Comparative Analysis of Transmembrane Regulators of the Filamentous Growth Mitogen-Activated Protein Kinase Pathway Uncovers Functional and Regulatory Differences

ABSTRACTFilamentous growth is a microbial differentiation response that involves the concerted action of multiple signaling pathways. In budding yeast, one pathway that regulates filamentous growth is a Cdc42p-dependent mitogen-activated protein kinase (MAPK) pathway. Several transmembrane (TM) proteins regulate the filamentous growth pathway, including the signaling mucin Msb2p, the tetraspan osmosensor Sho1p, and an adaptor Opy2p. The TM proteins were compared to identify common and unique features. Msb2p, Sho1p, and Opy2p associated by coimmunoprecipitation analysis but showed predominantly different localization patterns. The different localization patterns of the proteins resulted in part from different rates of turnover from the plasma membrane (PM). In particular, Msb2p (and Opy2p) were turned over rapidly compared to Sho1p. Msb2p signaled from the PM, and its turnover was a rate-limiting step in MAPK signaling. Genetic analysis identified unique phenotypes of cells overexpressing the TM proteins. Therefore, each TM regulator of the filamentous growth pathway has its own regulatory pattern and specific function in regulating filamentous growth. This specialization may be important for fine-tuning and potentially diversifying the filamentation response.

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Cdc42p-Interacting Protein Bem4p Regulates the Filamentous-Growth Mitogen-Activated Protein Kinase Pathway

Molecular and Cellular Biology ◽

10.1128/mcb.00850-14 ◽

2014 ◽

Vol 35 (2) ◽

pp. 417-436 ◽

Cited By ~ 13

Author(s):

Andrew Pitoniak ◽

Colin A. Chavel ◽

Jacky Chow ◽

Jeremy Smith ◽

Diawoye Camara ◽

...

Keyword(s):

Protein Kinase ◽

Mapk Pathway ◽

Filamentous Growth ◽

Cell Polarization ◽

Mitogen Activated Protein Kinase ◽

Specific Response ◽

Ph Domain ◽

Interacting Protein ◽

Mitogen Activated Protein ◽

Growth Pathway

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 inSaccharomyces 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.

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The tRNA Modification Complex Elongator Regulates the Cdc42-Dependent Mitogen-Activated Protein Kinase Pathway That Controls Filamentous Growth in Yeast

Eukaryotic Cell ◽

10.1128/ec.00015-09 ◽

2009 ◽

Vol 8 (9) ◽

pp. 1362-1372 ◽

Cited By ~ 20

Author(s):

Ummi Abdullah ◽

Paul J. Cullen

Keyword(s):

Protein Kinase ◽

Mapk Pathway ◽

Protein Translation ◽

Filamentous Growth ◽

Cell Polarization ◽

Mitogen Activated Protein Kinase ◽

Global Changes ◽

Trna Modification ◽

Mitogen Activated Protein ◽

Growth Pathway

ABSTRACT Signal transduction pathways control multiple aspects of cellular behavior, including global changes to the cell cycle, cell polarity, and gene expression, which can result in the formation of a new cell type. In the budding yeast Saccharomyces cerevisiae, the mitogen-activated protein kinase (MAPK) pathway that controls filamentous growth induces a dimorphic foraging response under nutrient-limiting conditions. How nutritional cues feed into MAPK activation remains an open question. Here we report a functional connection between the elongator tRNA modification complex (ELP genes) and activity of the filamentous growth pathway. Elongator was required for filamentous growth pathway signaling, and elp mutants were defective for invasive growth, cell polarization, and MAPK-dependent mat formation. Genetic suppression analysis showed that elongator functions at the level of Msb2p, the signaling mucin that operates at the head of the pathway, which led to the finding that elongator regulates the starvation-dependent expression of the MSB2 gene. The Elp complex was not required for activation of related pathways (pheromone response or high osmolarity glycerol response) that share components with the filamentous growth pathway. Because protein translation provides a rough metric of cellular nutritional status, elongator may convey nutritional information to the filamentous growth pathway at the level of MSB2 expression.

