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1.
We investigated the cellular localization of ectopically-expressed CIS, SOCS1, SOCS2 and SOCS3 proteins. We found that SOCS proteins localize to the nucleus where they reduce Stat3 proteins and that the presence of proteasome inhibitors increased SOCS nuclear localization. Our results indicate that increased nuclear localization resulted from increased levels of SOCS proteins in the cytoplasm. Finally, we demonstrate that the same effect occurs with endogenously-expressed SOCS proteins. These observations suggest that increased cytoplasmic levels of proteins in the SOCS family are regulated through nuclear translocation. 相似文献
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Hwang MN Ha TH Park J Shim J Lee H Kim YN Lee ES Yoon S 《Biochemical and biophysical research communications》2007,354(1):184-189
We investigated stability of the ectopically expressed the SOCS6 protein in HEK293T cells with PMA, which activates protein kinase C (PKC). The treatment of PMA could largely increase SOCS6 stability in HEK293T cells. But, we did not observe increased protein levels of SOCS3 or Erk1 with PMA. This result suggests that the increased stability of SOCS6 with PMA did not generally occur in other proteins. The stability of SOCS6 depended on the N-terminal region containing an unidentified domain. We then studied the role of signal pathways in SOCS6 stability with PMA. We found that both Erk and Pkcdelta activation were required for the increased SOCS6 stability by PMA. The Erk activation by PMA appeared to be downstream from the Pkcdelta activation. The increased SOCS6 stability and Erk activation by PMA were both conserved in another cell line, MCF7. In addition, we demonstrated that PMA, insulin, and PDGF increased both the stability of endogenous-expressed SOCS6 and Erk activation in MDA-MB231 cells. These observations suggest that Erk activation may be correlated in the cells with high expression of SOCS6. 相似文献
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Mutational analyses of the SOCS proteins suggest a dual domain requirement but distinct mechanisms for inhibition of LIF and IL-6 signal transduction. 总被引:27,自引:0,他引:27
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S E Nicholson T A Willson A Farley R Starr J G Zhang M Baca W S Alexander D Metcalf D J Hilton N A Nicola 《The EMBO journal》1999,18(2):375-385
SOCS-1 (suppressor of cytokine signaling-1) is a representative of a family of negative regulators of cytokine signaling (SOCS-1 to SOCS-7 and CIS) characterized by a highly conserved C-terminal SOCS box preceded by an SH2 domain. This study comprehensively examined the ability of several SOCS family members to negatively regulate the gp130 signaling pathway. SOCS-1 and SOCS-3 inhibited both interleukin-6 (IL-6)- and leukemia inhibitory factor (LIF)-induced macrophage differentiation of murine monocytic leukemic M1 cells and LIF induction of a Stat3-responsive reporter construct in 293T fibroblasts. Deletion of amino acids 51-78 in the N-terminal region of SOCS-1 prevented inhibition of LIF signaling. The SOCS-1 and SOCS-3 N-terminal regions were functionally interchangeable, but this did not extend to other SOCS family members. Mutation of SH2 domains abrogated the ability of both SOCS-1 and SOCS-3 to inhibit LIF signal transduction. Unlike SOCS-1, SOCS-3 was unable to inhibit JAK kinase activity in vitro, suggesting that SOCS-1 and SOCS-3 act on the JAK-STAT pathway in different ways. Thus, although inhibition of signaling by SOCS-1 and SOCS-3 requires both the SH2 and N-terminal domains, their mechanisms of action appear to be biochemically different. 相似文献
