Dephosphorylation of the linker regions of Smad1 and Smad2/3 by small C-terminal domain phosphatases has distinct outcomes for bone morphogenetic protein and transforming growth factor-beta pathways.
Smad proteins transduce bone morphogenetic protein (BMP) and transforming growth factor-beta (TGFbeta) signals upon phosphorylation of their C-terminal SXS motif by receptor kinases. The activity of Smad1 in the BMP pathway and Smad2/3 in the TGFbeta pathway is restricted by pathway cross-talk and feedback through protein kinases, including MAPK, CDK2/4, p38MAPK, JNK, and others. These kinases phosphorylate Smads 1-3 at the region that links the N-terminal DNA-binding domain and the C-terminal transcriptional domain. Phosphatases that dephosphorylate the linker region are therefore likely to play an integral part in the regulation of Smad activity. We reported previously that small C-terminal domain phosphatases 1, 2, and 3 (SCP1-3) dephosphorylate Smad1 C-terminal tail, thereby attenuating BMP signaling. Here we provide evidence that SCP1-3 also dephosphorylate the linker regions of Smad1 and Smad2/3 in vitro, in mammalian cells and in Xenopus embryos. Overexpression of SCP 1, 2, or 3 decreased linker phosphorylation of Smads 1, 2 and 3. Moreover, RNA interference-mediated knockdown of SCP1/2 increased the BMP-dependent phosphorylation of the Smad1 linker region as well as the C terminus. In contrast, SCP1/2 knockdown increased the TGFbeta-dependent linker phosphorylation of Smad2/3 but not the C-terminal phosphorylation. Consequently, SCP1/2 knockdown inhibited TGFbeta transcriptional responses, but it enhanced BMP transcriptional responses. Thus, by dephosphorylating Smad2/3 at the linker (inhibitory) but not the C-terminal (activating) site, the SCPs enhance TGFbeta signaling, and by dephosphorylating Smad1 at both sites, the SCPs reset Smad1 to the basal unphosphorylated state.
Sapkota
Gopal
G
Cancer Biology and Genetics Program, Howard Hughes Medical Institute, Memorial Sloan-Kettering Cancer Center, New York, New York 10021, USA.
Knockaert
Marie
M
Alarcón
Claudio
C
Montalvo
Ermelinda
E
Brivanlou
Ali H
AH
Massagué
Joan
J
D000818:Animals; D019485:Bone Morphogenetic Proteins / Q000502:physiology*; D002460:Cell Line; D045744:Cell Line, Tumor; D006801:Humans; D009687:Nuclear Proteins / Q000502:physiology*; D010446:Peptide Fragments / Q000502:physiology; D010749:Phosphoprotein Phosphatases / Q000502:physiology*; D010766:Phosphorylation; D017434:Protein Structure, Tertiary / Q000502:physiology; D015398:Signal Transduction / Q000502:physiology*; D051898:Smad1 Protein / Q000378:metabolism*; D051899:Smad2 Protein / Q000378:metabolism*; D051900:Smad3 Protein / Q000378:metabolism*; D053773:Transforming Growth Factor beta1 / Q000502:physiology*; D025521:Tumor Suppressor Proteins / Q000502:physiology; D014982:Xenopus laevis
D019485:Bone Morphogenetic Proteins; C486483:CTDSPL protein, human; D009687:Nuclear Proteins; D010446:Peptide Fragments; C497399:SMAD1 protein, human; C497782:SMAD2 protein, human; C497790:SMAD3 protein, human; D051898:Smad1 Protein; D051899:Smad2 Protein; D051900:Smad3 Protein; D053773:Transforming Growth Factor beta1; D025521:Tumor Suppressor Proteins; C413483:CTDSP1 protein, human; C108580:CTDSP2 protein, human; D010749:Phosphoprotein Phosphatases