Biomedical subjects

Jay M Baraban

Publications and source records attributed to Jay M Baraban.

11 recordsLinked to original sources

PubMed2006

Developmental expression of the SRF co-activator MAL in brain: role in regulating dendritic morphology.

The dynamic changes in dendritic morphology displayed by developing and mature neurons have stimulated interest in deciphering the signaling pathways involved. Recent studies have identified megakaryocytic acute leukemia (MAL), a serum response factor (SRF) co-activator, as a key component of a signaling pathway linking changes in the actin cytoskeleton to SRF-mediated transcription. To help define the role of this pathway in regulating dendritic morphology, we have characterized the pattern of MAL expression in the developing and adult brain, and have examined its role in regulating dendritic morphology in cultured cortical neurons. In histological studies of mouse brain, we found prominent expression of MAL in neurons in adult hippocampus and cerebral cortex. MAL immunostaining revealed localization of this protein in neuronal cell bodies and apical dendrites. During development, an increase in MAL expression occurs during the second post-natal week. Expression of dominant negative MAL constructs or MAL siRNA in cortical neurons grown in primary culture reduces the number of dendritic processes and decreases the basal level of SRF-mediated transcription. Taken together, these findings indicate that the MAL-SRF signaling pathway plays a key role in regulating dendritic morphology.

Animals↗

PubMed2005

Tech: a RhoA GEF selectively expressed in hippocampal and cortical neurons.

Recent studies implicating the Rho family of small G proteins in the regulation of neuronal morphology have focused attention on identifying key components of Rho signaling pathways in neurons. To this end, we have conducted studies aimed at defining the localization and function of Tech, a Rho guanine nucleotide exchange factor (GEF) family member that is highly enriched in brain. We have found that Tech is selectively expressed in cortical and hippocampal neurons with prominent Tech immunostaining apparent in the cell bodies and dendrites of these cells. In vitro studies with prototypical members of the major Rho subfamilies, RhoA, Rac1 and Cdc42, indicate that Tech binds selectively to and activates RhoA. To assess whether Tech may be involved in the regulation of neuronal morphology, we examined the effects of Tech constructs on the morphology of cortical neurons grown in primary culture. We found that a constitutively active Tech construct, Tech 245DeltaC, decreases the number of dendritic processes present on these neurons. This reduction appears to be mediated by activation of RhoA as it is blocked by insertion of a point mutation into the DH domain of Tech which blocks its ability to activate RhoA or coexpression of a dominant negative RhoA construct. As Tech protein levels increase during post-natal development and remain at peak levels into adulthood, these results indicate that Tech regulates RhoA signaling pathways in developing and mature forebrain neurons.

Animals↗

PubMed2005

Nuclear translocation of the SRF co-activator MAL in cortical neurons: role of RhoA signalling.

Although it is well established that RhoA signaling pathways play key roles in regulating neuronal morphology, their involvement in other aspects of neuronal function has received little attention. Recent studies have elucidated a novel intracellular signaling pathway used by RhoA to elicit activation of serum response factor (SRF)-mediated transcription. In this pathway, activation of RhoA triggers nuclear translocation of the SRF co-activator, megakaryocytic acute leukemia (MAL). In assessing whether RhoA regulates transcription in neurons via this pathway, we have found that a constitutively active form of Tech (transcript-enriched in cortex and hippocampus), a RhoA guanine nucleotide exchange factor (GEF) that is expressed in forebrain neurons, stimulates SRF reporter activity in extracts of primary cortical cultures and induces nuclear translocation of MAL in cortical neurons. Both of these responses appear to be mediated by Tech's activation of RhoA as they are not mimicked by a mutant Tech construct lacking RhoA GEF activity and are blocked by C3 transferase, a selective inhibitor of RhoA. Furthermore, Tech-induced increases in SRF activity are suppressed by a dominant negative MAL construct. These findings demonstrate that RhoA signaling pathways are able to regulate transcription in neurons by triggering translocation of the SRF co-activator MAL.

Active Transport, Cell Nucleus↗

PubMed2004

High affinity binding of the Translin/Trax complex to RNA does not require the presence of Y or H elements.

Translin and its partner protein, Trax, are components of an RNA binding complex that has been implicated in suppressing translation of several mRNAs by binding to Y and H cis elements contained in these transcripts. However, it is unclear which features of these elements are critical for conferring high affinity binding to the Translin/Trax complex, information that might be useful in identifying other candidate transcripts targeted by this complex. To help clarify this issue, we have assessed the effect of truncating or mutating a segment of the 3'UTR of the protamine-2 transcript which contains both Y and H elements and binds to this complex with high affinity. Our results indicate that high affinity binding to this segment is preserved following extensive mutation of the Y and H elements as long as clusters of G residues are retained. Thus, our findings indicate that the Translin/Trax complex recognizes clusters of G residues rather than RNA sequences that closely match the primary sequence of the Y and H elements. This revised view of the cis elements recognized by the Translin/Trax complex may be useful in future studies aimed at identifying endogenous RNA species targeted by this complex.

