TY - JOUR
T1 - PLCγ-dependent mTOR signalling controls IL-7-mediated early B cell development
AU - Yu, Mei
AU - Chen, Yuhong
AU - Zeng, Hu
AU - Zheng, Yongwei
AU - Fu, Guoping
AU - Zhu, Wen
AU - Broeckel, Ulrich
AU - Aggarwal, Praful
AU - Turner, Amy
AU - Neale, Geoffrey
AU - Guy, Cliff
AU - Zhu, Nan
AU - Chi, Hongbo
AU - Wen, Renren
AU - Wang, Demin
N1 - Funding Information:
Pro-B cells are highly responsive to IL-7 stimulation6, 7, 10, 11. Rag deficiency arrests B cell development at the pro-B cell stage. Rag-deficient pro-B cells express IL-7R, can be readily expanded in vitro, and are particularly suitable for the study of IL-7-mediated signaling and functions. Inhibition of the PLCγ pathway reduced IL-7-mediated oxygen consumption (oxidative phosphorylation) and extracellular acidification (glycolysis) in unexpanded or in vitro-expanded B cell progenitors, demonstrating a critical role of the PLCγ pathway in controlling IL-7-regulated cellular metabolism. Cell proliferation is supported by cellular metabolism, which is controlled by the mTOR signaling pathway52, 53, 61. Importantly, we discovered that the PLCγ pathway-regulated IL-7-induced activation of mTOR. Typically, activation of mTOR depends on the PI3K/Akt/Tsc/Rheb pathway53, 61, 62. Activation of the lipid kinase PI3K leads to production of PIP3 that recruits Akt to the plasma membrane, resulting in Akt activation through its dual phosphorylation by PDK1 and mTORC2. Activated Akt phosphorylates Tsc2, leading to functional inactivation of the Rheb GAP Tsc1/2 complex, and subsequent elevation of GTP-bound Rheb and activation of mTOR. However, Rheb deficiency had no effect on IL-7-induced mTOR activation in B cell progenitors, but disrupted BCR-induced mTOR activation in peripheral B cells, demonstrating that IL-7 uniquely activates mTOR through the PLCγ pathway independently of the conventional Akt/Tsc/Rheb pathway. In addition, inhibition of AKT blocks BCR-induced but not IL-7-induced mTOR activation. Consistently, single or double deficiency of Akt1 and Akt2 has no effect on IL-7-mediated early B cell development63. However, deficiency of PDK1 inhibits IL-7-mediated proliferation of B cell progenitors, which could be due to PDK1 targeting Akt and other signaling molecules, such as PKC64. PLCγ hydrolyzes PIP2 to generate DAG and IP3 that induces PKC activation and intracellular Ca2+ release, respectively26, 27. Inhibition of PKC but not chelation of intracellular calcium markedly decreased IL-7-induced mTOR activation. In addition, PKC agonist but not calcium ionophore markedly activated mTOR. Our data demonstrate that PKC, but not Ca2+-dependent downstream pathway of PLCγ has a central role in activating mTOR. Of note, PKC agonist activated mTOR without Akt activation in pro-B cells, supporting the notion that the PLCγ pathway activates mTOR independently of the Akt/Tsc/ Rheb pathway. Consistently, previous studies have shown that PKC is able to activate mTOR in mouse epithelial cells and glioma independently of Akt65, 66. Of note, the Ca2+-dependent pathway is required for classical PKC activation58, and thus this pathway may also contribute to mTOR activation through PKC. In fact, we found here that PKC agonist plus calcium further induced mTOR activation. These results demonstrate the PKC-dependent downstream pathway of PLCγ plays a major role in mTOR activation, whereas the Ca2+-dependent downstream pathway contributes to it. The importance of the PLCγ/PKC/ mTOR pathway in controlling early B cell development is supported by studies of mice with genetic disruption of this pathway via Raptor deletion67, 68. Raptor deficiency impairs early B cell development, leading to a marked reduction in peripheral B cells67, 68. Consistent with our current findings, B cell-specific Raptor deletion blocks early B cell development at the pre-pro-B cell stage, resulting in a decrease in early pro-B (fraction B)67.
Publisher Copyright:
© 2017 The Author(s).
PY - 2017/12/1
Y1 - 2017/12/1
N2 - The precise molecular mechanism underlying the regulation of early B cell lymphopoiesis is unclear. The PLCγ signaling pathway is critical for antigen receptor-mediated lymphocyte activation, but its function in cytokine signaling is unknown. Here we show that PLCγ1/PLCγ2 double deficiency in mice blocks early B cell development at the pre-pro-B cell stage and renders B cell progenitors unresponsive to IL-7. PLCγ pathway inhibition blocks IL-7-induced activation of mTOR, but not Stat5. The PLCγ pathway activates mTOR through the DAG/PKC signaling branch, independent of the conventional Akt/TSC/Rheb signaling axis. Inhibition of PLCγ/PKC-induced mTOR activation impairs IL-7-mediated B cell development. PLCγ1/PLCγ2 double-deficient B cell progenitors have reduced expression of genes related to B cell lineage, IL-7 signaling, and cell cycle. Thus, IL-7 receptor controls early B lymphopoiesis through activation of mTOR via PLCγ/DAG/PKC signaling, not via Akt/Rheb signaling.
AB - The precise molecular mechanism underlying the regulation of early B cell lymphopoiesis is unclear. The PLCγ signaling pathway is critical for antigen receptor-mediated lymphocyte activation, but its function in cytokine signaling is unknown. Here we show that PLCγ1/PLCγ2 double deficiency in mice blocks early B cell development at the pre-pro-B cell stage and renders B cell progenitors unresponsive to IL-7. PLCγ pathway inhibition blocks IL-7-induced activation of mTOR, but not Stat5. The PLCγ pathway activates mTOR through the DAG/PKC signaling branch, independent of the conventional Akt/TSC/Rheb signaling axis. Inhibition of PLCγ/PKC-induced mTOR activation impairs IL-7-mediated B cell development. PLCγ1/PLCγ2 double-deficient B cell progenitors have reduced expression of genes related to B cell lineage, IL-7 signaling, and cell cycle. Thus, IL-7 receptor controls early B lymphopoiesis through activation of mTOR via PLCγ/DAG/PKC signaling, not via Akt/Rheb signaling.
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U2 - 10.1038/s41467-017-01388-5
DO - 10.1038/s41467-017-01388-5
M3 - Article
C2 - 29133930
AN - SCOPUS:85034212045
VL - 8
JO - Nature Communications
JF - Nature Communications
SN - 2041-1723
IS - 1
M1 - 1457
ER -