Multiple signal transduction pathways mediated by 5-HT receptors

Among human serotonin (5-HT) receptor subtypes, each G protein-coupled receptor subtype is reported to have one G protein-signaling cascade. However, the signaling may not be as simple as previously thought to be. 5-HT5A receptors are probably the least well understood among the 5-HT receptors, but the authors found that 5-HT5A receptors couple to multiple signaling cascades. When the 5-HT5A receptors were expressed in undifferentiated C6 glioma cells, they modulated the level of second messengers. For example, activation of 5-HT5A receptors inhibited the adenylyl cyclase activity and subsequently reduced the cAMP level, as previously reported. In addition to this known signaling via Gi/Go, 5-HT5A receptors are coupled to the inhibition of ADP-ribosyl cyclase and cyclic ADP ribose formation. On the other hand, activation of 5-HT5A receptors transiently opened the K+ channels, presumably due to the increase in intracellular Ca2+ after formation of inositol (1,4,5) trisphosphate. The K+ currents were inhibited by both heparin and pretreatment with pertussis toxin, suggesting the cross-talk between Gi/Go protein and phopholipase C cascade. Thus, the authors results indicate that 5-HT5A receptors couple to multiple second messenger systems and may contribute to the complicated physiological and pathophysiological states. Although this multiple signaling has been reported only for 5-HT5A/5-HT1 receptors so far, it is possible that other 5-HT receptor subtypes bear similar complexity. As a result, in addition to the wide variety of expression patterns of each 5-HT receptor subtype, it is possible that multiple signal transduction systems may add complexity to the serotonergic system in brain function. The investigation of these serotonergic signaling and its impairment at cellular level may help to understand the symptoms of brain diseases.     

PSD—95对NMDA受体信号转导的整合作用

PSD－95是新近在谷氨酸能突触的突触后致密物（PSD）中发现的一种特殊蛋白质,含有3个N末端的PDZ结构域,一个SH3结构域和一个C末端的GK结构域。PSD－95通过不同结构域与其它蛋白相互作用,不仅能够串集NMDA受体及其信号通路中的相关蛋白分子,组成受体－信号分子－调节分子－靶分子复合物,还可通过突触前后粘附分子的相互作用,参与突触连接的形成和维持,在介导和整合NMDA受体信号转导中具有关键性作用。     

受体激酶介导的油菜素内酯信号转导途径

油菜素内酯（brassinosteroids,BRs）是一类重要的类固醇激素,参与调控植物生长发育的许多过程。结合应用遗传学、生物化学以及蛋白质组学等研究手段现已基本阐明了BR信号转导的主要过程。BRI1作为受体在细胞表面感知BR,BRI1抑制子BKI1从质膜上解离下来,使BRI1与其共受体BAK1结合。BRI1和BAK1通过顺序磷酸化将BR信号完全激活。活化的BRI1将BSK磷酸化激活,BSK活化BSU1,BSU1将BIN2去磷酸化使其失活,解除BIN2对BES1/BZR1的抑制功能。PP2A可以将BES1/BZR1去磷酸化激活,又可以将受体BRI1去磷酸化促使其降解。BR信号的传递最终使去磷酸化状态的BES1/BZR1在细胞内累积,激活BR信号通路下游的转录调控。     

Specific localization and timing in neuronal signal transduction mediated by protein-lipid interactions

A large number of signaling proteins translocate from the cytosol to the plasma membrane in response to receptor and electrical stimuli. The site of translocation to the plasma membrane and the "on" and "off" rates of the translocation process are critical for defining the specificity of the signaling response. In addition to targeting mechanisms based on protein-protein interactions, signaling proteins have evolved a large repertoire of covalent lipid modifications and lipid binding protein modules that regulate reversible membrane association. The time constants of these membrane interactions range from milliseconds to several hours. Here we discuss how diversity in lipid-based membrane anchoring and targeting motifs contributes to plasticity in neuronal signaling by providing local and regional control mechanisms as well as a means to transduce and integrate signals over a broad range of different time scales.     

Ganglioside GM3 inhibition of EGF receptor mediated signal transduction

Ganglioside GM3 is a membrane component that has been describedto modulate cell growth through inhibition of EGF receptor associatedtyrosine kinase. In order to determine if the inhibition ofcell growth by this ganglioside is specifically mediated throughEGF receptor signaling, the effects of GM3 on key enzymes implicatedin EGF signaling were determined and compared to another inhibitorof the EGF receptor kinase. Treatment of A1S cells in cultureby GM3 or a tyrosine kinase inhibitor, leflunomide, led to theinhibition of MAP kinase and PI3 kinase activities. There wasno detectable effect on phosphotyrosine phosphatases. In a cellfree system, however, GM3 had no effect on the activity of thesesignaling intermediates. Leflunomide was able to directly inhibitMAP kinase activity. GM3 and leflunomide were also found toact differently on the expression of the early immediate genes.The expression of c-fos and c-jun was inhibited by both GM3and leflunomide. The expression of c-myc, however, was onlyinhibited by leflunomide. These findings suggest that the actionof GM3 on cell growth and signaling is specifically mediatedby EGF receptor and that this ganglioside does not act directlyon the intracellular intermediates of EGF receptor signaling.In addition, soluble small molecule tyrosine kinase inhibitorssuch as leflunomide can directly affect the activity of MAPkinases and possibly other signaling intermediates. The directeffects of leflunomide on signaling intermediates may explainthe differential effects of leflunomide and GM3 on gene expressionand cell growth. cell growth epidermal growth factor gangliosides GM3 signal transduction     

