RESUMEN
Hippocampal neurons exhibit activation of both the conventional transmembrane adenylyl cyclases (tmACs) and the non-canonical soluble adenylyl cyclase (sAC) as sources of cyclic AMP (cAMP). These two cAMP sources play crucial roles in mediating signaling pathways downstream of CRHR1 in neuronal and neuroendocrine contexts. In this study, we investigate the involvement of both cAMP sources in the molecular mechanisms triggered by CRHR2α. Here we provide evidence demonstrating that UCN1 and UCN3 exert a neuritogenic effect on HT22-CRHR2α cells, which is solely dependent on the cAMP pool generated by sAC and PKA activity but independent of ERK1/2 activation. Through the characterization of the effectors implicated in neurite elongation, we found that CREB phosphorylation and c-Fos induction rely on PKA activity and ERK1/2 phosphorylation, underscoring the critical role of signaling pathway regulation. These findings strengthen the concept that localized cAMP microdomains actively participate in the regulation of these signaling processes.
Asunto(s)
Adenilil Ciclasas , Proteínas Quinasas Dependientes de AMP Cíclico , AMP Cíclico , Receptores de Hormona Liberadora de Corticotropina , Transducción de Señal , AMP Cíclico/metabolismo , Receptores de Hormona Liberadora de Corticotropina/metabolismo , Animales , Adenilil Ciclasas/metabolismo , Ratones , Fosforilación , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo , Urocortinas/metabolismo , Línea Celular , Neuritas/metabolismo , Proteínas Proto-Oncogénicas c-fos/metabolismo , Neuronas/metabolismoRESUMEN
Although transferrin (Tf) is a glycoprotein best known for its role in iron delivery, iron-independent functions have also been reported. Here, we assessed apoTf (aTf) treatment effects on Neuro-2a (N2a) cells, a mouse neuroblastoma cell line which, once differentiated, shares many properties with neurons, including process outgrowth, expression of selective neuronal markers, and electrical activity. We first examined the binding of Tf to its receptor (TfR) in our model and verified that, like neurons, N2a cells can internalize Tf from the culture medium. Next, studies on neuronal developmental parameters showed that Tf increases N2a survival through a decrease in apoptosis. Additionally, Tf accelerated the morphological development of N2a cells by promoting neurite outgrowth. These pro-differentiating effects were also observed in primary cultures of mouse cortical neurons treated with aTf, as neurons matured at a higher rate than controls and showed a decrease in the expression of early neuronal markers. Further experiments in iron-enriched and iron-deficient media showed that Tf preserved its pro-differentiation properties in N2a cells, with results hinting at a modulatory role for iron. Moreover, N2a-microglia co-cultures revealed an increase in IL-10 upon aTf treatment, which may be thought to favor N2a differentiation. Taken together, these findings suggest that Tf reduces cell death and favors the neuronal differentiation process, thus making Tf a promising candidate to be used in regenerative strategies for neurodegenerative diseases.
Asunto(s)
Neuronas , Transferrina , Ratones , Animales , Transferrina/química , Transferrina/metabolismo , Neuronas/metabolismo , Hierro/metabolismo , Línea Celular , Diferenciación CelularRESUMEN
Corticotropin releasing hormone (CRH) is crucial for basal and stress-initiated reactions in the hypothalamic-pituitary-adrenal axis (HPA) and extrahypothalamic brain circuits, where it acts as a neuromodulator to organize behavioral and humoral responses to stress. We review and describe cellular components and molecular mechanisms involved in CRH system signaling through G protein-coupled receptors (GPCRs) CRHR1 and CRHR2, under the current view of GPCR signaling from the plasma membrane but also from intracellular compartments, which establish the bases of signal resolution in space and time. Focus is placed on latest studies of CRHR1 signaling in physiologically significant contexts of the neurohormone function that disclosed new mechanistic features of cAMP production and ERK1/2 activation. We also introduce in a brief overview the pathophysiological function of the CRH system, underlining the need for a complete characterization of CRHRs signaling to design new and specific therapies for stress-related disorders.
