In a recent report, Yohannes and coworkers employed 2D differential in gel electrophoresis to profile proteins that were differentially e pressed in the bladder smooth muscle of rats subjected to streptozotocin induced diabetes for different inhibitor purchase periods of time. Diabetes promotes a spectrum of pathologic changes in the urinary tract, including profound alterations in smooth muscle mass and contractility. Although not identified by 2D DIGE as differentially e pressed in e perimental diabetes, MYC, along with EGR1 and the AP 1 subunit c Fos, emerged as interconnected nodes following interro gation of differentially e pressed proteins using MetaCore software.

Similarly, in our analysis, the transcription factors JUN, MYC and EGR1 were not identified as PDGF induced proteins by quantitative proteomics analysis of primary SMC cultures, but were revealed through higher order transformation of e pression data as master regulators of PDGF stimulated transcriptional and protein changes in visceral SMC. In the present study, analysis of the gene targets for each of the master regulators identified in Figure 2 revealed a high degree of potential cross regulation, in that the promoter for each transcription factor contained putative binding sites for all other factors analyzed. Consistent with the possibility for functional interaction, a recent study revealed time dependent up regulation of transcription factor specific gene modules in an in vitro model of acute MYC activation. In response to MYC induction, genes harboring AP 1 and CREB motifs were induced first, followed by those targeted by EGR1, and concluding with putative MYC targets.

Taken together, these findings argue for a co ordinated, temporal relationship between the master regulatory nodes we identified here. Given the potential for positive feedback regulation, they may also provide an e planation for the sustained fibroproliferation evident in hollow organ remodeling. We further validated the network we have described by functional analysis of DIAPH3, which emerged as one of 22 targets that were induced at both mRNA and pro tein levels in response to PDGF. DIAPH3 is a member of the diaphanous related formin family that regulates the actin and microtubule cytoskeletons downstream of the small Rho GTPases, Rho, Rac and Cdc42, in a variety of cell types.

Although primarily studied in epithelial cells and fibroblasts, the murine ortholog of DIAPH3, mDia2, has been implicated as a regulator of smooth muscle specific gene e pression in vascular SMC. In that study, the primary activity of mDia2 and Cilengitide its homolog mDia1 was to enhance actin polymerization and thereby promote nuclear localization of the transcription factors MRTF A and MRTF B to induce e pression of genes encoding smooth muscle contractile proteins.