As the MarR-type proteins can act as positive, negative, or bifunctional regulators, TcaR also acts as a multi-functional regulator. It is not only as a regulatory factor to affect the transcription of icaADBC [4] , the first regulator reported for cell wall-anchored proteins (SpA and sasF), but also as the regulator of sarS [18, 19] . We previously described the 3D structures of TcaR in its apo form and in complex with salicylate as well as several aminoglycoside and b-lactam antibiotics [17] . In this research, comparison of the native TcaR structures and those in complexes indicated that the regulation mechanism involves a large conformational shift in the DNA binding lobe. Several antimicrobial compounds inhibited TcaR-DNA interaction and further induced biofilm formation in S. epidermidis. In the present study, we found that TcaR could interact with ssDNA and the result demonstrated that TcaR shows a stronger preference toward GCrich ssDNA than to dsDNA by using EMSA, CD, and Biacore experiments. However, the detailed mechanism of the interaction between TcaR and ssDNA still remains to be elucidated. In order to investigate the regulation mechanism of the ssDNA binding ability of TcaR, we applied electron microscopy (EM) technique to reveal TcaR-ssDNA complex. Furthermore, we clarified the role of TcaR-ssDNA interaction by in vitro replication assay and in vivo plaque assay. Taken together, these results provide an in-depth investigation on the multiple functions of TcaR in S. epidermidis.