PP2A regulates BMP signalling by interacting with BMP receptor complexes and by dephosphorylating both the C-terminus and the linker region of Smad1

Abstract

Phosphorylation of Smads is a crucial regulatory step in the signal transduction pathway initiated by bone morphogenetic proteins (BMPs). Although the dephosphorylation events terminating the pathway in the nucleus have been characterized, little is known about the dephosphorylation of Smads in the cytoplasm. In a proteomic screen for proteins interacting with the BMP type-II receptor, we found the regulatory Bβ subunit of PP2A. PP2A is one of the major serine/threonine phosphatases involved in cell-cycle regulation and signal transduction. Here, we present data showing that the Bβ subunit of PP2A interacts with both BMP type-I and type-II receptors. Furthermore, we demonstrate that several B subunits can associate with the BMP type-II receptor, independently of the kinase activity of the receptor and the catalytic subunit of PP2A. By contrast, the PP2A catalytic subunit is required for PP2A function at the receptor complex. This function of PP2A is the dephosphorylation of Smad1, mainly in the linker region. PP2A-mediated dephosphorylation of the BMP-Smad linker region leads to increased nuclear translocation of Smads and overall amplification of the BMP signal. Although other phosphatases identified within the BMP pathway are all shown to inhibit signalling, PP2A is the first example for a signalling stimulatory phosphatase within this pathway.

Novel Protein Interactions with Endoglin and Activin Receptor-like Kinase 1: Potential Role in Vascular Networks

“…In vertebrates, the PPP2R2A and PPP2R2D subunits were shown to regulate TGF-␤/Activin/Nodal signaling in opposite ways (46), PPP2R2A enhancing signals by stabilizing ACVR1B and TGFBR1 basal levels, whereas PPP2R2D restricted receptor activity. Our data imply that the PPP2R2B subunit interacts preferentially with ACVRL1, and independently of its kinase activity, as previously reported for BMPR2 (47). We also confirm that BMPR2 and TGFBR2 associate with PPP2R2B and PPP2R2D.…”

Hypoxia-activated Smad3-specific Dephosphorylation by PP2A

“…BMP and TGF-␤ signaling are regulated by SCP1-3 phosphatases, which target the C-terminal phosphorylation of Smad1 and the linker phosphorylation of Smad2 and -3 (38,39). BMP receptor associates with one form of PP2A leading to dephosphorylation of BMP R-Smad, Smad1, in normoxia, and different PP2A forms can dephosphorylate different receptors of the TGF-␤ superfamily and Smad1 (11,12). Bengtson et al (12) show that PP2A dephosphorylates Smad1 linker region and only weakly the C terminus in normoxia.…”

“…BMP receptor associates with one form of PP2A leading to dephosphorylation of BMP R-Smad, Smad1, in normoxia, and different PP2A forms can dephosphorylate different receptors of the TGF-␤ superfamily and Smad1 (11,12). Bengtson et al (12) show that PP2A dephosphorylates Smad1 linker region and only weakly the C terminus in normoxia. They also reported that the linker region dephosphorylation leads to enhanced nuclear localization of Smad1 in normoxia.…”

“…Luciferase Assay-For reporter gene assay, HaCaT cells stably transfected with Smad3-specific luciferase reporter gene (CAGA 12 -Luc) and a constantly active TK-Renilla gene were used. The assay was performed with Dual-Luciferase reporter assay system (E1910, Promega) according to the manufacturer's protocol and measured with VICTOR TM 2 plate reader (PerkinElmer Life Sciences).…”

“…Smad7 has also been shown to mediate PP1C (protein phosphatase 1C) binding to the activated TGF-␤RI and subsequent inactivation of the receptor by dephosphorylation (10). Independently from I-Smads, PP2A can inhibit the TGF-␤ superfamily receptors and BMP R-Smad (11,12). Moreover, Smad2 and -3 activity may be attenuated by a negative feedback mechanism independently from the TGF-␤RI activity through dephosphorylation of the C-terminal serines by the PPM1A (PP2C) phosphatase (13).…”