Phosphorylation of Mnk1 by caspase-activated Pak2/gamma-PAK inhibits phosphorylation and interaction of eIF4G with Mnk.
The mitogen-activated protein kinase-interacting kinase 1 (Mnk1) is phosphorylated by caspase-cleaved protein kinase Pak2/gamma-PAK but not by Cdc42-activated Pak2. Phosphorylation of Mnk1 is rapid, reaching 1 mol/mol within 15 min of incubation with Pak2. A kinetic analysis of the phosphorylation of Mnk1 by Pak2 yields a K(m) of 0.6 microm and a V(max) of 14.9 pmol of (32)P/min/microg of Pak2. Two-dimensional tryptic phosphopeptide mapping of Mnk1 phosphorylated by Pak2 yields two distinct phosphopeptides. Analysis of the phosphopeptides by automated microsequencing and manual Edman degradation identified the sites in Mnk1 as Thr(22) and Ser(27). Mnk1, activated by phosphorylation with Erk2, phosphorylates the eukaryotic initiation factor (eIF) 4E and the eIF4G components of eIF4F. Phosphorylation of Mnk1 by Pak2 does not activate Mnk1, as measured with either eIF4E or eIF4F as substrate. Phosphorylation of Erk2-activated Mnk1 by Pak2 has no effect on phosphorylation of eIF4E but reduces phosphorylation of eIF4G by Mnk1 by up to 50%. Phosphorylation of Mnk1 by Pak2 inhibits binding of eIF4G peptides containing the Mnk1 binding site by up to 80%. When 293T cells are subjected to apoptotic induction by hydrogen peroxide, Mnk1 is phosphorylated at both Thr(22) and Ser(27). These results indicate a role for Pak2 in the down-regulation of translation initiation in apoptosis by phosphorylation of Mnk1.
Orton
Kevin C
KC
Department of Biochemistry, University of California, Riverside, Riverside, California 92521, USA.
Ling
Jun
J
Waskiewicz
Andrew J
AJ
Cooper
Jonathan A
JA
Merrick
William C
WC
Korneeva
Nadejda L
NL
Rhoads
Robert E
RE
Sonenberg
Nahum
N
Traugh
Jolinda A
JA
D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis; D001665:Binding Sites; D020169:Caspases / Q000378:metabolism*; D002460:Cell Line; D015536:Down-Regulation; D015180:Electrophoresis, Gel, Two-Dimensional; D004789:Enzyme Activation; D039561:Eukaryotic Initiation Factor-4E / Q000378:metabolism; D039562:Eukaryotic Initiation Factor-4F / Q000378:metabolism; D039603:Eukaryotic Initiation Factor-4G / Q000378:metabolism*; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D007313:Insecta; D047908:Intracellular Signaling Peptides and Proteins; D007700:Kinetics; D051379:Mice; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry; D010766:Phosphorylation; D011485:Protein Binding; D014176:Protein Biosynthesis; D020033:Protein Isoforms; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D011817:Rabbits; D012694:Serine / Q000737:chemistry; D013912:Threonine / Q000737:chemistry; D013997:Time Factors; D014357:Trypsin / Q000494:pharmacology; D020764:cdc42 GTP-Binding Protein / Q000378:metabolism; D054462:p21-Activated Kinases
D039561:Eukaryotic Initiation Factor-4E; D039562:Eukaryotic Initiation Factor-4F; D039603:Eukaryotic Initiation Factor-4G; D047908:Intracellular Signaling Peptides and Proteins; D010455:Peptides; D020033:Protein Isoforms; D013912:Threonine; D012694:Serine; D006861:Hydrogen Peroxide; D005982:Glutathione Transferase; C491395:MKNK1 protein, human; C106281:Mknk1 protein, mouse; C515907:PAK2 protein, human; C515909:Pak2 protein, mouse; D017346:Protein Serine-Threonine Kinases; D054462:p21-Activated Kinases; D014357:Trypsin; D020169:Caspases; D020764:cdc42 GTP-Binding Protein