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SH2-B, APS, and Lnk constitute a family of adapter proteins that modulate signaling by protein tyrosine kinases. These adapters contain an N-terminal dimerization region, a pleckstrin homology domain, and a C-terminal Src homology-2 (SH2) domain. SH2-B is recruited via its SH2 domain to various protein tyrosine kinases, including Janus kinase-2 (Jak2) and the insulin receptor. Here, we present the crystal structure at 2.35 A resolution of the SH2 domain of SH2-B in complex with a phosphopeptide representing the SH2-B recruitment site in Jak2 (pTyr813). The structure reveals a canonical SH2 domain-phosphopeptide binding mode, but with specific recognition of a glutamate at the +1 position relative to phosphotyrosine, in addition to recognition of a hydrophobic residue at the +3 position. Biochemical studies of SH2-B and APS demonstrate that, although the SH2 domains of these two adapter proteins share 79% sequence identity, the SH2-B SH2 domain binds preferentially to Jak2, whereas the APS SH2 domain has higher affinity for the insulin receptor. This differential specificity is attributable to the difference in the oligomeric states of the two SH2 domains: monomeric for SH2-B and dimeric for APS. 相似文献
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Wang X Tian QB Okano A Sakagami H Moon IS Kondo H Endo S Suzuki T 《Journal of neurochemistry》2005,92(3):647-659
We cloned a rat BAALC 1-6-8 isoform cDNA (GenBank Accession No. AB073318) that encoded a 22-kDa protein, and identified endogenous BAALC 1-6-8 protein in the brain. The gene was expressed widely in the frontal part of the brain, and the protein was localized to the synaptic sites and was increased in parallel with synaptogenesis. The protein interacted with the alpha, but not beta, subunit of Ca(2+)/calmodulin-dependent protein kinase II (CaMKIIalpha). The interaction occurred between the N-terminal 35-amino-acid region of BAALC 1-6-8 protein and the C-terminal end of the regulatory domain of CaMKIIalpha, which contains alpha isoform-specific sequence. Thus, the interaction may be CaMKIIalpha-specific. We also found that BAALC 1-6-8 protein, as well as CaMKIIalpha, was localized to lipid rafts and that both myristoylation and palmitoylation of BAALC 1-6-8 N-terminal portion were required for targeting of the protein into lipid rafts. These findings suggest that BAALC 1-6-8 protein play a synaptic role at the postsynaptic lipid raft possibly through interaction with CaMKIIalpha. 相似文献
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Chunguang Yan Jay Cao Min Wu Wei Zhang Tao Jiang Aihiko Yoshimura Hongwei Gao 《The Journal of biological chemistry》2010,285(48):37227-37239
Suppressor of cytokine signaling 3 (SOCS3) is an important intracellular protein that inhibits cytokine signaling in numerous cell types and has been implicated in several inflammatory diseases. However, the expression and function of SOCS3 in osteoblasts are not known. In this study, we demonstrated that SOCS3 expression was transiently induced by LPS in osteoblasts, and apparently contributed to the inhibition of IL-6 induction by LPS treatment. We found that tyrosine 204 of the SOCS box, the SH2 domain, and the N-terminal kinase inhibitory region (KIR) of SOCS3 were all involved in its IL-6 inhibition. Furthermore, we demonstrated that CCAAT/enhancer-binding protein (C/EBP) β was activated by LPS (increased DNA binding activity), and played a key role in LPS-induced IL-6 expression in osteoblasts. We further provided the evidence that SOCS3 functioned as a negative regulator for LPS response in osteoblasts by suppressing C/EBPβ DNA binding activity. In addition, tyrosine 204 of the SOCS box, the SH2 domain, and the N-terminal kinase inhibitory region (KIR) of SOCS3 were all required for its C/EBPβ inhibition. These findings suggest that SOCS3 by interfering with C/EBPβ activation may have an important regulatory role during bone-associated inflammatory responses. 相似文献
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Papazyan R Rozengurt E Rey O 《Biochemical and biophysical research communications》2006,342(3):685-689