Asunto(s)
Sistema Hipotálamo-Hipofisario , Sistema Hipófiso-Suprarrenal , Humanos , Sistema Hipotálamo-Hipofisario/metabolismo , Sistema Hipófiso-Suprarrenal/metabolismo , Hormona Liberadora de Corticotropina/metabolismo , Sistema Nervioso Central/metabolismo , EndocitosisRESUMEN
Depression and other psychiatric stress-related disorders are leading causes of disability worldwide. Up to date, treatments of mood disorders have limited success, most likely due to the multifactorial etiology of these conditions. Alterations in inflammatory processes have been identified as possible pathophysiological mechanisms in psychiatric conditions. Here, we review the main features of 2 systems involved in the control of these inflammatory pathways: the CRH system as a key regulator of the stress response and the ATP-gated ion-channel P2X7 receptor (P2X7R) involved in the control of immune functions. The pathophysiology of depression as a stress-related psychiatric disorder is depicted in terms of the impact of CRH and P2X7R function on inflammatory pathways in the brain. Understanding pathogenesis of affective disorders will lead to the development of therapies for treatment of depression and other stress-related diseases.
Asunto(s)
Hormona Liberadora de Corticotropina , Trastornos Mentales , Encéfalo/metabolismo , Hormona Liberadora de Corticotropina/metabolismo , Depresión , Humanos , Receptores Purinérgicos P2X7RESUMEN
The development of live-cell sensors for real-time measurement of signaling responses, with improved spatial and temporal resolution with respect to classical biochemical methods, has changed our understanding of cellular signaling. Examination of cAMP generation downstream activated GPCRs has shown that signaling responses can be short-lived (generated from the cell surface) or prolonged after receptor internalization. Class B secretin-like Corticotropin-releasing hormone receptor 1 (CRHR1) is a key player in stress pathophysiology. By monitoring real-time signaling in living cells, we uncovered cell context-dependent temporal characteristics of CRHR1-elicited cAMP responses and disclosed a specific link between cAMP generation and receptor signaling from internal compartments. We describe technical aspects and elaborate the protocols for cell line expression of Förster resonance energy transfer (FRET)-based biosensors to study the dynamics of cAMP and calcium signaling responses downstream activated CRHR1, live-cell imaging and analysis, and fluorescence flow cytometry to determine receptor levels at the cell surface.
Asunto(s)
Sistemas de Computación , Endocitosis , Transferencia Resonante de Energía de Fluorescencia/métodos , Receptores de Hormona Liberadora de Corticotropina/agonistas , Transducción de Señal , Animales , Calcio/metabolismo , Línea Celular , AMP Cíclico/metabolismo , Humanos , Ratones , Ratas , Receptores de Hormona Liberadora de Corticotropina/metabolismoRESUMEN
Class B G protein-coupled receptors (GPCRs) are involved in a variety of human pathophysiological states. These groups of membrane receptors are less studied than class A GPCRs due to the lack of structural information, delayed small molecule drug discovery, and scarce fluorescence detection tools available. The class B corticotropin-releasing hormone type 1 receptor (CRHR1) is a key player in the stress response whose dysregulation is critically involved in stress-related disorders: psychiatric conditions (i.e. depression, anxiety, and addictions), neuroendocrinological alterations, and neurodegenerative diseases. Here, we present a strategy to label GPCRs with a small fluorescent antagonist that permits the observation of the receptor in live cells through stochastic optical reconstruction microscopy (STORM) with 23 nm resolution. The marker, an aza-BODIPY derivative, was designed based on computational docking studies, then synthesized, and finally tested in biological cells. Experiments on hippocampal neurons demonstrate antagonist effects in similar concentrations as the well-established antagonist CP-376395. A quantitative analysis of two color STORM images enabled the determination of the binding affinity of the new marker in the cellular environment.
