Mapping of MST1 Kinase Sites of Phosphorylation

MST1 is a member of the Sterile-20 family of cytoskeletal, stress, and apoptotic kinases. MST1 is activated by phosphorylation at previously unidentified sites. This study examines the role of phosphorylation at several sites and effects on kinase activation. We define Thr183 in subdomain VIII as a primary site of phosphoactivation. Thr187 is also critical for kinase activity. Phosphorylation of MST1 in subdomain VIII was catalyzed by active MST1 via intermolecular autophosphorylation, enhanced by homodimerization. Active MST1 (wild-type or T183E), but not inactive Thr183/Thr187 mutants, was also highly autophosphorylated at the newly identified Thr177 and Thr387 residues. Cells expressing active MST1 were mostly detached, whereas with inactive MST1, adhesion was normal. Active MKK4, JNK, caspase-3, and caspase-9 were detected in the detached cells. These cells also contained all autophosphorylated and essentially all caspase-cleaved MST1. Similar phenotypes were elicited by a caspase-insensitive D326N mutant, suggesting that kinase activity, but not cleavage of MST1, is required. Interestingly, an S327E mutant mimicking Ser327 autophosphorylation was also caspase-insensitive, but only when expressed in caspase-3-deficient cells. Together, these data suggest a model whereby MST1 activation is induced by existing, active MST kinase, which phosphorylates Thr183 and possibly Thr187. Dimerization promotes greater phosphorylation. This leads to induction of the JNK signaling pathway, caspase activation, and apoptosis. Further activation of MST1 by caspase cleavage is best promoted by caspase-3, although this appears to be unnecessary for signaling and morphological responses. MST1 is a member of the Sterile-20 family of cytoskeletal, stress, and apoptotic kinases. MST1 is activated by phosphorylation at previously unidentified sites. This study examines the role of phosphorylation at several sites and effects on kinase activation. We define Thr183 in subdomain VIII as a primary site of phosphoactivation. Thr187 is also critical for kinase activity. Phosphorylation of MST1 in subdomain VIII was catalyzed by active MST1 via intermolecular autophosphorylation, enhanced by homodimerization. Active MST1 (wild-type or T183E), but not inactive Thr183/Thr187 mutants, was also highly autophosphorylated at the newly identified Thr177 and Thr387 residues. Cells expressing active MST1 were mostly detached, whereas with inactive MST1, adhesion was normal. Active MKK4, JNK, caspase-3, and caspase-9 were detected in the detached cells. These cells also contained all autophosphorylated and essentially all caspase-cleaved MST1. Similar phenotypes were elicited by a caspase-insensitive D326N mutant, suggesting that kinase activity, but not cleavage of MST1, is required. Interestingly, an S327E mutant mimicking Ser327 autophosphorylation was also caspase-insensitive, but only when expressed in caspase-3-deficient cells. Together, these data suggest a model whereby MST1 activation is induced by existing, active MST kinase, which phosphorylates Thr183 and possibly Thr187. Dimerization promotes greater phosphorylation. This leads to induction of the JNK signaling pathway, caspase activation, and apoptosis. Further activation of MST1 by caspase cleavage is best promoted by caspase-3, although this appears to be unnecessary for signaling and morphological responses. mitogen-activated protein kinase c-Jun N-terminal kinase mitogen-activated protein kinase/extracellular signal-regulated kinase kinase mitogen-activated protein kinase/extracellular signal-regulated kinase kinase kinase tumor necrosis factor p21-activated kinase mitogen-activated protein kinase kinase wild-type β-glycerophosphate/HEPES-buffered solution SAPK/Erk kinase stress-activated protein kinase poly(ADP-ribose) polymerase combined caspase-3, -6, and -7 activities extracellular signal-regulated kinase cycloheximide In mammalian cells, Sterile-20 (Ste20)-related kinases participate in the regulation of the cytoskeleton that controls cell morphology and motility, and in the regulation of apoptosis (1Dan I. Watanabe N.M. Kusumi A. Trends Cell Biol. 2001; 11: 220-230Abstract Full Text Full Text PDF PubMed Scopus (514) Google Scholar). These kinases share a conserved catalytic (kinase) domain at the amino terminus and a C-terminal regulatory region of great structural diversity, which interacts with signaling molecules that regulate the cytoskeleton. Currently, four closely related MST kinases have been described (5Creasy C.L. Chernoff J. J. Biol. Chem. 1995; 270: 21695-21700Abstract Full Text Full Text PDF PubMed Scopus (174) Google Scholar, 6Creasy C.L. Chernoff J. Gene (Amst.). 