We generated a set of GFP-tagged chimeras between protein kinase D2 (PKD2) and protein kinase D3 (PKD3) to examine in live cells the contribution of their C-terminal region to their intracellular localization. We found that the catalytic domain of PKD2 and PKD3 can localize to the nucleus when expressed without other kinase domains. However, when the C-terminal tail of PKD2 was added to its catalytic domain, the nuclear localization of the resulting protein was inhibited. In contrast, the nuclear localization of the CD of PKD3 was not inhibited by its C-terminal tail. Furthermore, the exchange of the C-terminal tail of PKD2 and PKD3 in the full-length proteins was sufficient to exchange their intracellular localization. Collectively, these data demonstrate that the short C-terminal tail of these kinases plays a critical role in determining their cytoplasmic/nuclear localization. 相似文献
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Analysis of leptin signalling in hematopoietic cells using an adapted MAPPIT strategy 总被引:2,自引:0,他引:2
Montoye T Piessevaux J Lavens D Wauman J Catteeuw D Vandekerckhove J Lemmens I Tavernier J 《FEBS letters》2006,580(13):3301-3307
The adipocyte-secreted hormone leptin participates in the regulation of hematopoiesis and enhances proliferation of hematopoietic cells. We used an adaptation of the MAPPIT mammalian two-hybrid method to study leptin signalling in a hematopoietic setting. We confirmed the known interactions of suppressor of cytokine signalling 3 (SOCS3) and STAT5 with the Y985 and Y1077 motifs of the leptin receptor, respectively. We also provide evidence for novel interactions at the Y1077 motif, including phospholipase C gamma and several members of the SOCS protein family, further underscoring the important role of the Y1077 motif in leptin signalling. 相似文献
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Ogura K Nagata K Horiuchi M Ebisui E Hasuda T Yuzawa S Nishida M Hatanaka H Inagaki F 《Journal of biomolecular NMR》2002,22(1):37-46
The three-dimensional structure of the N-terminal SH3 domain (residues 583–660) of murine Vav, which contains a tetra-proline sequence (Pro 607-Pro 610), was determined by NMR. The solution structure of the SH3 domain shows a typical SH3 fold, but it exists in two conformations due to cis-trans isomerization at the Gly614-Pro615 bond. The NMR structure of the P615G mutant, where Pro615 is replaced by glycine, reveals that the tetra-proline region is inserted into the RT-loop and binds to its own SH3 structure. The C-terminal SH3 domain of Grb2 specifically binds to the trans form of the N-terminal SH3 domain of Vav. The surface of Vav N-terminal SH3 which binds to Grb2 C-terminal SH3 was elucidated by chemical shift mapping experiments using NMR. The surface does not involve the tetra-proline region but involves the region comprising the n-src loop, the N-terminal and the C-terminal regions. This surface is located opposite to the tetra-proline containing region, consistent with that of our previous mutagenesis studies. 相似文献
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杜氏盐藻核基质结合区结合蛋白的细胞定位 总被引:1,自引:0,他引:1
为研究核基质结合区结合蛋白的功能及调控机制,PCR扩增杜氏盐藻MBP的cDNA全长序列及N端和C端序列,与绿色荧光蛋白基因融合构建真核表达载体,脂质体转染CHO细胞,Western blotting和荧光显微镜检测基因表达情况和细胞定位。结果显示:MBP及N端和C端融合蛋白成功在CHO细胞表达,MBP和C端部分定位于细胞核且聚集于核仁,N端部分分布整个细胞,说明MBP定位于细胞核且细胞定位信号位于C端,MBP可能与rRNA前体结合发挥作用。 相似文献
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Shenli Xi Yi Tie Kefeng Lu Xiushan Yin Guichun Xing Xiaofei Zheng Lingqiang Zhang 《FEBS letters》2010,584(6):1223-1230
The pleckstrin homology (PH) domain-containing protein casein kinase 2 interacting protein-1 (CKIP-1) plays an important role in regulation of bone formation and muscle differentiation. How CKIP-1 localization is determined remains largely unclear. We observed that isolated CKIP-1-PH domain was predominantly localized in the nucleus and the C-terminus of CKIP-1 counteracted its nuclear localization. The net charge of basic residues and a serine-rich motif within the PH domain plays a pivotal role in the localization switch of both full-length CKIP-1 and the isolated PH domain. We propose that the N-terminal PH domain and C-terminal auto-inhibitory region of CKIP-1 coordinate to determine its subcellular localization and the nucleus-plasma membrane shuttling. 相似文献