Asunto(s)
Simulación del Acoplamiento Molecular , Nanotecnología , Imagen Óptica , Receptores de Hormona Liberadora de Corticotropina/química , Biomarcadores/química , Colorantes Fluorescentes/síntesis química , Colorantes Fluorescentes/química , Colorantes Fluorescentes/farmacología , Humanos , Microscopía Fluorescente , Estructura Molecular , Receptores de Hormona Liberadora de Corticotropina/antagonistas & inhibidoresRESUMEN
Corticotropin-releasing hormone (CRH) is a key player of basal and stress-activated responses in the hypothalamic-pituitary-adrenal axis (HPA) and in extrahypothalamic circuits, where it functions as a neuromodulator to orchestrate humoral and behavioral adaptive responses to stress. This review describes molecular components and cellular mechanisms involved in CRH signaling downstream of its G protein-coupled receptors (GPCRs) CRHR1 and CRHR2 and summarizes recent findings that challenge the classical view of GPCR signaling and impact on our understanding of CRHRs function. Special emphasis is placed on recent studies of CRH signaling that revealed new mechanistic aspects of cAMP generation and ERK1/2 activation in physiologically relevant contexts of the neurohormone action. In addition, we present an overview of the pathophysiological role of the CRH system, which highlights the need for a precise definition of CRHRs signaling at molecular level to identify novel targets for pharmacological intervention in neuroendocrine tissues and specific brain areas involved in CRH-related disorders.
RESUMEN
Corticotropin-releasing hormone receptor 1 (CRHR1) activates the atypical soluble adenylyl cyclase (sAC) in addition to transmembrane adenylyl cyclases (tmACs). Both cAMP sources were shown to be required for the phosphorylation of ERK1/2 triggered by activated G protein coupled receptor (GPCR) CRHR1 in neuronal and neuroendocrine contexts. Here, we show that activated CRHR1 promotes growth arrest and neurite elongation in neuronal hippocampal cells (HT22-CRHR1 cells). By characterising CRHR1 signalling mechanisms involved in the neuritogenic effect, we demonstrate that neurite outgrowth in HT22-CRHR1 cells takes place by a sAC-dependent, ERK1/2-independent signalling cascade. Both tmACs and sAC are involved in corticotropin-releasing hormone (CRH)-mediated CREB phosphorylation and c-fos induction, but only sAC-generated cAMP pools are critical for the neuritogenic effect of CRH, further highlighting the engagement of two sources of cAMP downstream of the activation of a GPCR, and reinforcing the notion that restricted cAMP microdomains may regulate independent cellular processes.
Asunto(s)
Diferenciación Celular , AMP Cíclico/metabolismo , Células Piramidales/citología , Células Piramidales/metabolismo , Receptores de Hormona Liberadora de Corticotropina/metabolismo , Adenilil Ciclasas/sangre , Adenilil Ciclasas/metabolismo , Animales , Biomarcadores , Proteína de Unión a CREB/metabolismo , Puntos de Control del Ciclo Celular , Supervivencia Celular , Células Cultivadas , Hormona Liberadora de Corticotropina/metabolismo , Humanos , RatonesRESUMEN
RSUME (RWD-containing SUMO Enhancer) is a small protein that increases SUMO conjugation to proteins. To date, four splice variants that codify three RSUME isoforms have been described, which differ in their C-terminal end. Comparing the structure of the RSUME isoforms we found that, in addition to the previously described RWD domain in the N-terminal, all these RSUME variants also contain an intermediate domain. Only the longest RSUME isoform presents a C-terminal domain that is absent in the others. Given these differences, we used the shortest and longest RSUME variants for comparative studies. We found that the C-terminal domain is dispensable for the SUMO-conjugation enhancer properties of RSUME. We also demonstrate that these two RSUME variants are equally induced by hypoxia. The NF-κB signaling pathway is inhibited and the HIF-1 pathway is increased more efficiently by the longest RSUME, by means of a greater physical interaction of RSUME267 with the target proteins. In addition, the mRNA and protein levels of these isoforms differ in human glioma samples; while the shortest RSUME isoform is expressed in all the tumors analyzed, the longest variant is expressed in most but not all of them. The results presented here show a degree of redundancy of the RSUME variants on the SUMO pathway. However, the increased inhibition conferred by RSUME267 over the NF-κB signaling pathway, the increased activation over the HIF-1 pathway and the different expression of the RSUME isoforms suggest specific roles for each RSUME isoform which may be relevant in certain types of brain tumors that express RSUME, like human pituitary adenomas and gliomas.