1995; 167: 303-306Crossref PubMed Scopus (119) Google Scholar, 7Qian Z. Lin C. Espinosa R. LeBeau M. Rosner M.R. J. Biol. Chem. 2001; 276: 22439-22445Abstract Full Text Full Text PDF PubMed Scopus (60) Google Scholar, 8Schinkmann K. Blenis J. J. Biol. Chem. 1997; 272: 28695-28703Abstract Full Text Full Text PDF PubMed Scopus (94) Google Scholar, 9Taylor L.K. Wang H.C. Erikson R.L. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 10099-10104Crossref PubMed Scopus (141) Google Scholar). Most Ste20 group kinases activate mitogen-activated protein kinase (MAPK)1cascades in the signaling pathways between the cellular membrane and the nuclear compartment (2Hagemann C. Blank J.L. Cell. Signal. 2001; 13: 863-875Crossref PubMed Scopus (245) Google Scholar). In yeast, the mating pheromone receptor Ste20p phosphorylates and activates a MAPK kinase kinase, Ste11p, raising the possibility that mammalian homologs of Ste20p (e.g. MST1 kinase) also function as MAPK kinase kinase kinases (3Wu C. Whiteway M. Thomas D.Y. Leberer E. J. Biol. Chem. 1995; 270: 15984-15992Abstract Full Text Full Text PDF PubMed Scopus (150) Google Scholar, 4Graves J.D. Gotoh Y. Draves K.E. Ambrose D. Han D.K. Wright M. Chernoff J. Clark E.A. Krebs E.G. EMBO J. 1998; 17: 2224-2234Crossref PubMed Scopus (325) Google Scholar). MST1 was shown to act upstream of MAPK kinases that regulate p38 and JNK activities, probably acting via the MAPK kinase kinase MEKK1 (10Graves J.D. Draves K.E. Gotoh Y. Krebs E.G. Clark E.A. J. Biol. Chem. 2001; 276: 14909-14915Abstract Full Text Full Text PDF PubMed Scopus (136) Google Scholar). There is substantial evidence that MST1 promotes apoptosis, although role in this in cell of MST1 apoptosis and nuclear in and cells J.D. Gotoh Y. Draves K.E. Ambrose D. Han D.K. Wright M. Chernoff J. Clark E.A. Krebs E.G. EMBO J. 1998; 17: 2224-2234Crossref PubMed Scopus (325) Google Scholar, S. J.D. Gotoh Y. 2001; PubMed Scopus Google Scholar, S. J.D. Gotoh Y. Proc. Natl. Acad. Sci. U. S. A. 2001; PubMed Scopus Google whereas MST1 promotes nuclear cleavage in cells S. S. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus Google Scholar). of MST1 in all cell is cleavage by in to apoptotic to a to as MST1 kinase J.D. Gotoh Y. Draves K.E. Ambrose D. Han D.K. Wright M. Chernoff J. Clark E.A. Krebs E.G. EMBO J. 1998; 17: 2224-2234Crossref PubMed Scopus (325) Google Scholar, M. E. S. S. K. S. 1998; PubMed Scopus Google Scholar, J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, M. PubMed Scopus Google Scholar, R. Y. Acad. Sci. PubMed Scopus Google and S. S. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus Google Scholar, S. J.D. Gotoh Y. Proc. Natl. Acad. Sci. U. S. A. 2001; PubMed Scopus Google Scholar). a caspase cleavage site was identified in the MST1 that is in MST1 and in several (10Graves J.D. Draves K.E. Gotoh Y. Krebs E.G. Clark E.A. J. Biol. Chem. 2001; 276: 14909-14915Abstract Full Text Full Text PDF PubMed Scopus (136) Google Scholar). of this site on of the of a MST1 mutant or apoptosis or in and cells with apoptotic S. J.D. Gotoh Y. Proc. Natl. Acad. Sci. U. S. A. 2001; PubMed Scopus Google Scholar, M. PubMed Scopus Google Scholar). This is with of JNK and the mutant to apoptosis in and cells with or J.D. Gotoh Y. Draves K.E. Ambrose D. Han D.K. Wright M. Chernoff J. Clark E.A. Krebs E.G. EMBO J. 1998; 17: 2224-2234Crossref PubMed Scopus (325) Google Scholar, M. E. S. S. K. S. 1998; PubMed Scopus Google Scholar). for the cell and in MST function not been Interestingly, 1997; 276: PubMed Scopus Google Scholar, S. Ambrose Chernoff J. Biol. 1997; Full Text Full Text PDF PubMed Scopus Google MST1 or have a on cell and of caspase activation and to nuclear S. S. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus Google Scholar). This and the of MST1 kinase suggest a function to apoptosis. as and L.K. Wang H.C. Erikson R.L. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 10099-10104Crossref PubMed Scopus (141) Google MST1 kinase of MST1 been and is the activation of MST1. was that in to caspase MST1 phosphorylation at unidentified sites to kinase activation (10Graves J.D. Draves K.E. Gotoh Y. Krebs E.G. Clark E.A. J. Biol. Chem. 2001; 276: 14909-14915Abstract Full Text Full Text PDF PubMed Scopus (136) Google as been for A. Z. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, Z. R. J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar). and Chernoff (5Creasy C.L. Chernoff J. J. Biol. Chem. 1995; 270: 21695-21700Abstract Full Text Full Text PDF PubMed Scopus (174) Google that of MST1 with protein in in of activity, suggesting that phosphorylation sites in of activity. of this of of MST1 not been of the related to MST a of MST1 phosphorylation sites. We identified phosphorylation sites by a role for intermolecular autophosphorylation of the activation of MST1 in subdomain We identified Thr183 and Thr187 as for kinase and for effects on as and Phosphorylation at Thr177 the of MST1, suggesting an on the of the activation We also that MST1 phosphorylation on MST1 cleavage by apoptosis, but to cleavage by cells and cells were Cells were at in with and in a with cells and cells were by and in with and of cells was induced by with and cycloheximide for was with and MST1 was with polymerase and site and a was the sites of with the and MST1 N-terminal kinase domain to the caspase cleavage of MST1 was with a site and a This was the sites of MST1 was by the was with a the were by or cells were at and cells were at in and to for at or Cells were with of and of to the were for at protein were cells were on and with β-glycerophosphate/HEPES-buffered solution and of cells, the was with the and at to cells. Cell were in of to and at of cells, cells were the in of to and at Cells were in with and a J. 2001; PubMed Scopus (150) Google Scholar). were a of protein were and on or were which were with and in and were at with and Cell and p38 MAPK and polymerase at were in was with and enhanced an were in of and the and combined with primary and protein were at were four with and were by in and or at were the described previously J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google with the kinase contained in of were with of protein or the of and were and for kinase were to and and the and MST1, and were cells. were and the of protein were by with MST1 were on the were to which was as described were with kinase and were in of kinase with of or in for at kinase was by and was to was caspase-9 and were Cells were with solution and and in the by the were by and were to the with of protein were and activities were a at in of Cells were with and the as described the were with and cells were in for cells were at with and in MST1 was cells by for the and is to as of at by with the and was by described were expressed with an N-terminal of MST1 and amino in this study in cells were with MST1 or in and by and kinase kinase of the was substantial and was not by of or cells not of active cell and of cells. This was as a in the of cells and and by the in the of the wild-type kinase in the cell by but not by inactive shown is the cell induced by MST1 MST1 were with in cells. cells were and for activity. the of cells in was a was with cell of and cell were for kinase activities by kinase and caspase activation was by for MST1 and the MST1 caspase cleavage and the caspase cleavage of also the of MST1 cleavage and kinase to the effects of the on expressed and caspase-insensitive mutant kinases in cells. with a S. S. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus Google kinase was for MST1 induction of cell and and Interestingly, the morphological and cell by the and were elicited by the MST1 kinase to be and to morphological in to which MST1 caspase cleavage or of the C. M. J. J. 2001; PubMed Scopus Google Scholar). the between MST1 activity, caspase activity, and cell and and detached cells. In of were cells when with or when with as activities were or by of cells with induction of apoptosis, as by of the which was in all cell with or This with of and caspase-9 activities, which were by in cells of in cells expressing and was by to and with the This an in the of detached cells and an to in caspase MST1 was by kinase at the kinase were with of activities of and were by in cells. of the caspase cleavage was mostly in the cell and at to the In cells, the caspase-cleaved kinase of the of the was kinase was and the was to that of the combined and caspase-cleaved kinases. Together, these data suggest that of active MST1 leads to cell and caspase activation. cleavage of MST1 in and of is not for of MST1 subdomain VIII with of the Ste20 family and and kinases in MST1 of residues. with the in and and Thr183 with the in and Thr187 also conserved the Ste20 family although phosphorylation at these sites not been We to phosphorylation at these and to MST1 kinase all four with and or and and catalytic activities We to for phosphorylation at Thr177 and Thr183 to a phosphorylation at of J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). of Thr183 and Thr187 the catalytic of MST1 the to of that of This was the in which of the wild-type kinase J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). substantial was the of catalytic with the with a role for Thr183 phosphorylation in MST1 kinase activation, the mutant was with for a the mutant which is to the effects of in as J. D. 1995; Full Text Full Text PDF PubMed Scopus Google Scholar). in the of the mutant, of kinase of and and or not in or morphological and of cells. of the mostly active mutant morphological and that were to by the wild-type kinase and of and Thr177 not catalytic and in and morphological cell was to that with the kinase not MST1 was for the MST1 kinase S. S. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus Google although the were not identified of Thr177 and Thr183 the of the mutant of the and caspase-cleaved kinases This was in in the active mutant at with the to that of We the kinase, also a and a Interestingly, as the evidence of This that in with phosphorylation at phosphorylation in this were with that phosphorylation at Thr177 and Thr183 and to suggesting that these were In the mutant, by these was suggesting that kinase be for phosphorylation to at these sites. the not by with phosphorylation at the by whereas by was by with a role in autophosphorylation, also by inactive mutant a suggest that catalytic phosphorylation at active mutant was caspase-cleaved kinase as a and caspase-cleaved wild-type and kinases were with and was by and as with a for catalytic for phosphorylation at the mutant, the kinase, was by Interestingly, although the caspase-cleaved was a detected only in the this was the was Together, these data with MST1 activation via phosphorylation at the inactive of the Thr187 also suggest phosphorylation at this the was be for this phosphorylation at Thr177 not catalytic activity, but phosphorylation at Thr177 and Thr183 the activation was when the inactive kinase was suggesting autophosphorylation at these sites. the and the of activation via the cells MST1, which and to activate newly MST1. We intermolecular active kinase to inactive in This inactive mutant although phosphorylation sites of phosphorylation at Thr183 in possibly by MST kinase, a for in of Active and inactive kinases were expressed in cells as and and and These be in kinase were expressed MST1 were combined to active and inactive on the active kinase was combined with the inactive kinase and the inactive kinase was combined with the active kinase a combined inactive and kinase were on the as described and Active MST1 autophosphorylated or when was in the kinase as by of and was also the in This when inactive were with the active the of inactive in of substantial phosphorylation of which by intermolecular autophosphorylation, and Thr387 were as were in the of and phosphorylation was detected by phosphorylation of the inactive was of in induced by active MST1. inactive MST1 was by the active phosphorylation at Thr177 was by and phosphorylation at Thr183 was by kinase was also by the active This phosphorylation at Thr177 by and at Thr183 by of in phosphorylation at these sites of the mutant the phosphorylation in cells and the phosphorylation of the mutant in these cells. be with MST kinase, which be by the We also phosphorylation at which is of an by several kinases. this but only the inactive was identified by with all by this not In phosphorylation of the mutant at detected by was by by with the wild-type kinase kinase, which was detected by but only in kinase for this was identified as which is of an with this and the or to the MST1 kinase not at the critical Thr183 was by of and the cells. kinases were with were with and by which that only were by these were for phosphorylation at Thr183 In with of with in a in phosphorylation to at with inactive a in phosphorylation was of Together, these data suggest that MST1 is by intermolecular of the activation of MST1 at sites Thr177 and Thr183 as as the of Thr387 to was not been that p38 and JNK kinases be activated by MST1 when kinase is with and with or J.D. Gotoh Y. Draves K.E. Ambrose D. Han D.K. Wright M. Chernoff J. Clark E.A. Krebs E.G. EMBO J. 1998; 17: 2224-2234Crossref PubMed Scopus (325) Google Scholar). of p38 and JNK pathways was by of MST1 M. E. S. S. K. S. 1998; PubMed Scopus Google Scholar). In the that of MST1 in cells of cells. active MST1 was mostly in the We that signaling also the cell induced cell the mutant not a of the to that of cells, was We the and cells with and the mutant a of cells, cells with the mutant with were of cell the or cells of with the that active MST1 was mostly in detached cells and phosphorylation at Thr183 and Thr177 was detected only in the cell by the kinase was not in was substantial of the