Ligands of the EGF/Heregulin family control the growth of epithelial cells by binding to receptors of the erbB family. By searching a large database of cDNA sequences at Human Genome Sciences Inc. we have identified a new encoded protein sequence containing all the conserved elements of the EGF/Heregulin family. The same sequence has recently been independently identified as NRG-3. The EGF-like domain of NRG-3 was generated as a recombinant protein in E. coli and used to test the specificity of receptor binding. In human breast cancer cells and in 32D cells transfected by erbB family members, NRG-3 activated multiple erbB family members. These include EGF receptor (erbB1) and erbB4 when expressed individually and erbB2 and erbB3 when expressed together. Recombinant NRG-3 altered the growth of human breast cancer cells growing in vitro. NRG-3 was expressed in cell lines derived from breast cancer. These results indicate that NRG-3 is a potential regulator of normal and malignant breast epithelial cells in vivo.

Overexpression of ErbB2 and ErbB4 receptors in breast cancers may be accompanied by contrasting clinical outcomes. To investigate the molecular mechanisms contributing to these differences, we undertook a comparative study of gene expression regulated by the two receptors. Agonistic antibodies were employed to activate ErbB2 and ErbB4 in isolation from the other ErbBs in breast cancer cells. Gene expression profiling using a 16 755-gene oligonucleotide array was performed to identify transcriptional targets of receptor activation. Our results indicate that, in the same cell line, ErbB2 and ErbB4 activation influence gene transcription differentially. Although there are genes that are regulated by signaling from both receptors, there are also receptor-specific targets that are preferentially regulated by each receptor. We further show that two ligands acting via the same receptor homodimer may activate different subsets of genes. Many of the induced genes are hitherto unidentified targets of ErbB signaling. These include ErbB4 targets EPS15R, GATA4, and RAB2 and ErbB2-activated HRY/HES1 and PPAP2A. Targets of ErbB2 homodimer signaling may be especially important as markers in breast cancer, where ErbB2 homodimerization mediated by overexpression and ligand-independent activation is common.

ErbB4 is a predominant heterodimer for other ErbB receptors in late fetal lung development where it participates in regulating type II cell surfactant synthesis. To further elucidate the role of ErbB4 in pulmonary alveolar epithelial cell function, the authors hypothesized that ErbB4 participates in maintaining adult lung type II cell homeostasis. The authors used small interfering RNA (siRNA) to down-regulate endogenous, ErbB4 receptors in the adult rat lung epithelial L2 cell line and measured neuregulin 1beta (NRG1beta)-, and fibroblast conditioned medium (FCM)-induced effects on L2 cell surfactant phospholipid synthesis and proliferation. Under control conditions, total and phosphorylated ErbB4 were significantly increased after both NRG1beta and FCM treatment, as were surfactant phospholipids synthesis and cell proliferation. Down-regulation of ErbB4 with siRNA reduced stimulation of NRG1beta- and FCM-induced ErbB4 phosphorylation, decreased endogenous surfactant phospholipid synthesis, and blocked NRG1beta- and FCM-stimulated surfactant phospholipid synthesis. NRG1beta- and FCM-induced cell proliferation was not affected. The authors conclude that ErbB4 participates in maintaining adult lung alveolar epithelial cell surfactant synthesis and proliferation with development-specific functions.

Integrins are widely expressed cell surface molecules that mediate cell attachment to extracellular matrix (ECM) proteins. They also interact with molecules on their own membranes, and these cis-interactions play a crucial role in integrin-dependent cellular responses. We herein analysed what molecules interact with β1 integrin during biological events induced by cell attachment to different ECM proteins, using a recently established reaction, the enzyme-mediated activation of radical sources (EMARS). The interactions between β1 integrin and receptor tyrosine kinases including EGFR and ErbB4 reached a peak at 2 h after seeding HeLa S3 cells onto the ECM proteins. The peak of phosphorylation of ErbB4 (at 2 h after seeding the cells onto fibronectin) coincided with the peak of the interaction with β1 integrin, while that of EGFR (at 1 day) did not. Accompanying with these findings, suppression of cell migration by a pharmacological inhibitor of the ErbB family receptors, PD168393 and an anti-ErbB4 neutralizing antibody, 12D8 was observed at 2 h after seeding. Taken together, it is deduced that interactions between β1 integrin and ErbB4 occur in a spatiotemporally-regulated manner, and such interaction contributes to the integrin-dependent cell migration.

BACKGROUND

Members of the ErbB family of the receptor protein tyrosine kinase superfamily mediate heregulin (HRG)-induced cell responses. Here we investigated HRG activation of ErbB receptors, and the role of this activation in the development of the permeability barrier in airway epithelial cells (AECs).