Asunto(s)
Biología Computacional , Factores de Transcripción/química , Factores de Transcripción/metabolismo , Secuencia de Aminoácidos , Animales , Línea Celular , Regulación de la Expresión Génica , Humanos , Factor 1 Inducible por Hipoxia/metabolismo , Modelos Moleculares , Datos de Secuencia Molecular , FN-kappa B/metabolismo , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Estructura Secundaria de Proteína , Proteína SUMO-1/metabolismo , Transducción de Señal , Factores de Transcripción/genéticaRESUMEN
CRH is a key regulator of neuroendocrine, autonomic, and behavioral response to stress. CRH-stimulated CRH receptor 1 (CRHR1) activates ERK1/2 depending on intracellular context. In a previous work, we demonstrated that CRH activates ERK1/2 in limbic areas of the mouse brain (hippocampus and basolateral amygdala). ERK1/2 is an essential mediator of hippocampal physiological processes including emotional behavior, synaptic plasticity, learning, and memory. To elucidate the molecular mechanisms by which CRH activates ERK1/2 in hippocampal neurons, we used the mouse hippocampal cell line HT22. We document for the first time that ERK1/2 activation in response to CRH is biphasic, involving a first cAMP- and B-Raf-dependent early phase and a second phase that critically depends on CRHR1 internalization and ß-arrestin2. By means of mass-spectrometry-based screening, we identified B-Raf-associated proteins that coimmunoprecipitate with endogenous B-Raf after CRHR1 activation. Using molecular and pharmacological tools, the functional impact of selected B-Raf partners in CRH-dependent ERK1/2 activation was dissected. These results indicate that 14-3-3 proteins, protein kinase A, and Rap1, are essential for early CRH-induced ERK1/2 activation, whereas dynamin and vimentin are required for the CRHR1 internalization-dependent phase. Both phases of ERK1/2 activation depend on calcium influx and are affected by calcium/calmodulin-dependent protein kinase II inactivation. Thus, this report describes the dynamics and biphasic nature of ERK1/2 activation downstream neuronal CRHR1 and identifies several new critical components of the CRHR1 signaling machinery that selectively controls the early and late phases of ERK1/2 activation, thus providing new potential therapeutic targets for stress-related disorders.