Radiation-induced upregulation of gamma-glutamyltransferase in colon carcinoma cells is mediated through the Ras signal transduction pathway

The activity of gamma-glutamyltransferase (GGT) is frequently upregulated in tumor cells after oxidative stress and may thus increase the availability of amino acids needed for biosynthesis of the antioxidant glutathione. As gamma-radiation of tumor cells can result in oxidative stress, we investigated whether such treatments modulate the enzyme level in colon carcinoma CC531 cells. Radiation of these cells blocked cell proliferation, increased cellular size, initiated apoptosis and upregulated GGT activity and protein levels in a dose- and time-related manner. A slight but significant increase in the cellular level of reactive oxygen species (ROS) was found directly after radiation but appeared not to cause the GGT elevation. Thus, other mechanisms than cellular oxidative stress appear to be responsible for the radiation-induced upregulation of GGT. Stable transfection of activated Ras in a human colon carcinoma cell line expressing wild-type Ras resulted in an increased GGT level, while a reduced enzyme level was demonstrated in another cell line with constitutively activated Ras after stably transfection with a dominant-negative Ras mutant. Moreover, addition of specific protein kinase inhibitors that blocked downstream targets PI-3K and MEK1/2 of Ras, prior to and after radiation, attenuated the radiation-induced activation of GGT. These results support a role for Ras, being frequently activated after radiation, in regulating the level of GGT and also indicate that GGT participates in radioresistance.     

Evidence that signal transduction for osmoreception is mediated by stretch-activated ion channels in tilapia

Prolactin (PRL) playsa central role in the freshwater osmoregulation of teleost fish,including the tilapia (Oreochromis mossambicus). Consistentwith this action, PRL release from the tilapia pituitary increases asextracellular osmolality is reduced both in vitro and in vivo.Dispersed tilapia PRL cells were incubated in a perfusion chamber thatallowed simultaneous measurements of cell volume and PRL release.Intracellular Ca²⁺ concentrations were measured from fura2-loaded PRL cells treated in a similar way. Gadolinium(Gd³⁺), known to block stretch-activated cation channels,inhibited hyposmotically induced PRL release in a dose-related mannerwithout preventing cell swelling. Nifedipine, an L-typeCa²⁺ channel blocker, did not prevent the increase in PRLrelease during hyposmotic stimulation. A high, depolarizingconcentration of KCl induced a transient and marked increase ofintracellular Ca²⁺ and release of PRL but did not preventthe rise in intracellular Ca²⁺ and PRL release evoked byexposure to hyposmotic medium. These findings suggest that a decreasein extracellular osmolality stimulates PRL release through the openingof stretch-activated ion channels, which allow extracellularCa²⁺ to enter the cell when it swells.

     

Self assembly of the transmembrane domain promotes signal transduction through the erythropoietin receptor

Hematopoietic cytokine receptors, such as the erythropoietin receptor (EpoR), are single membrane-spanning proteins. Signal transduction through EpoR is crucial for the formation of mature erythrocytes. Structural evidence shows that in the unliganded form EpoR exists as a preformed homodimer in an open scissor-like conformation precluding the activation of signaling. In contrast to the extracellular domain of the growth hormone receptor (GHR), the structure of the agonist-bound EpoR extracellular region shows only minimal contacts between the membrane-proximal regions. This evidence suggests that the domains facilitating receptor dimerization may differ between cytokine receptors. We show that the EpoR transmembrane domain (TM) has a strong potential to self interact in a bacterial reporter system. Abolishing self assembly of the EpoR TM by a double point mutation (Leu 240-Leu 241 mutated to Gly-Pro) impairs signal transduction by EpoR in hematopoietic cells and the formation of erythroid colonies upon reconstitution in erythroid progenitor cells from EpoR(-/-) mice. Interestingly, inhibiting TM self assembly in the constitutively active mutant EpoR R129C abrogates formation of disulfide-linked receptor homodimers and consequently results in the loss of ligand-independent signal transduction. Thus, efficient signal transduction through EpoR and possibly other preformed receptor oligomers may be determined by the dynamics of TM self assembly.     

