The interactome of the amyloid β precursor protein family members is shaped by phosphorylation of their intracellular domains

<p>Abstract</p> <p>Background</p> <p>Brain tissue from patients with Alzheimer's disease has shown an increase of phosphorylation of Tyr-682, located on the conserved Y682ENPTY motif, and Thr-668 residues, both in the intracellular domain (AID) of amyloid β precursor protein (APP), although the role of these two residues is not yet known.</p> <p>Results</p> <p>Here, we report that the phosphorylation status of Tyr-682, and in some cases Thr-668, shapes the APP interactome. It creates a docking site for SH2-domain containing proteins, such as ShcA, ShcB, ShcC, Grb7, Grb2, as well as adapter proteins, such as Crk and Nck, that regulate important biological processes, cytosolic tyrosine kinases, such as Abl, Lyn and Src, which regulate signal transduction pathways, and enzymes that control phosphatidylinositols levels and signaling, such as PLC-γ. At the same time, it either reduces (like for JIP1, NUMB, NUMBL and ARH) or abolishes (like for Fe65, Fe65L1 and Fe65L2) binding of other APP interactors. Phosphorylation of Thr-668, unlike Tyr-682, does not seem to affect APP's ability to interact with the various proteins, with Pin1 and X11 being the exclusions. We also found that there are some differences between the interactions to AID and to ALID1 and ALID2, its two homologues.</p> <p>Conclusion</p> <p>Our data indicates that APP can regulate diverse cellular processes and that, vice versa, a network of signaling events can impact APP processing. Our results also suggest that phosphorylation of the APP Intracellular Domain will dramatically shape the APP interactome and, consequently, will regulate APP processing, APP transport and APP/AID-mediated functions.</p

The Mst1 and Mst2 kinases control activation of rho family GTPases and thymic egress of mature thymocytes

The Mst1 kinase is an important regulator of murine T cell adhesion, migration, proliferation, and apoptosis. In this study, we analyze mice lacking both Mst1 and Mst2 in hematopoietic cells. Compared with wild-type mice, these double knockout (DKO) mice exhibit a severe reduction in the number of mature T cells in the circulation and in secondary lymphoid organs (SLOs). CD4+CD8− and CD4−CD8+ single-positive (SP) thymocytes in DKO mice resemble mature T cells of wild-type mice but undergo excessive apoptosis, and their egress from the thymus is reduced by >90%. Even when placed directly in the circulation, DKO SP thymocytes failed to enter SLOs. In SP thymocytes, deficiency of Mst1 and Mst2 abolished sphingosine-1 phosphate– and CCL21-induced Mob1 phosphorylation, Rac1 and RhoA GTP charging, and subsequent cell migration. When phosphorylated by Mst1 or Mst2, Mob1 binds and activates the Rac1 guanyl nucleotide exchanger Dock8, which is abundant in the thymus. Thus, the Mst1 and Mst2 kinases control Rho GTPase activation and the migratory responses of SP thymocytes

Growth Factor Receptor-bound Protein 2 Interaction with the Tyrosine-phosphorylated Tail of Amyloid β Precursor Protein Is Mediated by Its Src Homology 2 Domain

The sequential processing of the familial disease gene product amyloid beta precursor protein (AbetaPP) by beta- and gamma-secretases generates amyloid beta, which is considered to be the pathogenic factor of Alzheimer's disease, and the AID peptide (AbetaPP intracellular domain). The AID peptide acts as a positive regulator of apoptosis and modulates transcription and calcium release. To gain clues about the molecular mechanisms regulating the function of AbetaPP and AID, proteins interacting with the AID region of AbetaPP have been isolated using the yeast two-hybrid system. Recent evidence indicates that AbetaPP undergoes post-translational modification events in the AID region and that phosphorylation might regulate its affinity for interacting proteins. To test this possibility and to uncover AbetaPP-binding partners whose interaction depends on AbetaPP phosphorylation, we used a proteomic approach. Here we describe a protein, growth factor receptor-bound protein 2 (Grb2), that specifically binds AbetaPP, phosphorylated in Tyr(682). Furthermore, we show that this interaction is direct and that Grb2 binds to phospho-AbetaPP via its Src homology 2 region. Together with the evidence that Grb2 is in complex with AbetaPP in human brains and that these complexes are augmented in brains from Alzheimer's cases, our data indicate that Grb2 may mediate some biological and possibly pathological AbetaPP-AID function

Hyperphosphorylation of JNK-interacting Protein 1, a Protein Associated with Alzheimer Disease

