF beta stimulates the interaction of km23-2 (DYNLRB2) with TGF beta receptor II (T beta RII) but also that TGF beta regulates the interaction between km23-2 and endogenous T beta RII in vivo. In addition, TGF beta treatment causes km23-2 phosphorylation, whereas a kinase-deficient form of T beta RII prevents km23-2 phosphorylation. In contrast to the km23-1 isoform, blockade of km23-2 expression using small interfering RNAs (siRNAs) decreased key TGF beta/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 TGF beta/Smad3 responses under similar conditions. Further, km23-2 was required for TGF beta stimulation of Smad3-dependent Smad-binding element (SBE)2-Luc transcriptional activity, but not for TGF beta 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 TGF beta responses, immunoprecipitation (IP)/blot analyses were performed, which demonstrate that TGF beta stimulated preferential complex formation of km23-2 with Smad3, relative to Smad2. Collectively, our findings indicate that km23-2 is required for Smad3-dependent TGF beta signaling. More importantly, we demonstrate that km23-2 has functions in TGF beta 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 TGF beta signaling. J. Cell. Physiol. 221: 707-715, 2009. (C) 2009 Wiley-Liss, Inc.