IgD receptor-mediated signal transduction in T cells

Upregulation of immunoglobulin D-specific receptors (IgD-R) on CD4+ T cells may facilitate their interaction with specific carbohydrate moieties uniquely associated with membrane IgD on B cells. Previous studies have shown that upregulation of IgD-R facilitates cognate T-B cell interactions by mediating bidirectional signaling resulting in increased antibody responses and clonal expansion of antigen-specific T cells. Murine T hybridoma cells, 7C5, constitutively express IgD-R, as has been confirmed by staining with biotinylated IgD. Earlier studies have shown that inhibitors of protein tyrosine kinase (PTK) completely prevented upregulation of IgD-R in response to oligomeric IgD, suggesting that cross-linking of IgD-R may induce signal transduction and functional consequences through one or more PTK activation pathways, leading to upregulation of IgD-R. In the present study we show that cross-linking of IgD-R by oligomeric IgD indeed results in (a) T cell activation as seen by tyrosine phosphorylation of several intracellular proteins, (b) tyrosine phosphorylation of p56 Lck and PLC-gamma in 7C5 T hybridoma cells, and (c) phosphorylation of an approximately 29-kDa band that exhibits strong affinity for IgD. We analyzed tyrosine phosphorylation of p56 Lck and PLC-gamma in BALB/c splenic T cells that were exposed to oligomeric IgD both in vivo and in vitro. In vitro cross-linking as well as in vivo followed by in vitro cross-linking of IgD-R resulted in enhanced phosphorylation of p56 Lck and moderate tyrosine phosphorylation of PLC-gamma. These results suggest that interactions between IgD-R and IgD mediate signal transduction and support our previous findings that IgD-R+ T cells enhance cognate T cell-B cell interactions and antibody production.     

Modulation of cell-cycle regulatory signaling network by 2-methoxyestradiol in prostate cancer cells is mediated through multiple signal transduction pathways

2-Methoxyestradiol (2-ME(2)), a promising anticancer drug, induces growth arrest and apoptosis in various androgen-dependent (LNCaP) and -independent (DU145 and PC-3) prostate cancer cell lines. Moreover, flow cytometric analysis indicated a novel dual impact of 2-ME(2) on the cell division cycle of prostate cancer cells. Chronic exposure of high doses of 2-ME(2) enhance the accumulation of cells in S and G2/M phases, while cell numbers in the G1 phase were reduced significantly by this treatment. Because cyclin B1 overexpression, induction of cdc2 phosphorylation, and its regulatory proteins wee1 and phospho-cdc25C (interphase and mitotic forms) by 2-ME(2) treatment correlated with the induction of apoptosis, growth arrest at the G2/M phase, and accumulation of the S phase, we reasoned that cyclin B1 and cdc2 phosphorylation and its upstream regulatory molecular networks may be associated with the ultimate impacts of 2-ME(2). Because phosphorylation of cdc2 and upregulation of wee1 by 2-ME(2) can be abolished by both extracellular receptor kinase (ERK) inhibitor (U0126) and c-Jun N-terminal kinase (JNK) inhibitor (SP600125), our studies indicate that the 2-ME(2)-induced upregulation of wee1 and subsequent cdc2 phosphorylation are mediated through mitogen-activated protein kinase (MAPK)-ERK-JNK signaling pathways.     

Sphingosine 1-phosphate signal transduction in muscle cells

Sphingosine 1-phosphate (S1P) is a powerful bioactive sphingolipid recently recognized to act as extracellular ligand for various subtypes of G protein-coupled receptors belonging to the S1P family. In our study, focused on mouse skeletal muscle cells, we showed that S1P activated enzymes crucial for membrane signal transduction, such as phospholipase D (PLD) and protein kinase C; it promoted also a significant increase of cytosolic Ca2+ via ligation to S1P2 and S1P3 receptor subtypes. Interestingly, myogenic differentiation was found to be accompanied by a profound variation of S1P receptor expression levels and the progressive uncoupling of S1P from PLD activation, suggesting that this signaling pathway is exclusively required for S1P action on proliferating cells.     

Hyperoxia-induced signal transduction pathways in pulmonary epithelial cells

Mechanical ventilation with hyperoxia is necessary to treat critically ill patients. However, prolonged exposure to hyperoxia leads to the generation of excessive reactive oxygen species (ROS), which can cause acute inflammatory lung injury. One of the major effects of hyperoxia is the injury and death of pulmonary epithelium, which is accompanied by increased levels of pulmonary proinflammatory cytokines and excessive leukocyte infiltration. A thorough understanding of the signaling pathways leading to pulmonary epithelial cell injury/death may provide some insights into the pathogenesis of hyperoxia-induced acute inflammatory lung injury. This review focuses on epithelial responses to hyperoxia and some of the major factors regulating pathways to epithelial cell injury/death, and proinflammatory responses on exposure to hyperoxia. We discuss in detail some of the most interesting players, such as NF-kappaB, that can modulate both proinflammatory responses and cell injury/death of lung epithelial cells. A better appreciation for the functions of these factors will no doubt help us to delineate the pathways to hyperoxic cell death and proinflammatory responses.     

Regulation of signal transduction by endocytosis

Endocytosis of ligand-activated receptors has generally been considered a mechanism to attenuate signaling. There is now a growing body of evidence suggesting that this process is much more sophisticated and that endocytic membrane trafficking regulates both the intensity of signaling and the co-localization of activated receptors with downstream signaling molecules.