bSignaling mucins are evolutionarily conserved regulators of signal transduction pathways. The signaling mucin Msb2p regulates the Cdc42p-dependent mitogen-activated protein kinase (MAPK) pathway that controls filamentous growth in yeast. The cleavage and release of the glycosylated inhibitory domain of Msb2p is required for MAPK activation. We show here that proteolytic processing of Msb2p was induced by underglycosylation of its extracellular domain. Cleavage of underglycosylated Msb2p required the unfolded protein response (UPR), a quality control (QC) pathway that operates in the endoplasmic reticulum (ER). The UPR regulator Ire1p, which detects misfolded/underglycosylated proteins in the ER, controlled Msb2p cleavage by regulating transcriptional induction of Yps1p, the major protease that processes Msb2p. Accordingly, the UPR was required for differentiation to the filamentous cell type. Cleavage of Msb2p occurred in conditional trafficking mutants that trap secretory cargo in the endomembrane system. Processed Msb2p was delivered to the plasma membrane, and its turnover by the ubiquitin ligase Rsp5p and ESCRT attenuated the filamentous-growth pathway. We speculate that the QC pathways broadly regulate signaling glycoproteins and their cognate pathways by recognizing altered glycosylation patterns that can occur in response to extrinsic cues. Signaling mucins are evolutionarily conserved regulators of signal transduction pathways (1-4). Signaling mucins are composed of a highly glycosylated extracellular domain that contains a mucin homology domain (MHD), which is defined by tandem repeats rich in Ser/Thr/Pro residues. The extracellular domain is connected by a single-pass transmembrane (TM) alpha helix to a cytosolic signaling domain, which associates with a diverse array of proteins that regulate mitogen-activated protein kinase (MAPK) pathways, Akt, -catenin, and other pathways (5-8). Signaling mucins are overexpressed in different cancers, where they contribute to cell proliferation and metastasis (6). They are diagnostic biomarkers for cancers (9) and targets for immunotherapies (10,11). Therefore, the mechanisms by which signaling mucins and related glycoproteins are regulated is of intense interest.In the budding yeast Saccharomyces cerevisiae, the mucin-like glycoprotein Msb2p regulates the MAPK pathway that controls filamentous growth, a cell differentiation behavior that occurs in response to nutrient limitation (12-14). The extracellular domain of Msb2p is extensively glycosylated. Msb2p is modified by Nlinked and O-linked glycosylation and contains a canonical MHD that is itself highly glycosylated (15, 16). In a landmark study, Yang et al. identified Pmt4p as the major O-mannosyltransferase for Msb2p (17). Pmt4p is a member of an evolutionarily conserved protein mannosyl transferase (Pmt) gene family (2, 18). Msb2p also contains a cytosolic signaling domain. The cytosolic domain of Msb2p associates with the Rho GTPase Cdc42p (15), which is a ubiquitous regulator of cell polarity and signaling...
The ubiquitous Rho (Ras homology) GTPase Cdc42p can function in different settings to regulate cell polarity and cellular signaling. How Cdc42p and other proteins are directed to function in a particular context remains unclear. We show that the Cdc42p-interacting protein Bem4p regulates the mitogen-activated protein kinase (MAPK) pathway that controls filamentous growth in Saccharomyces cerevisiae. Bem4p controlled the filamentous-growth pathway but not other MAPK pathways (mating or high-osmolarity glycerol response [HOG]) that also require Cdc42p and other shared components. Bem4p associated with the plasma membrane (PM) protein, Sho1p, to regulate MAPK activity and cell polarization under nutrient-limiting conditions that favor filamentous growth. Bem4p also interacted with the major activator of Cdc42p, the guanine nucleotide exchange factor (GEF) Cdc24p, which we show also regulates the filamentous-growth pathway. Bem4p interacted with the pleckstrin homology (PH) domain of Cdc24p, which functions in an autoinhibitory capacity, and was required, along with other pathway regulators, to maintain Cdc24p at polarized sites during filamentous growth. Bem4p also interacted with the MAPK kinase kinase (MAPKKK) Ste11p. Thus, Bem4p is a new regulator of the filamentous-growth MAPK pathway and binds to general proteins, like Cdc42p and Ste11p, to promote a pathway-specific response.
