A-Kinase Anchoring Proteins Interact with Phosphodiesterases in T Lymphocyte Cell Lines

Abstract

The cAMP protein kinase A (PKA) pathway in T cells conveys an inhibitory signal to suppress inflammation. This study was performed to understand the mechanisms involved in cAMP-mediated signaling in T lymphocytes. A-kinase anchoring proteins (AKAPs) bind and target PKA to various subcellular locations. AKAPs also bind other signaling molecules such as cyclic nucleotide phosphodiesterases (PDEs) that hydrolyze cAMP in the cell. PDE4 and PDE7 have important roles in T cell activation. Based on this information, we hypothesized that AKAPs associate with PDEs in T lymphocytes. Immunoprecipitation of Jurkat cell lysates with Abs against both the regulatory subunit of PKA (RIIα) and specific AKAPs resulted in increased PDE activity associated with RIIα and AKAP95, AKAP149, and myeloid translocation gene (MTG) compared with control (IgG). Immunoprecipitation and pull-down analyses demonstrate that PDE4A binds to AKAP149, AKAP95, and MTG, but not AKAP79, whereas PDE7A was found to bind only MTG. Further analysis of MTG/PDE association illustrated that PDE4A and PDE7A bind residues 1–344 of MTG16b. Confocal analysis of HuT 78 cells stained with anti-PDE7A showed overlapping staining patterns with the Golgi marker GM130, suggesting that PDE7A is located in the Golgi. The staining pattern of PDE7A also showed similarity to the staining pattern of MTG, supporting the immunoprecipitation data and suggesting that MTG may interact with PDE7A in the Golgi. In summary, these data suggest that AKAPs interact with both PKA and PDE in T lymphocytes and thus are a key component of the signaling complex regulating T cell activation.

Differential Expression and Function of Phosphodiesterase 4 (PDE4) Subtypes in Human Primary CD4+ T Cells: Predominant Role of PDE4D

“…Additionally, Tasken and coworkers (31) confirmed in human primary T cells that PKA-dependent phosphorylation increases PDE4 activity, a mechanism that was already described in several cell systems (42, 48 -51). The well-characterized interactions of PDE4 isoforms with A-kinase anchoring proteins were also found in T cell lines, enabling a local feedback regulation as control for cAMP concentration (52).…”

AKAP signaling complexes: getting to the heart of the matter

“…PDE4D3 has been detected in the AKAP350 (AKAP450-Yotiao) complex at the centrosomal area of Sertoli cells [35], and other members of the PDE family have been shown to associate with AKAP complexes [21]. For example, in T cells, PDE4A is associated with several AKAPs, including AKAP95, AKAP149 and MTG16b, which has also been shown to target another PDE, PDE7A [21]. Although the functional implications of these interactions have not been fully investigated, the activity of these PDEs might be controlled by AKAPassociated kinases and phosphatases.…”

“…Importantly, the PDE4D3 isozyme has been shown to be part of signaling complexes that target PKA. These two cAMPresponsive enzymes (PDE and PKA) are found in complex with a growing number of AKAPs [20,21], thereby enabling tight regulation of cAMP.…”

Mammalian Cyclic Nucleotide Phosphodiesterases: Molecular Mechanisms and Physiological Functions

“…AKAP450, which anchors to the Golgi, interacts with PDE4A and AKAP7 and forms a complex with PDE4D to regulate osmotic water permeability (358). PDE7A has also been shown to interact with an AKAP, MTG (myeloid translocation gene) in the Golgi to form a regulatory module in the T cell (12). PDE3A has been shown to complex with another AKAP, BIG (BFA-inhibited GEP), which likely contributes to the cAMP-mediated regulation of ADP-ribosylation factors (293).…”