sentence: No additional peaks were detected when the Ste5 fragment was incubated with Pkc1 , indicating that S185 is the dominant phosphorylation target in the RING-H2 domain.To test whether Pkc1 and Ste5 directly interact in vivo , we used the M-track protein – protein proximity assay ( Zuzuarregui et al ., 2012 ; Brezovich et al ., 2015 ) designed to capture short-lived interactions .