A fundamental problem in cell biology is to understand how spatial information is recognized and integrated into morphogenetic responses. Budding yeast undergoes differentiation to filamentous growth, which involves changes in cell polarity through mechanisms that remain obscure. Here we define a regulatory input where spatial landmarks (bud-site–selection proteins) regulate the MAPK pathway that controls filamentous growth (fMAPK pathway). The bud-site GTPase Rsr1p regulated the fMAPK pathway through Cdc24p, the guanine nucleotide exchange factor for the polarity establishment GTPase Cdc42p. Positional landmarks that direct Rsr1p to bud sites conditionally regulated the fMAPK pathway, corresponding to their roles in regulating bud-site selection. Therefore, cell differentiation is achieved in part by the reorganization of polarity at bud sites. In line with this conclusion, dynamic changes in budding pattern during filamentous growth induced corresponding changes in fMAPK activity. Intrinsic compromise of bud-site selection also impacted fMAPK activity. Therefore, a surveillance mechanism monitors spatial position in response to extrinsic and intrinsic stress and modulates the response through a differentiation MAPK pathway.
Clinical studies combining radiation and immunotherapy have shown promising response rates, strengthening efforts to sensitize tumors to immune-mediated attack. Thus, there is an ongoing surge in trials using preconditioning regimens with immunotherapy. Yet, due to the scarcity of resected tumors treated in situ with radiotherapy, there has been little investigation of radiation’s sole contributions to local and systemic antitumor immunity in patients. Without this access, translational studies have been limited to evaluating circulating immune subsets and systemic remodeling of peripheral T cell receptor repertoires. This constraint has left gaps in how radiation impacts intratumoral responses and whether tumor-resident T cell clones are amplified following treatment. Therefore, to interrogate the immune impact of radiation on the tumor microenvironment and test the hypothesis that radiation initiates local and systemic expansion of tumor-resident clones, we analyzed renal cell carcinomas from patients treated with stereotactic body radiation therapy. Transcriptomic comparisons were evaluated by bulk RNA sequencing. T cell receptor sequencing monitored repertoires during treatment. Pathway analysis showed radiation-specific enrichment of immune-related processes, and T cell receptor sequencing revealed increased clonality in radiation-treated tumors. The frequency of identified, tumor-enriched clonotypes was tracked across serial blood samples. We observed increased abundance of tumor-enriched clonotypes at 2 wk postradiation compared with pretreatment levels; however, this expansion was not sustained, and levels contracted toward baseline by 4 wk posttreatment. Taken together, these results indicate robust intratumoral immune remodeling and a window of tumor-resident T cell expansion following radiation that may be leveraged for the rational design of combinatorial strategies.
All cells establish and maintain an axis of polarity that is critical for cell shape and progression through the cell cycle. A well-studied example of polarity establishment is bud emergence in the yeast Saccharomyces cerevisiae, which is controlled by the Rho GTPase Cdc42p. The prevailing view of bud emergence does not account for regulation by extrinsic cues. Here, we show that the filamentous growth mitogen activated protein kinase (fMAPK) pathway regulates bud emergence under nutrient-limiting conditions. The fMAPK pathway regulated the expression of polarity targets including the gene encoding a direct effector of Cdc42p, Gic2p. The fMAPK pathway also stimulated GTP-Cdc42p levels, which is a critical determinant of polarity establishment. The fMAPK pathway activity was spatially restricted to bud sites and active during the period of the cell cycle leading up to bud emergence. Time-lapse fluorescence microscopy showed that the fMAPK pathway stimulated the rate of bud emergence during filamentous growth. Unregulated activation of the fMAPK pathway induced multiple rounds of symmetry breaking inside the growing bud. Collectively, our findings identify a new regulatory aspect of bud emergence that sensitizes this essential cellular process to external cues.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
customersupport@researchsolutions.com
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
This site is protected by reCAPTCHA and the Google Privacy Policy and Terms of Service apply.
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.