Because of the importance to fully understand transcription regulation, several new approaches have been recently developed to profiling PDI specificity. Bulyk and colleagues developed a ds-DNA microarray that covers the entire DNA 10-mer space and applied it to determine consensus motifs of TFs (
Berger and Bulyk, 2009). Wolfe and colleagues applied the yeast/bacteria one-hybrid approach for the same purpose (
Meng and Wolfe, 2006). Although highly successful, such DNA-centered approaches require knowledge as which TFs to be studied. Because of this limitation, it becomes quite challenging when the task is to identify potential proteins that specifically recognized a DNA motif of interest, such as motifs predicted to be functional as part of the effort in the ENCODE project (The ENCODE (ENCyclopedia Of DNA Elements) Project, 2004). To solve this problem, Hu et al. (
2009) fabricated a protein microarray composed of 4191 full-length human proteins, most of which are TFs, DNA- and RNA binding proteins, chromatin-associated proteins, protein kinases, and mitochondrial proteins. They probed the microarray with 400 predicted and 60 known DNA motifs. 17718 PDIs were thus identified. Many known PDIs for TFs were successfully recovered. A large number of new PDIs for annotated and predicted TFs were also identified. Over 300 proteins that were not previously annotated as TFs were found to show sequence-specific PDIs, including RNA binding proteins, mitochondrial proteins, and protein kinases (
Hu et al., 2009;
Xie et al., 2010). The most surprising finding is from Erk2, a well studied MAP kinase. Using a series of well-designed
in vitro and
in vivo approaches, they demonstrated that this kinase had a capability of binding DNA independent of its protein kinase activity. Further results revealed it act as a transcription repressor of transcripts induced by interferon gamma signaling (
Hu et al., 2009). Again, this study proved the unprecedented unbiased global screening capability of protein microarray for novel functions of the un-characterized proteins as well as the moonlighting function of the well-studied proteins.