Asunto(s)
Hormona Liberadora de Corticotropina/farmacología , Endocitosis/efectos de los fármacos , Hipocampo/enzimología , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Proteínas Proto-Oncogénicas B-raf/metabolismo , Receptores de Hormona Liberadora de Corticotropina/metabolismo , Adenilil Ciclasas/metabolismo , Animales , Arrestinas/metabolismo , Calcio/metabolismo , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/metabolismo , AMP Cíclico/metabolismo , Activación Enzimática/efectos de los fármacos , Hipocampo/citología , Humanos , Ratones , Modelos Biológicos , Ratas , Transducción de Señal/efectos de los fármacos , Fracciones Subcelulares/efectos de los fármacos , Fracciones Subcelulares/metabolismo , Factores de Tiempo , Vimentina/metabolismo , beta-ArrestinasRESUMEN
Corticotropin-releasing hormone (CRH) plays a key role in adjusting the basal and stress-activated hypothalamic-pituitary-adrenal axis (HPA). CRH is also widely distributed in extrahypothalamic circuits, where it acts as a neuroregulator to integrate the complex neuroendocrine, autonomic, and behavioral adaptive response to stress. Hyperactive and/or dysregulated CRH circuits are involved in neuroendocrinological disturbances and stress-related mood disorders such as anxiety and depression. This review describes the main physiological features of the CRH network and summarizes recent relevant information concerning the molecular mechanism of CRH action obtained from signal transduction studies using cells and wild-type and transgenic mice lines. Special focus is placed on the MAPK signaling pathways triggered by CRH through the CRH receptor 1 that plays an essential role in CRH action in pituitary corticotrophs and in specific brain structures. Recent findings underpin the concept of specific CRH-signaling pathways restricted to specific anatomical areas. Understanding CRH action at molecular levels will not only provide insight into the precise CRH mechanism of action, but will also be instrumental in identifying novel targets for pharmacological intervention in neuroendocrine tissues and specific brain areas involved in CRH-related disorders.
Asunto(s)
Hormona Liberadora de Corticotropina/fisiología , Sistema Hipotálamo-Hipofisario/fisiología , Sistema Hipófiso-Suprarrenal/fisiología , Animales , Sistema Nervioso Central/fisiología , Humanos , Ratones , Modelos Animales , Receptores de Hormona Liberadora de Corticotropina/fisiología , Transducción de Señal/fisiologíaRESUMEN
B-Raf links a variety of extracellular stimuli downstream of cell surface receptors, constituting a determining factor in the ability of neurons to activate ERK. A detailed study of the B-Raf interactome is necessary to clarify the intricacy of B-Raf-dependent signal transduction. We used a mouse hippocampal cell line (HT22) that expresses B-Raf at high levels, to identify B-Raf associated proteins under endogenous expression conditions, avoiding artificial interactions from overexpression studies. We used stringent procedures to co-immunoprecipitate proteins that specifically associate with endogenous B-Raf with the help of gel electrophoresis separation and off-line LC-MALDI-MS/MS proteomic analysis. Our stringent protein identification criteria allowed confident identification of B-Raf interacting proteins under non-stimulating conditions. The presence of previously reported B-Raf interactors among the list of proteins identified confirms the quality of proteomic data. We identified tubulin and actin as B-Raf interactors for the first time, among structural and accessory proteins of cell cytoskeleton, molecular chaperones (Hsc70, GRP78), and cellular components involved in aspects of mRNA metabolism and translation. Interactions were validated in HT22 cells and in the neuronal cell line Neuro-2a providing further evidence that the identified proteins are B-Raf interactors, which constitute a basis for understanding MAPK pathway regulation in neurons.
Asunto(s)
Hipocampo/metabolismo , Neuronas/citología , Neuronas/metabolismo , Proteómica/métodos , Proteínas Proto-Oncogénicas B-raf/metabolismo , Animales , Western Blotting , Células Cultivadas , Cromatografía Liquida , Citoesqueleto/química , Citoesqueleto/metabolismo , Electroforesis en Gel Bidimensional , Chaperón BiP del Retículo Endoplásmico , Hipocampo/química , Ratones , Chaperonas Moleculares/química , Chaperonas Moleculares/metabolismo , Neuronas/química , Proteínas Proto-Oncogénicas B-raf/química , ARN Mensajero/metabolismo , Espectrometría de Masa por Láser de Matriz Asistida de Ionización DesorciónRESUMEN
A classical view of the neuroendocrine-immune network assumes bidirectional interactions where pro-inflammatory cytokines influence hypothalamic-pituitary-adrenal (HPA) axis-derived hormones that subsequently affect cytokines in a permanently servo-controlled circle. Nevertheless, this picture has been continuously evolving over the last years as a result of the discovery of redundant expression and extended functions of many of the molecules implicated. Thus, cytokines are not only expressed in cells of the immune system but also in the central nervous system, and many hormones present at hypothalamic-pituitary level are also functionally expressed in the brain as well as in other peripheral organs, including immune cells. Because of this intermingled network of molecules redundantly expressed, the elucidation of the unique roles of HPA axis-related molecules at every level of complexity is one of the major challenges in the field. Genetic engineering in the mouse offers the most convincing method for dissecting in vivo the specific roles of distinct molecules acting in complex networks. Thus, various immunological, behavioral, and signal transduction studies performed with different HPA axis-related mutant mouse lines to delineate the roles of beta-endorphin, the type 1 receptor of corticotropin-releasing hormone (CRHR1), and its ligand CRH will be discussed here.