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Role of Phosphatidylinositol Phosphate Signaling in the Regulation of the Filamentous-Growth Mitogen-Activated Protein Kinase Pathway

Eukaryotic Cell ◽

10.1128/ec.00013-15 ◽

2015 ◽

Vol 14 (4) ◽

pp. 427-440 ◽

Cited By ~ 9

Author(s):

Hema Adhikari ◽

Paul J. Cullen

Keyword(s):

Protein Kinase ◽

Mapk Pathway ◽

Filamentous Growth ◽

Mapk Signaling ◽

Mitogen Activated Protein Kinase ◽

Hog Pathway ◽

Mapk Activity ◽

Conditional Allele ◽

Mitogen Activated Protein

ABSTRACTReversible phosphorylation of the phospholipid phosphatidylinositol (PI) is a key event in the determination of organelle identity and an underlying regulatory feature in many biological processes. Here, we investigated the role of PI signaling in the regulation of the mitogen-activated protein kinase (MAPK) pathway that controls filamentous growth in yeast. Lipid kinases that generate phosphatidylinositol 4-phosphate [PI(4)P] at the Golgi (Pik1p) or PI(4,5)P2 at the plasma membrane (PM) (Mss4p and Stt4p) were required for filamentous-growth MAPK pathway signaling. Introduction of a conditional allele ofPIK1(pik1-83) into the filamentous (Σ1278b) background reduced MAPK activity and caused defects in invasive growth and biofilm/mat formation. MAPK regulatory proteins that function at the PM, including Msb2p, Sho1p, and Cdc42p, were mislocalized in thepik1-83mutant, which may account for the signaling defects of the PI(4)P kinase mutants. Other PI kinases (Fab1p and Vps34p), and combinations of PIP (synaptojanin-type) phosphatases, also influenced the filamentous-growth MAPK pathway. Loss of these proteins caused defects in cell polarity, which may underlie the MAPK signaling defect seen in these mutants. In line with this possibility, disruption of the actin cytoskeleton by latrunculin A (LatA) dampened the filamentous-growth pathway. Various PIP signaling mutants were also defective for axial budding in haploid cells, cell wall construction, or proper regulation of the high-osmolarity glycerol response (HOG) pathway. Altogether, the study extends the roles of PI signaling to a differentiation MAPK pathway and other cellular processes.

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APPL1 mediates adiponectin-stimulated p38 MAPK activation by scaffolding the TAK1-MKK3-p38 MAPK pathway

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00427.2010 ◽

2011 ◽

Vol 300 (1) ◽

pp. E103-E110 ◽

Cited By ~ 59

Author(s):

Xiaoban Xin ◽

Lijun Zhou ◽

Caleb M. Reyes ◽

Feng Liu ◽

Lily Q. Dong

Keyword(s):

Protein Kinase ◽

P38 Mapk ◽

Mapk Pathway ◽

Adaptor Protein ◽

Mapk Signaling ◽

Mitogen Activated Protein Kinase ◽

Scaffolding Protein ◽

Mapk Activation ◽

P38 Mapk Pathway ◽

Mitogen Activated Protein

The adaptor protein APPL1 mediates the stimulatory effect of adiponectin on p38 mitogen-activated protein kinase (MAPK) signaling, yet the underlying mechanism remains unclear. Here we show that, in C2C12 cells, overexpression or suppression of APPL1 enhanced or suppressed, respectively, adiponectin-stimulated p38 MAPK upstream kinase cascade, consisting of transforming growth factor-β-activated kinase 1 (TAK1) and mitogen-activated protein kinase kinase 3 (MKK3). In vitro affinity binding and coimmunoprecipitation experiments revealed that TAK1 and MKK3 bind to different regions of APPL1, suggesting that APPL1 functions as a scaffolding protein to facilitate adiponectin-stimulated p38 MAPK activation. Interestingly, suppressing APPL1 had no effect on TNFα-stimulated p38 MAPK phosphorylation in C2C12 myotubes, indicating that the stimulatory effect of APPL1 on p38 MAPK activation is selective. Taken together, our study demonstrated that the TAK1-MKK3 cascade mediates adiponectin signaling and uncovers a scaffolding role of APPL1 in regulating the TAK1-MKK3-p38 MAPK pathway, specifically in response to adiponectin stimulation.

