Thyrotropin-releasing hormone and GTP activate inositol trisphosphate formation in membranes isolated from rat pituitary cells.
Thyrotropin-releasing hormone and GTP activate inositol trisphosphate formation in membranes isolated from rat pituitary cells.
复制标题
促甲状腺激素释放激素和 GTP 激活从大鼠垂体细胞分离的膜中形成肌醇三磷酸。
DOI：
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发表时间：
1986
影响因子：
4.8
通讯作者：
M. Gershengorn
中科院分区：
文献类型：
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作者：
R. Straub;M. Gershengorn
磷脂酶C促进磷脂酰肌醇4，5-二磷酸(PtdIns(4，5)P2)的水解制得三磷酸肌醇(InsP3)和1，2-二酰甘油，这似乎是包括促甲状腺激素释放激素(TRH)在内的一系列细胞-表面相互作用刺激的信号转导的第一步。从大鼠垂体(GH3)细胞分离的膜悬液中，用[~3H]肌醇预先标记到同位素稳态，并与ATP孵育，聚磷脂酰肌醇(4，5)P2和[~3H]磷脂酰肌醇4-磷酸，以及肌醇多聚磷酸盐[~3H]InsP3和[~3H]肌醇二磷酸在悬浮液中积累。TRH和GTP以时间和浓度依赖的方式刺激[~3H]肌醇多聚磷酸盐的积累，10-30 nM的TRH和3微米的GTP的作用达到一半。GTP的一个非水解性类似物也刺激了[~3H]肌醇多聚磷酸的积累。此外，TRH和GTP共同作用时，其作用大于相加作用。将孵育缓冲液中的游离钙离子浓度固定在20 nM，低于胞浆内的游离钙离子浓度，并不能抑制TRH和GTP刺激的[~3H]肌醇多聚磷酸的积累。ATP是基础和刺激积累[~3H]肌醇多磷酸所必需的，且不能替代ATP的非水解性类似物。这些结果表明，TRH和GTP协同作用刺激InsP3在脑垂体膜悬浮液中的积累，而ATP很可能是合成多磷肌醇的底物，是这一作用所必需的。这些发现表明，鸟嘌呤核苷酸结合的调节蛋白参与了TRH受体与磷脂酶C的偶联，磷脂酶C可以降解PtdIns(4，5)P2。
Stimulation of the hydrolysis of phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2) by a phospholipase C to produce inositol trisphosphate (InsP3) and 1,2-diacylglycerol appears to be the initial step in signal transduction for a number of cell-surface interacting stimuli, including thyrotropin-releasing hormone (TRH). In suspensions of membranes isolated from rat pituitary (GH3) cells that were prelabeled to isotopic steady state with [3H]inositol and incubated with ATP, [3H] PtdIns(4,5)P2, and [3H]phosphatidylinositol 4-phosphate, the polyphosphoinositides, and [3H]InsP3 and [3H]inositol bisphosphate, the inositol polyphosphates, accumulated. TRH and GTP stimulated the accumulation of [3H]inositol polyphosphates in time- and concentration-dependent manners; half-maximal effects occurred with 10-30 nM TRH and with 3 microM GTP. A nonhydrolyzable analog of GTP also stimulated [3H] inositol polyphosphate accumulation. Moreover, when TRH and GTP were added together their effects were more than additive. Fixing the free Ca2+ concentration in the incubation buffer at 20 nM, a value below that present in the cytoplasm in vivo did not inhibit stimulation by TRH and GTP of [3H]inositol polyphosphate accumulation. ATP was necessary for basal and stimulated accumulation of [3H]inositol polyphosphates, and a nonhydrolyzable analog of ATP could not substitute for ATP. These data demonstrate that TRH and GTP act synergistically to stimulate the accumulation of InsP3 in suspensions of pituitary membranes and that ATP, most likely acting as substrate for polyphosphoinositide synthesis, was necessary for this effect. These findings suggest that a guanine nucleotide-binding regulatory protein is involved in coupling the TRH receptor to a phospholipase C that hydrolyzes PtdIns(4,5)P2.