3' Untranslated Regions↗

PubMed2003

Opiate withdrawal induces Narp in the extended amygdala.

The negative affective states associated with drug withdrawal produce long-lasting behavioral effects thought to play a central role in the development and maintenance of dependence. However, little is known about the molecular mechanisms mediating the long-term effects of drug withdrawal. Neuronal activity-regulated pentraxin (Narp) is a secreted neuronal immediate early gene (IEG) product that regulates AMPA receptor clustering at synapses. As both IEGs and changes in AMPA receptor trafficking mediate enduring forms of neuronal plasticity, we have assessed whether Narp could be involved in the molecular adaptations accompanying drug withdrawal. To this end, we checked the effect of opiate withdrawal on Narp expression in the extended amygdala, a brain region closely linked to the aversive effects of drug withdrawal. We found a marked increase in the number of Narp-positive cells in this region following opiate withdrawal triggered by either low doses of opiate antagonists or by 'natural withdrawal', removal of the morphine pellets used to induce dependence. In contrast, Arc, another 'effector' IEG, was not induced by opiate withdrawal. As expected, pretreatment of animals with clonidine, which blocks opiate withdrawal, suppresses Narp induction in this paradigm. These results implicate Narp in mediating the long-term, aversive behavioral effects induced by opiate withdrawal.

Amygdala↗

PubMed2003

ERK and p38 inhibit the expression of 4E-BP1 repressor of translation through induction of Egr-1.

4E-BP1 plays a major role in translation by inhibiting cap-dependent translation initiation. Several reports have investigated the regulation of 4E-BP1 phosphorylation, which varies along with cell differentiation and upon various stimulations, but very little is known about the regulation of its expression. In a first part, we show that the expression of 4E-BP1 protein and transcript decreases in hematopoietic cell lines cultivated in the presence of phorbol 12-myristate 13-acetate (PMA). This decrease depends on the activation of the ERK/mitogen-activated protein kinases. 4E-BP1 expression also decreases when the p38/mitogen-activated protein kinase pathway is activated by granulocyte/macrophage colony-stimulating factor but to a lesser extent than with PMA. In a second part, we examine how 4e-bp1 promoter activity is regulated. PMA and granulocyte/macrophage colony-stimulating factor induce Egr-1 expression through ERK and p38 activation, respectively. Using a dominant negative mutant of Egr, ZnEgr, we show that this transcription factor is responsible for the inhibition of 4e-bp1 promoter activity. In a third part we show that histidine decarboxylase, whose activity and expression are inversely correlated with 4E-BP1 expression, is a potential target for the translational machinery. These data (i) are the first evidence of a new role of ERK and p38 on the translational machinery and (ii) demonstrate that 4E-BP1 is a new target for Egr-1.

Adaptor Proteins, Signal Transducing↗

PubMed2002

A dominant negative Egr inhibitor blocks nerve growth factor-induced neurite outgrowth by suppressing c-Jun activation: role of an Egr/c-Jun complex.

Members of the Egr family of transcription factors are rapidly and robustly induced by neurotransmitters and neurotrophins and have been implicated in mediating enduring changes in neuronal function elicited by these stimuli. Because we have found in previous studies that a dominant negative inhibitor of Egr action, the Egr zinc finger domain (ZnEgr), blocks NGF-induced neurite outgrowth in PC12 cells, we have used this preparation to help identify the downstream targets of Egr proteins involved in plasticity. Our investigation into the mechanism of action of ZnEgr indicates that it blocks NGF-induced neurite outgrowth by suppressing activation of c-Jun, a critical step in the signaling pathway mediating this response. Although we had assumed that ZnEgr exerts its effects by binding to the Egr response element (ERE) and thereby blocking target gene regulation by Egr proteins, this classical mode of action appears to be too slow to mediate the effects of Egr proteins on c-Jun activation. In evaluating alternative ERE-independent mechanisms of Egr (and ZnEgr) action, we found that Egr1 and c-Jun coprecipitate and that ZnEgr disrupts formation of the Egr1/c-Jun complex. Furthermore, mutations of ZnEgr that greatly impair or abolish its ability to bind to the ERE do not block its ability to suppress c-Jun activation or neurite outgrowth induced by NGF. Accordingly, our studies indicate that Egr and ZnEgr proteins regulate c-Jun activation via a novel mechanism, protein-protein interaction with c-Jun, rather than via their classical mode of action, binding to the ERE.

Cell Line↗

PubMed2002

Prominent expression of Narp in central vestibular pathways: selective effect of labyrinth ablation.

