The transferrin receptor CD71 regulates type II CD38, revealing tight topological compartmentalization of intracellular cyclic ADP-ribose production
The transferrin receptor CD71 regulates type II CD38, revealing tight topological compartmentalization of intracellular cyclic ADP-ribose production
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转铁蛋白受体 CD71 调节 II 型 CD38，揭示细胞内环状 ADP-核糖产生的紧密拓扑区隔
DOI：
10.1074/jbc.ra119.010010
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发表时间：
2019-10-18
影响因子：
4.8
通讯作者：
Zhao, Yong Juan
中科院分区：
文献类型：
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作者：
Deng, Qi Wen;Zhang, Jingzi;Zhao, Yong Juan
CD38分子（CD38）催化了钙的钙化信使循环ADP-核糖（CADPR）的生物发生。 CD38具有双膜取向，III型CD38及其催化结构域面向细胞质，具有较低的丰度，但有效地循环胞质NAD产生CADPR。细胞表面II型CD38在细胞CADPR产生中的作用尚不清楚。在这里，我们调制了II型CD38表达，并评估了该调制对CADPR水平的影响。我们开发了一种基于CD38纳米病的光活化交联探针，并将其与MS分析结合在一起，发现细胞表面CD38与CD71相互作用。 CD71敲低的CD38水平增加了，CD38敲除偶数增加了CD71，并且都可以被粘结并共免疫沉淀。我们构建了一个嵌合体，该嵌合体包括CD71的N末端片段和一个CD38纳米型，以模拟CD71的配体性质。该嵌合体的过表达通过溶酶体引起CD38的大幅度降低。值得注意的是，细胞CADPR水平并未相应降低。 Bafilomycin介导的溶酶体降解的阻断通过将其捕获到溶酶体中，从而大大升高了活性II型CD38，但也没有增加CADPR水平。通过表达ER构建构建体，以防止其转运到细胞表面的ER构建体在内质网（ER）中保留II型CD38。这些结果提供了首先和直接的证据，表明CADPR生物发生发生在细胞质中，主要由III型CD38催化，而在ER或溶酶体或细胞表面上划分的II型CD38仅对CADPR生物发生贡献。
The CD38 molecule (CD38) catalyzes biogenesis of the calcium-mobilizing messenger cyclic ADP-ribose (cADPR). CD38 has dual membrane orientations, and type III CD38, with its catalytic domain facing the cytosol, has low abundance but is efficient in cyclizing cytosolic NAD to produce cADPR. The role of cell surface type II CD38 in cellular cADPR production is unknown. Here we modulated type II CD38 expression and assessed the effects of this modulation on cADPR levels. We developed a photoactivatable cross-linking probe based on a CD38 nanobody, and, combining it with MS analysis, we discovered that cell surface CD38 interacts with CD71. CD71 knockdown increased CD38 levels, and CD38 knockout reciprocally increased CD71, and both could be cocapped and coimmunoprecipitated. We constructed a chimera comprising the N-terminal segment of CD71 and a CD38 nanobody to mimic CD71's ligand property. Overexpression of this chimera induced a dramatically large decrease in CD38 via lysosomes. Remarkably, cellular cADPR levels did not decrease correspondingly. Bafilomycin-mediated blockade of lysosomal degradation greatly elevated active type II CD38 by trapping it in the lysosomes but also did not increase cADPR levels. Retention of type II CD38 in the endoplasmic reticulum (ER) by expressing an ER construct that prevented its transport to the cell surface likewise did not change cADPR levels. These results provide first and direct evidence that cADPR biogenesis occurs in the cytosol and is catalyzed mainly by type III CD38 and that type II CD38, compartmentalized in the ER or lysosomes or on the cell surface, contributes only minimally to cADPR biogenesis.