The c-Jun N-terminal kinase (JNK) group of mitogen-activated protein (MAP) kinases are activated by pleiotropic signals including environmental stresses, growth factors, and hormones. JNK-interacting protein 1 (JIP1) is a scaffold protein that assembles and facilitates the activation of the mixed lineage kinase-dependent JNK module and also establishes an interaction with beta-amyloid precursor protein that has been partially characterized. Here we show that, similarly to other proteins involved in various neurological diseases, JIP1 becomes hyperphosphorylated following activation of stress-activated and MAP kinases. By immobilized metal affinity chromatography and a combined microcapillary LC/MALDI-TOF/ESI-ion trap mass spectrometry approach, we identified 35 sites of mitotic phosphorylation within JIP1, among which eight were present within (Ser/Thr)-Pro sequence. This motif is modified by various kinases in aggregates of the microtubule-associated protein tau, which generates typical intraneuronal lesions occurring in Alzheimer disease. Most of the post-translational modifications found were located within the JNK, MAP kinase kinase, and RAC-alpha Ser/Thr protein kinase binding regions; no modifications occurred in protein Src homology 3 and phosphotyrosine interaction domains, which are essential for binding to kinesin, beta-amyloid precursor protein, and MAP kinase kinase kinase. Protein phosphorylation is known to affect stability and protein-protein interactions. Thus, the findings that JIP1 is extensively phosphorylated after activation of stress-activated and MAP kinases indicate that these signaling pathways might modulate JIP1 signaling by regulating its stability and association with some, but not all, interacting proteins

The Mst1 Kinase Is Required for Follicular B Cell Homing and B-1 B Cell Development

The Mst1 and 2 cytosolic serine/threonine protein kinases are the mammalian orthologs of the Drosophila Hippo protein. Mst1 has been shown previously to participate in T-cell and B-cell trafficking and the migration of lymphocytes into secondary lymphoid organs in a cell intrinsic manner. We show here that the absence of Mst1 alone only modestly impacts B cell homing to lymph nodes. The absence of both Mst1 and 2 in hematopoietic cells results in relatively normal B cell development in the bone marrow and does not impact migration of immature B cells to the spleen. However, follicular B cells lacking both Mst1 and Mst2 mature in the splenic white pulp but are unable to recirculate to lymph nodes or to the bone marrow. These cells also cannot traffic efficiently to the splenic red pulp. The inability of late transitional and follicular B cells lacking Mst 1 and 2 to migrate to the red pulp explains their failure to differentiate into marginal zone B cell precursors and marginal zone B cells. Mst1 and Mst2 are therefore required for follicular B cells to acquire the ability to recirculate and also to migrate to the splenic red pulp in order to generate marginal zone B cells. In addition B-1 a B cell development is defective in the absence of Mst1

The transcriptional coactivator TAZ regulates reciprocal differentiation of T(h)17 cells and T(reg) cells

自身免疫性疾病是一类机体对自身抗原发生免疫反应而导致自身多器官、组织受累的慢性炎症性疾病。目前大量研究表明机体内促炎症的TH17细胞和抑制炎症Treg细胞在类群数量和活化状态的失衡是造成自身免疫疾病的主要致病因素。陈兰芬教授和周大旺教授团队的前期研究发现小鼠中Hippo信号通路中激酶Mst1/2缺失导致免疫缺陷，机体易受病原体感染并伴随着严重自身免疫疾病。该研究揭示了Hippo 信号通路转录共激活因子TAZ在决定CD4+初始T细胞分化为促进炎症的TH17效应细胞和抑制免疫反应的Treg调节性细胞过程中发挥着关键作用，拓展了当前对于Hippo信号通路的相关研究内容。 陈兰芬，博士，厦门大学生命科学学院教授。【Abstraact】An imbalance in the lineages of immunosuppressive regulatory T cells (Treg cells) and the inflammatory TH17 subset of helper T cells leads to the development of autoimmune and/or inflammatory disease. Here we found that TAZ, a coactivator of TEAD transcription factors of Hippo signaling, was expressed under T H17 cell–inducing conditions and was required for TH17 differentiation and TH17 cell–mediated inflammatory diseases. TAZ was a critical co-activator of the TH17-defining transcription factor RORγt. In addition, TAZ attenuated Treg cell development by decreasing acetylation of the Treg cell master regulator Foxp3 mediated by the histone acetyltransferase Tip60, which targeted Foxp3 for proteasomal degradation. In contrast, under T regcell–skewing conditions, TEAD1 expression and sequestration of TAZ from the transcription factors RORγt and Foxp3 promoted Treg cell differentiation. Furthermore, deficiency in TAZ or overexpression of TEAD1 induced Treg cell differentiation, whereas expression of a transgene encoding TAZ or activation of TAZ directed TH17 cell differentiation. Our results demonstrate a pivotal role for TAZ in regulating the differentiation of Treg cells and TH17 cells.J. Avruch for comments on the manuscript.Supported by the National Basic Research Program (973) of China (2015CB910502 to L.C.), the National Natural Science Foundation of China (81422018 to L.C.; 31625010 and U1505224 to D.Z.; U1405225 and 81372617 to L.C.; J1310027 to D.Z.; 81472229 to L.H.; and 31600698 to J. Geng), the 111 Projects (B12001 and B06016), China's 1000 Young Talents Program (D.Z., and L.C.), the Fundamental Research Funds for the Central Universities of China-Xiamen University (20720160071 to D.Z. and 20720160054 to L.H.) and Major disease research projects of Xiamen (3502Z20149029 to L.C.)