Asunto(s)
Conducta/fisiología , Sistema Hipotálamo-Hipofisario/inmunología , Sistema Hipófiso-Suprarrenal/inmunología , Transducción de Señal , Animales , Sistema Hipotálamo-Hipofisario/enzimología , Ratones , Ratones Transgénicos , Especificidad de Órganos , Sistema Hipófiso-Suprarrenal/enzimologíaRESUMEN
Members of group I KT-HAK-KUP transporters play an important role in K+ acquisition by plant roots, a process that is strongly affected by salt stress. A PCR-based random mutagenesis approach on HvHAK1 allowed identification of V366I and R591C substitutions, which confer enhanced K+-capture, and improved NaCl, LiCl and NH4Cl tolerance, to yeast cells. Improved K+-capture was linked to an enhanced Vmax. Results reveal an intrinsic protective effect of K+, and assign an important role to the 8th transmembrane domain, as well as the C-terminus, in determining the maximum capacity for the transport of K+ in KT-HAK-KUP transporters.
Asunto(s)
Proteínas de Transporte de Catión/metabolismo , Proteínas de Plantas/metabolismo , Potasio/metabolismo , Salinidad , Sustitución de Aminoácidos , Arginina/genética , Arginina/metabolismo , Proteínas de Transporte de Catión/química , Proteínas de Transporte de Catión/genética , Cisteína/genética , Cisteína/metabolismo , Transporte Iónico , Isoleucina/genética , Isoleucina/metabolismo , Presión Osmótica , Proteínas de Plantas/química , Proteínas de Plantas/genética , Mutación Puntual , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Sodio/metabolismo , Cloruro de Sodio/metabolismo , Valina/genética , Valina/metabolismoRESUMEN
SUMO conjugation to proteins is involved in the regulation of diverse cellular functions. We have identified a protein, RWD-containing sumoylation enhancer (RSUME), that enhances overall SUMO-1, -2, and -3 conjugation by interacting with the SUMO conjugase Ubc9. RSUME increases noncovalent binding of SUMO-1 to Ubc9 and enhances Ubc9 thioester formation and SUMO polymerization. RSUME enhances the sumoylation of IkB in vitro and in cultured cells, leading to an inhibition of NF-kB transcriptional activity. RSUME is induced by hypoxia and enhances the sumoylation of HIF-1alpha, promoting its stabilization and transcriptional activity during hypoxia. Disruption of the RWD domain structure of RSUME demonstrates that this domain is critical for RSUME action. Together, these findings point to a central role of RSUME in the regulation of sumoylation and, hence, several critical regulatory pathways in mammalian cells.
