Using a newly devised Ubc9fusion directed sumoylation approach, we examined the functional consequences of specific enhancement of Tec1 sumoylation and found that sumoylation has an inhibitory role on Tec1 activity. Together these findings revealed a previously unknown mechanism by which Kss1 controls the transcriptional activity of Tec1 and validated UFDS as a useful approach for Soyosaponin-Aa studying protein sumoylation. What could be the mechanism by which Kss1 regulates the sumoylation level of Tec1? One possibility is that Kss1 can directly phosphorylate Tec1 and phosphorylation targets Tec1 for desumoylation. It has been demonstrated previously that Tec1 can be phosphorylated by Fus3, the MAP kinase of the pheromone signaling pathway, but whether Tec1 is also a substrate of Kss1 remains to be determined. Another possibility is that Kss1 might have a regulatory role for the machinery that controls Tec1 sumoylation in vivo. It is not without precedent that a MAP kinase can regulate the activity of enzymes critical for ubiquitin and ubiquitin-like modifications. For instance, it has been reported previously that JNK can regulate the activity of an E3 ubiquitin ligase Itch. The possibility that Kss1 may inhibit the main components of the sumoylation pathway such as Ubc9 is unlikely, however. Under the same condition that we detected stimulusdependent decrease of Tec1 sumoylation, the global level of protein sumoylation is increased, indicating that Kss1 does not have a general role of inhibiting protein sumoylation. One often utilized strategy for studying the function of protein sumoylation is Glycitein determining the consequences of diminishing or enhancing the sumoylation level of the protein. Identifying and mutating the acceptor lysine residues is one of the commonly used approaches for blocking sumoylation. However, certain limitations are associated with this approach. For instance, a number of modifications such as ubiquitination, acetylation and methylation can occur on lysine residue. Therefore it is not necessarily appropriate to attribute the phenotype of a sumoylation site mutant solely to a change in sumoylation.