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Identification of RAS-Mitogen-Activated Protein Kinase Signaling Pathway Modulators in an ERF1 Redistribution® Screen

CrossRef Listing of Deleted DOIs ◽

10.1177/1087057106287136 ◽

2006 ◽

Vol 11 (4) ◽

pp. 423-434 ◽

Cited By ~ 12

Author(s):

Charlotta Grånäs ◽

Betina Kerstin Lundholt ◽

Frosty Loechel ◽

Hans-Christian Pedersen ◽

Sara Petersen Bjørn ◽

...

Keyword(s):

Protein Kinase ◽

Signaling Pathway ◽

Kinase Inhibitors ◽

Mapk Pathway ◽

Mapk Signaling ◽

Mitogen Activated Protein Kinase ◽

Mitogen Activated Protein ◽

A Cell ◽

Protein Kinase Signaling

The RAS-mitogen-activated protein kinase (MAPK) signaling pathway has a central role in regulating the proliferation and survival of both normal and tumor cells. This pathway has been 1 focus area for the development of anticancer drugs, resulting in several compounds, primarily kinase inhibitors, in clinical testing. The authors have undertaken a cell-based, high-throughput screen using a novel ERF1 Redistribution® assay to identify compounds that modulate the signaling pathway. The hit compounds were subsequently tested for activity in a functional cell proliferation assay designed to selectively detect compounds inhibiting the proliferation of MAPK pathway-dependent cancer cells. The authors report the identification of 2 cell membrane-permeable compounds that exhibit activity in the ERF1 Redistribution® assay and selectively inhibit proliferation of MAPK pathway-dependent malignant melanoma cells at similar potencies (IC50 =< 5 μM). These compounds have drug-like structures and are negative in RAF, MEK, and ERK in vitro kinase assays. Drugs belonging to these compound classes may prove useful for treating cancers caused by excessive MAPK pathway signaling. The results also show that cell-based, high-content Redistribution® screens can detect compounds with different modes of action and reveal novel targets in a pathway known to be disease relevant.

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Role of the Unfolded Protein Response in Regulating the Mucin-Dependent Filamentous-Growth Mitogen-Activated Protein Kinase Pathway

Molecular and Cellular Biology ◽

10.1128/mcb.01501-14 ◽

2015 ◽

Vol 35 (8) ◽

pp. 1414-1432 ◽

Cited By ~ 26

Author(s):

Hema Adhikari ◽

Nadia Vadaie ◽

Jacky Chow ◽

Lauren M. Caccamise ◽

Colin A. Chavel ◽

...

Keyword(s):

Protein Kinase ◽

Unfolded Protein Response ◽

Mapk Pathway ◽

Filamentous Growth ◽

Proteolytic Processing ◽

Mitogen Activated Protein Kinase ◽

Unfolded Protein ◽

Mitogen Activated Protein ◽

The Unfolded Protein Response ◽

Protein Response

Signaling 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.

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Graded Mitogen-Activated Protein Kinase Activity Precedes Switch-Like c-Fos Induction in Mammalian Cells

Molecular and Cellular Biology ◽

10.1128/mcb.25.11.4676-4682.2005 ◽

2005 ◽

Vol 25 (11) ◽

pp. 4676-4682 ◽

Cited By ~ 71

Author(s):

Jeffrey P. MacKeigan ◽

Leon O. Murphy ◽

Christopher A. Dimitri ◽

John Blenis

Keyword(s):

