Ras is an indispensable coregulator of the class IB phosphoinositide 3-kinase p87/p110γ
Ras is an indispensable coregulator of the class IB phosphoinositide 3-kinase p87/p110γ
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DOI：
10.1073/pnas.0905506106
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发表时间：
2009-12-01
影响因子：
11.1
通讯作者：
Nuernberg, Bernd
中科院分区：
文献类型：
--
作者：
Kurig, Barbara;Shymanets, Aliaksei;Nuernberg, Bernd
I-B类磷脂酰肌醇3-激酶γ（PI 3 K γ）引起各种免疫和心血管反应;然而，这种信号异质性的分子基础尚不清楚。PI 3 K γ由催化性p110 γ和调节性p87（PIKAP）（p87，也称为p84）或p101亚基组成。通常认为，高表达p87和p101在受体诱导的和G蛋白β γ（G β γ）介导的PI 3 K γ调节中表现出冗余功能。在这里，我们研究了受体依赖性p87/p110 γ激活的分子机制。通过分析HEK 293细胞、p110 γ（-/-）小鼠骨髓来源的肥大细胞（BMMCs）中表达的GFP标记蛋白以及重组于脂质囊泡的纯化重组蛋白，我们阐明了p87依赖的G蛋白偶联受体（GPCR）诱导的PI 3 K γ激活的新途径。尽管p101与G β γ强烈相互作用，从而介导PI 3 K γ膜募集和刺激，但p87仅表现出弱相互作用，导致适度的激酶活化和缺乏膜募集。令人惊讶的是，Ras-GTP通过与p110 γ的直接相互作用取代了p87/p110 γ缺失的G β γ依赖性膜募集，这表明Ras对于p87/p110 γ的激活是不可或缺的。因此，Ras信号的干扰确实选择性地阻断p87/p110 γ，但不是p101/p110 γ，激酶活性在HEK 293和BMMC细胞，揭示了一个重要的串扰之间的单体和三聚体G蛋白的p87/p110 γ激活。我们的数据显示了p87和p101的不同信号需求，赋予PI 3 K γ信号特异性，这可能为治疗干预开辟新的可能性。
Class I-B phosphoinositide 3-kinase gamma (PI3K gamma) elicits various immunologic and cardiovascular responses; however, the molecular basis for this signal heterogeneity is unclear. PI3K gamma consists of a catalytic p110 gamma and a regulatory p87(PIKAP) (p87, also p84) or p101 subunit. Hitherto p87 and p101 are generally assumed to exhibit redundant functions in receptor-induced and G protein beta gamma (G beta gamma)-mediated PI3K gamma regulation. Here we investigated the molecular mechanism for receptor-dependent p87/p110 gamma activation. By analyzing GFP-tagged proteins expressed in HEK293 cells, PI3K gamma-complemented bone marrow-derived mast cells (BMMCs) from p110 gamma(-/-) mice, and purified recombinant proteins reconstituted to lipid vesicles, we elucidated a novel pathway of p87-dependent, G protein-coupled receptor (GPCR)-induced PI3K gamma activation. Although p101 strongly interacted with G beta gamma, thereby mediating PI3K gamma membrane recruitment and stimulation, p87 exhibited only a weak interaction, resulting in modest kinase activation and lack of membrane recruitment. Surprisingly, Ras-GTP substituted the missing G beta gamma-dependent membrane recruitment of p87/p110 gamma by direct interaction with p110 gamma, suggesting the indispensability of Ras for activation of p87/p110 gamma. Consequently, interference with Ras signaling indeed selectively blocked p87/p110 gamma, but not p101/p110 gamma, kinase activity in HEK293 and BMMC cells, revealing an important crosstalk between monomeric and trimeric G proteins for p87/p110 gamma activation. Our data display distinct signaling requirements of p87 and p101, conferring signaling specificity to PI3K gamma that could open up new possibilities for therapeutic intervention.