Asunto(s)
Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Proteína SUMO-1/metabolismo , Factores de Transcripción/fisiología , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Hipoxia de la Célula , Línea Celular Tumoral , Chlorocebus aethiops , Humanos , Quinasa I-kappa B/metabolismo , Datos de Secuencia Molecular , FN-kappa B/metabolismo , Especificidad de Órganos , Unión Proteica , Proteínas Modificadoras Pequeñas Relacionadas con Ubiquitina/metabolismo , Factores de Transcripción/genética , Enzimas Ubiquitina-Conjugadoras/metabolismo , Ubiquitinas/metabolismoRESUMEN
Highly sophisticated mechanisms confer on the immune system the capacity to respond with a certain degree of autonomy. However, the final outcome of an immune response depends on the interaction of the immune system with other systems. The immune and neuroendocrine systems have an intimate cross-communication that makes possible a satisfactory response to environmental changes. Part of this interaction occurs through cytokines and steroid hormones. The last step of this cross-talk is the molecular level. As a model of interaction, this review focuses on the gp130 cytokine family. These cytokines, as well as their receptors, are expressed in pituitary cells. They regulate hormone production as well as growth of pituitary cells. During acute or chronic inflammation or infection, systemic, hypothalamic and hypophyseal gp130 cytokines act on anterior pituitary cells, integrating the neuroendocrine-immune response. Disruptions of these pathways may lead not only to abnormal growth of pituitary cells but also to immune disorders, for which, based on recent findings, targeting these cytokines might be a novel therapeutic approach.
Asunto(s)
Receptor gp130 de Citocinas/fisiología , Citocinas/fisiología , Hormonas/fisiología , Neuroinmunomodulación/fisiología , Transducción de Señal/inmunología , Animales , Humanos , Sistemas Neurosecretores/fisiologíaRESUMEN
The anterior pituitary can develop benign tumors of different sizes, classified as micro- and macroadenomas, frequently associated with high levels of hormone production, leading to different associated syndromes like Cushing's disease, acromegaly or prolactinomas. Much work has been done in order to understand the signaling pathways and the factors and hormones involved in the pituitary tumorigenic process. In recent years, much evidence has been collected and it is now well documented that cytokines of the gp130 family, such as interleukin-6, that use gp130 as a common signaling protein stimulate not only the proliferation but also the hormone secretion of pituitary cells. Experiments in vivo have shown that the overexpression of the gp130 receptor resulted in pituitary abnormal growth. Moreover, it has been recently described that bone morphogenetic protein-4 (BMP-4), a member of the TGF-beta family, has a stimulatory role on lactosomatotropic cells promoting the development of prolactinomas but it has an inhibitory action on the corticotropic lineage. This inhibitory action prevents Cushing's disease progression. Furthermore, BMP-4 mediates the antiproliferative action of retinoic acid in these cells. The present review highlights the most recent work about gp130 and TGF-beta cytokine families and their role in pituitary tumorigenesis.
Asunto(s)
Proteínas Morfogenéticas Óseas/fisiología , Receptor gp130 de Citocinas/fisiología , Citocinas/fisiología , Hipófisis/fisiología , Animales , Proteína Morfogenética Ósea 4 , Humanos , Modelos Biológicos , Familia de Multigenes/fisiologíaRESUMEN
YFR041C/ERJ5 was identified in Saccharomyces cerevisiae as a gene regulated by the unfolded protein response pathway (UPR). The open reading frame of the gene has a J domain characteristic of the DnaJ chaperone family of proteins that regulate the activity of Hsp70 chaperones. We determined the expression and topology of Erj5p, a type I membrane protein with a J domain in the lumen of the endoplasmic reticulum (ER) that colocalizes with Kar2p, the major Hsp70 in the yeast ER. We identified synthetic interactions of Deltaerj5 with mutations in genes involved in protein folding in the ER (kar2-159, Deltascj1Deltajem1) and in the induction of the unfolded protein response (Deltaire1). Loss of Erj5p in yeast cells with impaired ER protein folding capacity increased sensitivity to agents that cause ER stress. We identified the ERJ5 mRNA and confirmed that agents that promote accumulation of misfolded proteins in the ER regulate its abundance. We found that loss of the non-essential ERJ5 gene leads to a constitutively induced UPR, indicating that ERJ5 is required for maintenance of an optimal folding environment in the yeast ER.