Protein Kinase ◽

Cell Fate ◽

Oocyte Maturation ◽

Mammalian Cells ◽

Mapk Pathway ◽

Mapk Signaling ◽

Mitogen Activated Protein Kinase ◽

Cell Fate Decisions ◽

Graded Response ◽

Mitogen Activated Protein

ABSTRACT The mitogen-activated protein kinase (MAPK) pathway is an evolutionarily conserved signaling module that controls important cell fate decisions in a variety of physiological contexts. During Xenopus oocyte maturation, the MAPK cascade converts an increasing progesterone stimulus into a switch-like, all-or-nothing response. While the importance of such switch-like behavior is widely discussed in the literature, it is not known whether the MAPK pathway in mammalian cells exhibits a switch-like or graded response. For this study, we used flow cytometry and immunofluorescence to generate single-cell measurements of MAPK signaling in Swiss 3T3 fibroblasts. In contrast to the case in Xenopus oocytes, we found that ERK activation in individual mammalian cells is not ultrasensitive and shows a graded response to changes in agonist concentration. Thus, the conserved MAPK signaling module exhibits different systems-level properties in different cellular contexts. Furthermore, the graded ERK response was converted into a more switch-like behavior at the level of immediate-early gene induction and cell cycle progression. Thus, while MAPK signaling is involved in all-or-nothing cell fate decisions for both Xenopus oocyte maturation and mammalian fibroblast proliferation, the underlying mechanisms responsible for the switch-like nature of the cellular responses are different in these two systems, with the mechanism appearing to lie downstream of the kinase cascade in mammalian fibroblasts.

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Rho2 Is a Target of the Farnesyltransferase Cpp1 and Acts Upstream of Pmk1 Mitogen-activated Protein Kinase Signaling in Fission Yeast

Molecular Biology of the Cell ◽

10.1091/mbc.e06-08-0688 ◽

2006 ◽

Vol 17 (12) ◽

pp. 5028-5037 ◽

Cited By ~ 52

Author(s):

Yan Ma ◽

Takayoshi Kuno ◽

Ayako Kita ◽

Yuta Asayama ◽

Reiko Sugiura

Keyword(s):

Protein Kinase ◽

Fission Yeast ◽

Mapk Pathway ◽

Chloride Ion ◽

Mapk Signaling ◽

Small Gtpase ◽

Mitogen Activated Protein Kinase ◽

Protein Farnesyltransferase ◽

Mitogen Activated Protein ◽

Novel Target

We have previously demonstrated that knockout of the calcineurin gene or inhibition of calcineurin activity by immunosuppressants resulted in hypersensitivity to Cl− in fission yeast. We also demonstrated that knockout of the components of the Pmk1 mitogen-activated protein kinase (MAPK) pathway, such as Pmk1 or Pek1 complemented the hypersensitivity to Cl−. Using this interaction between calcineurin and Pmk1 MAPK, here we developed a genetic screen that aims to identify new regulators of the Pmk1 signaling and isolated vic (viable in the presence of immunosuppressant and chloride ion) mutants. One of the mutants, vic1-1, carried a missense mutation in the cpp1+ gene encoding a β subunit of the protein farnesyltransferase, which caused an amino acid substitution of aspartate 155 of Cpp1 to asparagine (Cpp1D155N). Analysis of the mutant strain revealed that Rho2 is a novel target of Cpp1. Moreover, Cpp1 and Rho2 act upstream of Pck2–Pmk1 MAPK signaling pathway, thereby resulting in the vic phenotype upon their mutations. Interestingly, compared with other substrates of Cpp1, defects of Rho2 function were more phenotypically manifested by the Cpp1D155N mutation. Together, our results demonstrate that Cpp1 is a key component of the Pck2–Pmk1 signaling through the spatial control of the small GTPase Rho2.

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Proprotein Convertase Subtilisin/Kexin Type 9 Promotes Gastric Cancer Metastasis and Suppresses Apoptosis by Facilitating MAPK Signaling Pathway Through HSP70 Up-Regulation

Frontiers in Oncology ◽

10.3389/fonc.2020.609663 ◽

2021 ◽

Vol 10 ◽

Author(s):

Beili Xu ◽

Shuyu Li ◽

Yong Fang ◽

Yanting Zou ◽

Dongqiang Song ◽

...

