. albicans Cek1 pathway. Msb2 is the head receptor protein that feeds into the signaling cascade of the S. cerevisiae Kss1 MAPK pathway, and is structurally very 
similar to C. albicans Msb2. Msb2 has since been identified as the head sensor protein of the Cek1 MAPK pathway and is needed for the successful execution of most Cek1 mediated functions in C. albicans, such as invasive growth and cell wall biosynthesis. However, the mechanism by which Msb2 transmits signals to the Cek1 pathway is not known. Msb2 belongs to a class of glycoproteins called signaling mucins. These mucins are characterized by a highly glycosylated extracellular domain containing a Ser/Thr/Pro-rich mucin homology domain , one transmembrane region, and a cytoplasmic domain that connects to and regulates cytosolic signaling molecules. Signaling mucins are processed within their extracellular domains, which results in the release of the external glycodomain from the cells. For S. cerevisiae Msb2, processing is required for MAPK activation. The 19380825 aspartic protease Yps1 was shown to proteolytically cleave ScMsb2, and in the absence of Yps1, the Kss1 MAPK pathway activation was inhibited, underscoring the functional importance of this cleavage in the signaling process. Mass spectroscopy analysis of the C. albicans secretome identified a soluble form of Msb2 with peptide fragments originating from within the cleavage domain, suggesting post-translational processing of Msb2. Moreover, a recent study showed that the heavily glycosylated extracellular domain of Msb2 is shed into the medium and that Cek1 phosphorylation is absent in cells harboring truncated versions of Msb2. But 2 Sap Mediated Processing of C. albicans Msb2 whether C. albicans Msb2 regulates Cek1 signaling in a cleavagedependent fashion is not well understood. Here we show that the extracellular domain of Msb2 is shed in response to specific environmental cues, and that this shedding is tightly linked to Cek1 phosphorylation. Furthermore, we identify a role for the secreted aspartic protease family of proteins in the proteolytic processing and cleavage of Msb2. We also present evidence that processing of Msb2 is required for facilitation of various Cek1 signaling-mediated virulence traits, such as biofilm formation and exposure of cell surface immunogens. The results presented here allow us to postulate that Msb2 is required for C. albicans virulence because its proteolytic processing is necessary for the activation of the Cek1 MAPK pathway. possibility of septation defects which may account for the slightly delayed growth rate of msb2D/D cells. However, msb2D/D cells readily formed true hyphae, and the extent of germination did not differ between msb2D/D and wild type CAI4 8250835 cells. Msb2 is shed from cells in response to environmental Astragalus polysaccharide site conditions and shedding correlates with Cek1 phosphorylation and germination Msb2 is known to activate the Cek1 pathway in response to inhibition of glycosylation and cell wall stress. We examined the possibility that Msb2 has a broader role in response to environmental cues. To confirm that msb2D/D cells could not activate the Cek1 pathway, we compared Cek1 phosphorylation in cells lacking Msb2 with WT cells. As expected, msb2D/D cells did not show phosphorylation of Cek1, while activation of a related MAPK pathway, the cell wall integrity pathway remained intact. Also, the Msb2-HA strain containing a single HA tagged Msb2 allele was fully functional in terms of its ability to pho
