Protein Kinase A (PKA) Phosphorylation of Shp2 Protein Inhibits Its Phosphatase Activity and Modulates Ligand Specificity.
Protein Kinase A (PKA) Phosphorylation of Shp2 Protein Inhibits Its Phosphatase Activity and Modulates Ligand Specificity.
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DOI：
10.1074/jbc.m115.642983
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发表时间：
2015-05-08
期刊：
影响因子：
--
通讯作者：
Carnegie GK
中科院分区：
文献类型：
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作者：
Burmeister BT;Wang L;Gold MG;Skidgel RA;O'Bryan JP;Carnegie GK
背景：AKAP-Lbc支架协调心脏肥大信号传导。结果：PKA磷酸化的Shp 2在Thr-73/Ser-189抑制酪氨酸磷酸化配体结合和PTP活性。结论：AKAP-Lbc整合PKA和Shp 2信号传导，慢性β-肾上腺素能刺激后肥厚心脏中AKAP-Lbc相关的Shp 2活性降低。意义：PKA抑制AKAP-Lbc锚定的Shp 2活性是一种以前未被认识到的可能促进心脏肥大的机制。病理性心脏肥大（由应激诱导的心肌细胞生长导致的心脏质量增加）是导致心力衰竭的主要因素。含Src同源2结构域的磷酸酶（Shp 2）对心脏功能至关重要，因为导致Shp 2催化活性丧失的突变与先天性心脏缺陷和肥大相关。我们确定了一种新的机制，Shp 2抑制，可能会促进心脏肥大。我们证明，Shp 2是蛋白激酶A锚定蛋白（AKAP）-Lbc复合物的一个组成部分。AKAP-Lbc促进Shp 2的PKA磷酸化，其抑制Shp 2磷酸酶活性。我们确定了两个关键的氨基酸在Shp 2的磷酸化PKA。Thr-73在Shp 2的N-末端SH 2结构域内为螺旋αB提供螺旋帽，而Ser-189在C-末端SH 2结构域内占据等同位置。利用双突变PKA磷酸缺陷（T73 A/S189 A）和磷酸模拟（T73 D/S189 D）的结构，在体外结合试验，磷酸酶活性测定，我们证明，这些残基的磷酸化破坏Shp 2与酪氨酸磷酸化配体的相互作用，并抑制其蛋白酪氨酸磷酸酶活性。总之，我们的数据表明AKAP-Lbc整合了心脏中的PKA和Shp 2信号传导，并且AKAP-Lbc相关的Shp 2活性在肥大心脏中响应于慢性β-肾上腺素能刺激和PKA活化而降低。因此，虽然诱导心脏肥大是一个多方面的过程，但通过AKAP-Lbc锚定PKA抑制Shp 2活性是一种以前未认识到的机制，可能会促进这种代偿反应。
Background: The AKAP-Lbc scaffold coordinates cardiac hypertrophic signaling. Results: PKA phosphorylation of Shp2 at Thr-73/Ser-189 inhibits tyrosine-phosphorylated ligand binding and PTP activity. Conclusion: AKAP-Lbc integrates PKA and Shp2 signaling, and AKAP-Lbc-associated Shp2 activity is reduced in hypertrophic hearts following chronic β-adrenergic stimulation. Significance: PKA inhibition of AKAP-Lbc-anchored Shp2 activity is a previously unrecognized mechanism that may promote cardiac hypertrophy. Pathological cardiac hypertrophy (an increase in cardiac mass resulting from stress-induced cardiac myocyte growth) is a major factor underlying heart failure. Src homology 2 domain-containing phosphatase (Shp2) is critical for cardiac function because mutations resulting in loss of Shp2 catalytic activity are associated with congenital cardiac defects and hypertrophy. We identified a novel mechanism of Shp2 inhibition that may promote cardiac hypertrophy. We demonstrate that Shp2 is a component of the protein kinase A anchoring protein (AKAP)-Lbc complex. AKAP-Lbc facilitates PKA phosphorylation of Shp2, which inhibits Shp2 phosphatase activity. We identified two key amino acids in Shp2 that are phosphorylated by PKA. Thr-73 contributes a helix cap to helix αB within the N-terminal SH2 domain of Shp2, whereas Ser-189 occupies an equivalent position within the C-terminal SH2 domain. Utilizing double mutant PKA phosphodeficient (T73A/S189A) and phosphomimetic (T73D/S189D) constructs, in vitro binding assays, and phosphatase activity assays, we demonstrate that phosphorylation of these residues disrupts Shp2 interaction with tyrosine-phosphorylated ligands and inhibits its protein-tyrosine phosphatase activity. Overall, our data indicate that AKAP-Lbc integrates PKA and Shp2 signaling in the heart and that AKAP-Lbc-associated Shp2 activity is reduced in hypertrophic hearts in response to chronic β-adrenergic stimulation and PKA activation. Therefore, although induction of cardiac hypertrophy is a multifaceted process, inhibition of Shp2 activity through AKAP-Lbc-anchored PKA is a previously unrecognized mechanism that may promote this compensatory response.