Recent in vitro studies demonstrated that Narp, a secreted immediate early gene (IEG) product, induces AMPA receptor clustering. Accordingly, Narp has been implicated in mediating activity-dependent changes in synaptic efficacy. To help define the role of Narp in vivo, we conducted immunohistochemical studies of Narp in rat brain. Unexpectedly, we found robust Narp expression in several discrete areas linked to the vestibular system: the anterodorsal nucleus (ADN) of the thalamus, which relays head orientation information to the cortex, the lateral vestibulospinal (Deiters') nucleus and Purkinje cells in the flocculonodular lobe of the cerebellum. Although strong Narp expression in Deiters' nucleus and the cerebellum was present consistently, Narp expression in the ADN displayed a high degree of variability among animals. To check if this variability in ADN Narp expression reflects its dependence on fluctuating levels of vestibular input, we monitored Narp immunostaining following bilateral labyrinth ablation. This procedure significantly suppressed Narp immunostaining in the ADN, indicating that it is stimulated by naturally occurring vestibular input. In contrast, labyrinth ablation did not affect Narp staining in Deiters' nucleus or the flocculonodular lobe of the cerebellum, presumably because these areas are driven by inputs from multiple systems. As previous studies implicate Narp in synaptic plasticity, these findings suggest that this IEG may mediate ongoing adjustments in synaptic strength or connectivity in several pathways linked to the vestibular system.

Animals↗

PubMed2002

Prominent Narp expression in projection pathways and terminal fields.

Narp (neuronal activity regulated pentraxin) is a secreted immediate early gene product that is induced by synaptic activity. Recent studies have indicated that Narp may be an extracellular aggregating factor for AMPA receptors. Immunohistochemical studies have revealed prominent expression of Narp in the mossy fiber pathway of the dentate gyrus, suggesting it may be released pre-synaptically. However, in vitro studies using recombinant Narp indicate that Narp may act when expressed by either pre- or post-synaptic elements. To help define Narp's mode of action, we have examined its localization in the habenula-interpeduncular pathway which also displays robust Narp expression. Focusing on this pathway as well as hippocampal and cortical Narp expression, we found prominent Narp staining in projection pathways and terminal fields. In contrast, Narp expression in dendrites was minimal in these neuronal populations. These findings indicate that, under physiological conditions, Narp is targeted to the synapse from pre- rather than post-synaptic elements. Our results also suggest that future studies focusing on these projection pathways that express high levels of Narp, in vivo, may help to understand the regulation and function of endogenous Narp.

Animals↗

PubMed2002

Selective expression of Narp, a secreted neuronal pentraxin, in orexin neurons.

Recent studies have provided compelling evidence demonstrating that orexin (also known as hypocretin) neurons play a central role in the pathophysiology of narcolepsy. However, targeted deletion of orexin does not fully mimic the functional deficits induced by selective ablation of these neurons; implying that other secreted signaling molecules expressed in these neurons mediate key aspects of their function. In this study, we demonstrate that orexin neurons display robust expression of neuronal activity-regulated pentraxin (Narp), a secreted neuronal pentraxin, implicated in regulating clustering of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) receptors. Furthermore, we have found that hypothalamic melanin-concentrating hormone (MCH) neurons, which form a peptidergic pathway thought to oppose the effects of the orexin system, express another neuronal pentraxin, NP1. Thus, these findings suggest that these pathways utilize neuronal pentraxins, in addition to neuropeptides, as synaptic signaling molecules.

Animals↗

PubMed2002

Trax is a component of the Translin-containing RNA binding complex.

Translin is a nucleic acid binding protein that has been implicated in regulating the targeting and translation of dendritic RNA. In previous studies, we found that Translin and its partner protein, Trax, are components of a gel-shift complex that is highly enriched in brain extracts. In those studies, we employed a DNA oligonucleotide, GS1, as a probe to label the complex. Translin has also been identified as a component of a gel-shift complex detected using an RNA oligonucleotide probe, derived from the 3' UTR of protamine-2 mRNA. Although we had assumed that these probes labeled the same complex, recent studies indicate that association of Trax with Translin suppresses its RNA binding activity. As these findings challenge this assumption and suggest that the native RNA binding complex does not contain Trax, we have re-examined this issue. We have found that the gel-shift complexes labeled with either GS1 or protamine-2 probes are "supershifted" by addition of Trax antibodies, indicating that both are heteromeric Translin/Trax complexes. In addition, cross-competition studies provide additional evidence that these probes label the same complex. Furthermore, analysis of recombinant Translin/Trax complexes generated by co-transfection of Trax with Translin in hEK293T demonstrates that they are labeled with either probe. Although recombinant Translin forms a homomeric nucleic acid binding complex in vitro, our findings indicate that both Trax and Translin are components of the native gel-shift complex labeled with either GS1 or protamine-2 probes.

Animals↗