title: Mechanism of Mpk1 mitogen-activated protein kinase binding to the Swi4 transcription factor and its regulation by a novel caffeine-induced phosphorylation.
abstract: The Mpk1 mitogen-activated protein kinase (MAPK) of the cell wall integrity signaling pathway uses a noncatalytic mechanism to activate the SBF (Swi4/Swi6) transcription factor. Active Mpk1 forms a complex with Swi4, the DNA-binding subunit of SBF, conferring the ability to bind DNA. Because SBF activation is independent of Mpk1 catalytic activity but requires Mpk1 to be in an active conformation, we sought to understand how Mpk1 interacts with Swi4. Mutational analysis revealed that binding and activation of Swi4 by Mpk1 requires an intact D-motif-binding site, a docking surface common to MAPKs that resides distal to the phosphorylation loop but does not require the substrate-binding site, revealing a novel mechanism for MAPK target regulation. Additionally, we found that Mpk1 binds near the autoinhibitory C terminus of Swi4, suggesting an activation mechanism in which Mpk1 substitutes for Swi6 in promoting Swi4 DNA binding. Finally, we show that caffeine is an atypical activator of cell wall integrity signaling, because it induces phosphorylation of the Mpk1 C-terminal extension at Ser423 and Ser428. These phosphorylations were dependent on the DNA damage checkpoint kinases, Mec1/Tel1 and Rad53. Phosphorylation of Ser423 specifically blocked SBF activation by preventing Mpk1 association with Swi4, revealing a novel mechanism for regulating MAPK target specificity.
authors.affiliation: Department of Biochemistry and Molecular Biology, Johns Hopkins Bloomberg School of Public Health, Baltimore, Maryland 21205, USA.
authors.forename: Andrew W
authors.initials: AW
authors.lastname: Truman
authors.forename: Ki-Young
authors.initials: KY
authors.lastname: Kim
authors.forename: David E
authors.initials: DE
authors.lastname: Levin
mesh_terms: D000483:Alleles; D002110:Caffeine / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D002473:Cell Wall / Q000378:metabolism; D064447:Checkpoint Kinase 2; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005964:Glucosyltransferases / Q000235:genetics / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D009154:Mutation; D010726:Phosphodiesterase Inhibitors / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012694:Serine / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
chemical_list: D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D010726:Phosphodiesterase Inhibitors; C062214:SWI4 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D002110:Caffeine; D012694:Serine; D005964:Glucosyltransferases; C095724:GSC2 protein, S cerevisiae; D064447:Checkpoint Kinase 2; C088937:MEC1 protein, S cerevisiae; D017346:Protein Serine-Threonine Kinases; C513623:TEL1 protein, S cerevisiae; D020928:Mitogen-Activated Protein Kinases; C072724:SLT2 protein, S cerevisiae; C067022:RAD53 protein, S cerevisiae