Asunto(s)
Retículo Endoplásmico/metabolismo , Proteínas de la Membrana/química , Proteínas de la Membrana/metabolismo , Pliegue de Proteína , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/genética , Alelos , Secuencia de Aminoácidos , Northern Blotting , Eliminación de Gen , Regulación Fúngica de la Expresión Génica , Genes Fúngicos , Proteínas del Choque Térmico HSP40/química , Proteínas de la Membrana/genética , Chaperonas Moleculares/metabolismo , Datos de Secuencia Molecular , Estructura Terciaria de Proteína , ARN Mensajero/genética , ARN Mensajero/metabolismo , Proteínas Recombinantes de Fusión/metabolismo , Saccharomyces cerevisiae/citología , Saccharomyces cerevisiae/crecimiento & desarrollo , Proteínas de Saccharomyces cerevisiae/genética , Alineación de SecuenciaRESUMEN
Highly sophisticated mechanisms confer upon the immune system the capacity to respond with a certain degree of autonomy. However, the final outcome of an adaptative immune response depends on the interaction with other systems of the organism. The immune-neuroendocrine systems have an intimate cross-communication, making possible a satisfactory response to environmental changes. Part of this interaction occurs through cytokines and steroid hormones. The last step of this crosstalk is at the molecular level. In this article we will focus on the physical and functional interrelationship between cytokine signaling pathway-activated transcription factors (TFs) and steroid receptors in different cell models, where the signals triggered by cytokines and steroid hormones have major roles: (1) the ligand-dependent-activated glucocorticoid receptor (GR) influence the genetic program that specifies lineage commitment in T helper (Th) cell differentiation. How posttranslational modifications of several TFs as well as nuclear hormone receptors could be implicated in the molecular crosstalk between the immune-neuroendocrine messengers is discussed. (2) glucocorticoid (GC) antagonism on the TCR-induced T cell apoptosis. (3) estrogen receptor/TGF-beta family proteins molecular interaction implicated on pituitary prolactinomas pathogenesis. The functional crosstalk at the molecular level between immune and steroids signals is essential to determine an integrative response to both mediators (which in the last instance results in a new gene activation/repression profile) and constitutes the ultimate integrative level of interaction between the immune and neuroendocrine systems.
Asunto(s)
Citocinas/inmunología , Neuroinmunomodulación/inmunología , Sistemas Neurosecretores/inmunología , Esteroides/inmunología , Citocinas/metabolismo , Humanos , Sistemas Neurosecretores/metabolismo , Esteroides/metabolismoRESUMEN
The transcript levels of Candida albicans TPK1 and TPK2 genes, encoding PKA catalytic subunits, as well as phosphotransferase activity, were measured in the parental strain CAI4 and in homozygous tpk1Delta and tpk2Delta mutants during vegetative growth and during yeast-to-mycelial transition in N-acetylglucosamine liquid inducing medium at 37 degrees C. We observed two TPK2 transcripts, a major one of 1.8 kb and a minor one of 1.4 kb, and established by 3'-RACE that they originate from the recognition of the three polyadenylation signals present in the 3' untranslated region of the gene. During vegetative growth of CAI4 strain, the expression profiles of TPK1 and TPK2 varied similarly, reaching maximal expression at the late logarithmic phase. TPK1 mRNA levels were lower than those of TPK2 at all stages measured. In the corresponding homozygous tpk mutants, mRNA levels and the expression patterns of TPK1 and TPK2 were similar to those of CAI4, suggesting that the loss of one catalytic isoform is not compensated by overexpression of the other. Changes in PKA specific activity roughly correlated with fluctuations of mRNA expression levels. During yeast-to-mycelial transition, a sharp increase in TPK1 mRNA levels and in PKA-specific activity correlated with the onset of germ-tube formation in strain tpk2Delta. We also showed that tpk1Delta strain exhibited a delayed morphogenetic shift in comparison with CAI4 and tpk2Delta strains in several liquid inducing media, reinforcing the idea that Tpk1p is important for faster germ-tube appearance.