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Ankyrin repeat and SOCS box 3 (ASB3) mediates ubiquitination and degradation of tumor necrosis factor receptor II
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Ankyrin repeat and SOCS box (ASB) family members have a C-terminal SOCS box and an N-terminal ankyrin-related sequence of variable repeats belonging to the SOCS superfamily. While SH2-domain-bearing SOCS proteins are mainly involved in the negative feedback regulation of the protein tyrosine kinase-STAT pathway in response to a variety of cytokines, the roles of ASB family members remain largely unknown. To investigate ASB functions, we screened for ASB3-interacting factors by using antibody array technology and identified tumor necrosis factor receptor II (TNF-R2) as an ASB3 binding target. ASB3 expression and activities are required for (i) TNF-R2 ubiquitination both in vivo and in vitro, (ii) TNF-R2 proteolysis via the proteasome pathway, and (iii) the inhibition of TNF-R2-mediated Jun N-terminal protein kinase (JNK) activation. While the ankyrin repeats of ASB3 interact with the C-terminal 37 amino acids of TNF-R2, the SOCS box of ASB3 is responsible for recruiting the E3 ubiquitin ligase adaptors Elongins-B/C, leading to TNF-R2 ubiquitination on multiple lysine residues within its C-terminal region. Downregulation of ASB3 expression by a small interfering RNA inhibited TNF-R2 degradation and potentiated TNF-R2-mediated cytotoxicity. The data presented here implicate ASB3 as a negative regulator of TNF-R2-mediated cellular responses to TNF-alpha by direct targeting of TNF-R2 for ubiquitination and proteasome-mediated degradation. 相似文献
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Terasaki AG Suzuki H Nishioka T Matsuzawa E Katsuki M Nakagawa H Miyamoto S Ohashi K 《Biochemical and biophysical research communications》2004,313(1):48-54
From eluates of F-actin affinity chromatography of chicken brain, we identified a novel actin-binding protein (lasp-2) whose gene was predicted in silico. We cloned cDNA of chicken lasp-2 and analyzed its structure, expression, activity, and localization with lasp-1 (LIM and SH3 protein 1), a previously identified actin-binding protein closely related to lasp-2. Chicken lasp-2 showed high homology to mammalian putative lasp-2. Both chicken lasp-1 and chicken lasp-2 have N-terminal LIM domains, C-terminal SH3 domains, and internal nebulin repeats. However, lasp-2 is greatly different from lasp-1 in the sequence between the second nebulin repeat and a SH3 domain, and the region is conserved in chicken, mouse, and human. As expected from its structural similarity to lasp-1, lasp-2 possessed actin-binding activity and localized with actin filament in filopodia of neuroblastoma. In contrast to lasp-1, which is widely distributed in non-muscle tissues, lasp-2 was highly expressed in brain. 相似文献
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The adapter 3BP2 is involved in leukocyte signaling downstream Src/Syk-kinases coupled immunoreceptors. Here, we show that 3BP2 directly interacts with the endocytic scaffold protein CIN85 and the actin-binding protein HIP-55. 3BP2 co-localized with CIN85 and HIP-55 in T cell rafts and at the T cell/APC synapse, an active zone of receptors and proteins recycling. A binding region of CIN85 SH3 domains on 3BP2 was mapped to a PVPTPR motif in the first proline-rich region of 3BP2, whereas the C-terminal SH3 domain of HIP-55 bound a more distal proline-rich domain of 3BP2. Together, our data suggest an unexpected role of 3BP2 in endocytic and cytoskeletal regulation through its interaction with CIN85 and HIP-55. 相似文献