Impeded Nedd4-1-Mediated Ras Degradation Underlies Ras-Driven Tumorigenesis

RAS genes are among the most frequently mutated proto-oncogenes in cancer. However, how Ras stability is regulated remains largely unknown. Here, we report a regulatory loop involving the E3 ligase Nedd4-1, Ras, and PTEN. We found that Ras signaling stimulates the expression of Nedd4-1, which in turn acts as an E3 ubiquitin ligase that regulates Ras levels. Importantly, Ras activation, either by oncogenic mutations or by epidermal growth factor (EGF) signaling, prevents Nedd4-1-mediated Ras ubiquitination. This leads to Ras-induced Nedd4-1 overexpression, and subsequent degradation of the tumor suppressor PTEN in both human cancer samples and cancer cells. Our study thus unravels the molecular mechanisms underlying the interplay of Ras, Nedd4-1, and PTEN and suggests a basis for the high prevalence of Ras-activating mutations and EGF hypersignaling in cancer. © 2014 The Authors

ATR/Chk1 signaling induces autophagy through sumoylated RhoB-mediated lysosomal translocation of TSC2 after DNA damage

RhoB作为抑癌蛋白通过诱导肿瘤细胞的凋亡在抑制肿瘤的发生发展中发挥着重要作用，并且与肿瘤耐药性密切相关，但关于RhoB如何促进细胞死亡的分子机理的研究仍不清楚。在本研究中，该团队发现在DNA单链损伤情况下，ATR-Chk1信号通路的激活，会使RhoB被Chk1磷酸化，该磷酸化修饰会使RhoB从细胞质膜解离下来进入细胞质中，进而被SUMO化修饰。SUMO化修饰后的RhoB会与TSC2形成复合物，并将TSC2复合物带到溶酶体上，引起细胞自噬的发生。 该文共同第一作者为刘明冬、曾涛玲和张新，通讯作者为王洪睿教授和赵同金教授。【Abstract】DNA damage can induce autophagy; however, the underlying mechanism remains largely unknown. Here we report that DNA damage leads to autophagy through ATR/Chk1/RhoB-mediated lysosomal recruitment of TSC complex and subsequent mTORC1 inhibition. DNA damage caused by ultraviolet light (UV) or alkylating agent methyl methanesulphonate (MMS) results in phosphorylation of small GTPase RhoB by Chk1. Phosphorylation of RhoB enhances its interaction with the TSC2, and promotes its sumoylation by PIAS1, which is required for RhoB/TSC complex to translocate to lysosomes. As a result, mTORC1 is inhibited, and autophagy is activated. Knockout of RhoB severely attenuates lysosomal translocation of TSC complex and the DNA damage-induced autophagy. Reintroducing wild-type but not sumoylation-resistant RhoB into RhoB−/− cells restores the onset of autophagy. Hence, our study identifies a molecular mechanism for translocation of TSC complex to lysosomes in response to DNA damage, which depends on ATR/Chk1-mediated RhoB phosphorylation and sumoylation.This work was supported by the National Natural Science Foundation of China (U1605222, 81472459, 31671223), the National Key Research and Development Project of China (2016YFC1302400, 2016YFA0502003), the Fundamental Research Funds for the Central Universities (20720140550, 20720160070), the National Science Foundation for Fostering Talents in Basic Research of the National Natural Science Foundation of China (J1310027), the Project 111 sponsored by the State Bureau of Foreign Experts and Ministry of Education (B12001), the National Natural Science Foundation of China (31601132) to T.Z., the National Natural Science Foundation of China (81402290) to Q.L., and the National Natural Science Foundation of China (U1405223) to X.D