Keyword(s):

Gastric Cancer ◽

Protein Kinase ◽

Signaling Pathway ◽

Western Blotting ◽

Mapk Pathway ◽

Serum Levels ◽

Mapk Signaling ◽

Mitogen Activated Protein Kinase ◽

Proprotein Convertase ◽

Mitogen Activated Protein

ObjectiveTo examine the effect of proprotein convertase subtilisin/kexin type 9 (PCSK9) on gastric cancer (GC) progression and prognosis, and to explore the underlying mechanism.MethodsPCSK9 expression levels in human GC tissues were determined by quantitative real-time PCR, western blotting, and immunohistochemical assay. PCSK9 serum levels were detected by enzyme-linked immunosorbent assay. The relationships of PCSK9 and GC progression and survival were analyzed using the Chi-square test, Kaplan-Meier analysis, and Cox proportional hazards model. The effect of PCSK9 on cell invasion, migration, and apoptosis were determined in human GC cell lines and mouse xenograft model separately using PCSK9 knockdown and overexpression strategies. The PCSK9 interacting molecules, screened by co-immunoprecipitation combined with LC-MS/MS, were identified by immunofluorescence localization and western blotting. Additionally, the mitogen-activated protein kinase (MAPK) pathway was assessed by western blotting.ResultsPCSK9 mRNA and protein levels were significantly elevated in GC tissues compared with the paired normal tissues at our medical center (P < 0.001). Notably, the up-regulation of PCSK9 expression in GC tissues was related to tumor progression and poor survival. GC patients had higher serum levels of PCSK9 than the age-matched healthy controls (P < 0.001); PCSK9 promoted invasive and migratory ability and inhibited apoptosis in GC cells with no apparent affection in cell proliferation. The silencing of PCSK9 reversed these effects, suppressing tumor metastasis in vitro and in vivo. Furthermore, PCSK9 maintained these functions through up-regulating heat shock protein 70 (HSP70), ultimately facilitating the mitogen-activated protein kinase (MAPK) pathway.ConclusionCollectively, our data revealed that high PCSK9 expression levels in GC tissue were correlated with GC progression and poor prognosis and that PCSK9 could promote GC metastasis and suppress apoptosis by facilitating MAPK signaling pathway through HSP70 up-regulation. PCSK9 may represent a novel potential therapeutic target in GC.

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PKCζ-Mitogen-Activated Protein Kinase Signaling Mediates Crotalphine-Induced Antinociception

Toxins ◽

10.3390/toxins13120912 ◽

2021 ◽

Vol 13 (12) ◽

pp. 912

Author(s):

Bárbara G. de Freitas ◽

Natália G. Hösch ◽

Leandro M. Pereira ◽

Tereza C. Barbosa ◽

Gisele Picolo ◽

...

Keyword(s):

Spinal Cord ◽

Protein Kinase ◽

Intracellular Signaling ◽

Mapk Pathway ◽

Antinociceptive Effect ◽

Primary Cultures ◽

Receptor Activation ◽

Mapk Signaling ◽

Mitogen Activated Protein Kinase ◽

Mitogen Activated Protein

Crotalphine (CRP) is a structural analogue to a peptide that was first identified in the crude venom from the South American rattlesnake Crotalus durissus terrificus. This peptide induces a potent and long-lasting antinociceptive effect that is mediated by the activation of peripheral opioid receptors. The opioid receptor activation regulates a variety of intracellular signaling, including the mitogen-activated protein kinase (MAPK) pathway. Using primary cultures of sensory neurons, it was demonstrated that crotalphine increases the level of activated ERK1/2 and JNK-MAPKs and this increase is dependent on the activation of protein kinase Cζ (PKCζ). However, whether PKCζ-MAPK signaling is critical for crotalphine-induced antinociception is unknown. Here, we biochemically demonstrated that the systemic crotalphine activates ERK1/2 and JNK and decreases the phosphorylation of p38 in the lumbar spinal cord. The in vivo pharmacological inhibition of spinal ERK1/2 and JNK, but not of p38, blocks the antinociceptive effect of crotalphine. Of interest, the administration of a PKCζ pseudosubstrate (PKCζ inhibitor) prevents crotalphine-induced ERK activation in the spinal cord, followed by the abolishment of crotalphine-induced analgesia. Together, our results demonstrate that the PKCζ-ERK signaling pathway is involved in crotalphine-induced analgesia. Our study opens a perspective for the PKCζ-MAPK axis as a target for pain control.

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