caspase-cleaved kinase in the cells. although the not cell the activated detected in this of cells the inactive MST1 of kinase was also detected by kinase to the of cells this of was not when were of and cells and active MST1 (wild-type and active detached cells, the of the expressed kinase was detected in combined and cells. We the cells for activities of the kinases p38 and JNK with the that these kinases were activated only in detached the for phosphorylation were in cells expressing inactive or the not suggesting that and not MST1 activity, activation of the MST1 cell p38 activation is a of MST kinase activity. of active and JNK also that was to the cell in to the in JNK phosphorylation was detected only in cells expressing active MST1. We also phosphorylation of the JNK at activation the to substantial of active JNK in the a of active was detected in these cells. this was in the cells expressing In of cells, activation of and JNK was also the p38 was not In related the and elicited a to that of the wild-type kinase, whereas the induced by the controls that induced by not Similar were cells for Together, these data suggest that MST1 activates the JNK signaling in and cells. of p38 to on the of cells on MST1 kinase MST1 of cells, p38 activation be the of this activation of and mutants, this appears to be a function of kinase In cells expressing active MST1, in the cell is substantial cleavage of the kinase to the -6, and were previously shown to MST1 at the in (10Graves J.D. Draves K.E. Gotoh Y. Krebs E.G. Clark E.A. J. Biol. Chem. 2001; 276: 14909-14915Abstract Full Text Full Text PDF PubMed Scopus (136) Google S. S. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus Google Scholar). the caspase cleavage of MST1 in cells, cells, which Y. C. 2001; PubMed Scopus Google and cells, which the of caspase-3, cells apoptosis in to C. M. J. J. 2001; PubMed Scopus Google Scholar). caspase-3, -6, and -7 activities, cleavage of a In of cells, was the of was by in and cells. In with cells, cells of the caspase in the and induced and cleavage was in a cleavage of was by and this was by the caspase at Ser327 in MST1 previously been in of caspase cleavage in J.D. Draves K.E. Gotoh Y. Krebs E.G. Clark E.A. J. Biol. Chem. 2001; 276: 14909-14915Abstract Full Text Full Text PDF PubMed Scopus (136) Google Scholar). the role of Ser327 phosphorylation in cleavage by in cells, the of caspase cleavage and in and of caspase as of expressed MST1 was essentially to that of and was as and in cells and to and in cells. This that autophosphorylation at Ser327 in is or that phosphorylation at this site not caspase in with to was in cells all to the D326N cleavage of the S327E mutant was in cells to and This was with was in apoptotic cells but cleavage of was detected In of in cells was in the autophosphorylation at Ser327 cleavage in the of activity. of wild-type or active MST1 was previously shown to morphological and cell S. S. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus Google Scholar). was MST1 cleavage is a for these study a role for the and kinases in cell have a role for in the activation of subdomain VIII and in the of catalytic activity. MST1 been identified as an via of subdomain phosphorylation at possibly to a have the role of in caspase to activate MST1 to and L.K. Wang H.C. Erikson R.L. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 10099-10104Crossref PubMed Scopus (141) Google Scholar). the apoptotic role of is MST1 have to MST1 kinase by in (5Creasy C.L. Chernoff J. J. Biol. Chem. 1995; 270: 21695-21700Abstract Full Text Full Text PDF PubMed Scopus (174) Google J.D. Draves K.E. Gotoh Y. Krebs E.G. Clark E.A. J. Biol. Chem. 2001; 276: 14909-14915Abstract Full Text Full Text PDF PubMed Scopus (136) Google unidentified in of kinase activity. that for activities via a region subdomain VIII the In or autophosphorylation at in the C-terminal kinase domain a role in kinase J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). Phosphorylation of at to and a kinase activation by J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). this substantial activity, suggesting or regulation of MST1 Thr183 to role in kinase activation, catalytic to the mimicking a MST1 kinase activity. This and the of phosphorylation at this site via phosphorylation suggest Thr183 as a site of a phosphorylation of to of R. Z. J. PubMed Scopus Google was to with and MST kinases in or A. J. 2001; PubMed Scopus Google Scholar). We this to phosphorylation at Thr183 of MST1 in the wild-type this was in and S. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google described a role for Thr183 phosphorylation in of MST1 whereby Thr183 phosphorylation nuclear of MST1. was as to or not a catalytic activity. that phosphorylation or with an at this site is critical for activity. In identified Thr187 as for MST1 activity, although was not by an sites the MST1 activation and were also as for but was to be critical for kinase activity. is that the caspase-cleaved wild-type and kinases as a whereas This autophosphorylation at this site in the We identified Thr387 the domain C.L. Chernoff J. J. Biol. Chem. 1996; Full Text Full Text PDF PubMed Scopus Google as a site for at Thr387 not kinase or morphological and cell not and intermolecular autophosphorylation at this and sites although Ser327 as the primary site for MST1 autophosphorylation in an autophosphorylation by (10Graves J.D. Draves K.E. Gotoh Y. Krebs E.G. Clark E.A. J. Biol. Chem. 2001; 276: 14909-14915Abstract Full Text Full Text PDF PubMed Scopus (136) Google Scholar). In of is that phosphorylation at Ser327 is in the whereas sites identified Ser327 the in In the in was detected inactive as a phosphorylation in was detected in the to the intermolecular activities of MST kinases. this in of phosphorylation at and this a for of in with active MST1, substantial in phosphorylation at and Thr387 the This that MST1 is of intermolecular at all of these and this was in a phosphorylation at Thr183 by of active MST1 phosphorylation at all identified sites is is for Thr183 an at this which the kinase We that intermolecular phosphorylation at Thr183 possibly by with MST kinase, the for activation of the newly is that the phosphorylation at Thr177 and Thr183 in the inactive kinase was that in the kinase, which is of with MST kinases to the in phosphorylation at Thr177 and Thr183 was to and in the site of MST1 controls an apoptotic of MST1 in (10Graves J.D. Draves K.E. Gotoh Y. Krebs E.G. Clark E.A. J. Biol. Chem. 2001; 276: 14909-14915Abstract Full Text Full Text PDF PubMed Scopus (136) Google and in cells a of caspase MST1 cellular S. S. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus Google and in the which be with the S. J.D. Gotoh Y. 2001; PubMed Scopus Google Scholar). phosphorylation at Ser327 in cells is which role in to caspase that the S327E mutant was to this was only in caspase-3-deficient cells. the in cells or expressing caspase-3, cleavage of the S327E mutant with the of with the of caspase activity. cells substantial to and cleavage of the S327E mutant at all In cells, in the or substantial cleavage of is to that not MST1 cleavage in these cells. activate caspase-9 E. J. K. K. Cell 2001; PubMed Scopus Google which MST1 in (10Graves J.D. Draves K.E. Gotoh Y. Krebs E.G. Clark E.A. J. Biol. Chem. 2001; 276: 14909-14915Abstract Full Text Full Text PDF PubMed Scopus (136) Google Scholar). was in cells expressing We have that MST1 mutants, which of autophosphorylation at Ser327 or were in the but not This that of cleavage of the kinase MST1 is to caspase J.D. Gotoh Y. Draves K.E. Ambrose D. Han D.K. Wright M. Chernoff J. Clark E.A. Krebs E.G. EMBO J. 1998; 17: 2224-2234Crossref PubMed Scopus (325) Google or to caspase activation induced by S. S. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus Google Scholar). In MST1 induced caspase activation, although only in the detached cells expressing MST1. is to that the caspase-insensitive D326N mutant to morphological and caspase cleavage in and of is not a for this the detached cells expressing active MST1 greater caspase controls of caspase activation. substantial effects were the of active MST1 to the of detached cells and the caspase the cell caspase activation appears to be a combined of cell to This and J.D. Gotoh Y. Draves K.E. Ambrose D. Han D.K. Wright M. Chernoff J. Clark E.A. Krebs E.G. EMBO J. 1998; 17: 2224-2234Crossref PubMed Scopus (325) Google have that activated MST1 of cleavage by to JNK and p38 kinases. In and cells, and JNK phosphorylation was and this was with the of MST1 of previously J.D. Gotoh Y. Draves K.E. Ambrose D. Han D.K. Wright M. Chernoff J. Clark E.A. Krebs E.G. EMBO J. 1998; 17: 2224-2234Crossref PubMed Scopus (325) Google was not in the p38 activation with cell and not with MST1 or activity. in this JNK activation appears to be a of MST1 kinase activity, whereas p38 activation to be an to in cell In for the that and the MST1 activation for kinase activation. Phosphorylation at Thr183 possibly is by intermolecular autophosphorylation the MST1 These highly conserved all of the MST kinase family and suggesting for this group of protein kinases. MST1 cell and JNK activation of caspase MST1 is to caspase cleavage at only and when is