METHODS

Two airway epithelial-like cell lines, Calu-3 and 16HBE were exposed to HRG or no stimulus and were evaluated with respect to their paracellular permeability as determined by transepithelial electric resistance (TER) and fluorescein isothiocyanate (FITC)-dextran flux. Tight junctions (TJs) were assessed by immunocytochemical localization of occludin and zonula occludens-1.

RESULTS

HRG promoted the development of the permeability barrier and TJ formation by monolayers of Calu-3 and 16HBE cells. Calu-3 cells expressed ErbB1, ErbB2, and ErbB3, but not ErbB4, on their surface. ErbB3 knockdown by small interference RNA (siRNA) blunted the effects of HRG on the permeability barrier. ErbB3 is known as a kinase-dead receptor and relies on other members of the family for its phosphorylation. To identify its heterodimerization partner, we knocked down the expression of other ErbB family receptors. We found that HRG's effect on the permeability barrier could be significantly attenuated by transfecting cells with ErbB2 siRNA but not with EGFR siRNA.

CONCLUSIONS

These results indicate that HRG activation of ErbB2/ErbB3 heterodimers is essential for regulation of the permeability barrier in AECs.

Neuron-derived neuregulins have been implicated in the regulation of glial cell function and survival. This factor family and its receptors may therefore be assumed to be of importance for the cellular response to traumatic injury. In this study we have examined the distribution of mRNA for neuregulin 1 (NRG1), ErbB3 and ErbB4-receptor tyrosine kinases after a ventral funiculus lesion in the lumbar spinal cord (VFL). The techniques used were in situ hybridization and immunohistochemistry. The survival times were 1-21 days. The spinal cords from normal adult and embryonic rats were used as controls. For comparison, sections from the olfactory bulb of perinatal and adult rats were also included in the study. Expression of NRG1 mRNA was observed in motoneurons in the intact spinal cord. A decrease in the labeling for NRG1 mRNA was seen during the first 5 days after VFL but then became slightly upregulated at 3 weeks after the lesion. A high labeling signal for ErbB3-mRNA was observed in the ventral and dorsal roots of E16 and E18 embryos. Labeling for ErbB3-mRNA was strong in the affected ventral root at 3 days after the VFL, reached a maximum at 1 week and was still upregulated after 3 weeks. Increased labeling for ErbB3 was also noted in scattered cells in the scar tissue 1-3 weeks after the VFL. These findings were verified with immunohistochemistry for ErbB3. A strong labeling for ErbB3 in the olfactory nerve fiber layer and olfactory nerve bundles was observed in rats of all ages examined. ErbB4 had strong expression in the embryonic spinal cord, but no evidence for lesion-induced regulation of ErbB4 receptors could be found after the VFL. Our data show that ErbB3 in the ventral roots was upregulated after a VFL and that NRG1 mRNA was initially downregulated in the motoneurons. The lesion-induced changes in the expression of NRG1 and ErbB3 in the injured spinal cord and denervated ventral root can be assumed to be of importance for axonal growth and the regulation of glial cell survival.

ErbB4 is unusual among receptor tyrosine kinases because some isoforms can be efficiently cleaved at the plasma membrane to release a soluble intracellular domain. The cleavage product has high kinase activity and homes to the nucleus. A screen for proteins that associate with the ErbB4 intracellular domain identified candidate interactors including ITCH, WWP2, Nucleolin, and Krab-associated protein 1 (Kap1). Kap1 binds to multiple isoforms of ErbB4 but does not require ErbB4 kinase activity for binding, nor is it an ErbB4 substrate. Kap1 reduces ERBB4 transcription and either directly or indirectly modulates the expression of genes that are themselves regulated by ErbB4. Upregulation of ErbB4 and suppression of MDM2 jointly enhance and accelerate the accumulation of p21(CIP1) in response to DNA damage. Overall, these findings further substantiate the role of ErbB4 in conjoint regulation of growth factor signaling and DNA damage responses.

The four members of the ErbB family of receptor tyrosine kinases are involved in development and tumorigenesis of the mammary gland. Whereas the epidermal growth factor receptor, ErbB2 and ErbB3 are positively associated with various cancers, clinical studies of ErbB4 in breast cancer are contradictory. Results from tissue culture analyses and some clinical studies suggested that ErbB4 is either a tumor suppressor or is a negative regulator of ErbB2-driven tumors. Neu-Cre-ErbB4(flox/null) mice in which ErbB4 was inactivated by Cre-lox-mediated recombination in the mammary gland developed MMTV-Neu-driven mammary tumors with a similar latency period to mice with one or two wild-type ErbB4 alleles. Moreover, there was no difference in the histologies of tumors that developed, nor in the propensity to form lung metastases. Taken together these results suggest that ErbB4 is not a potent, highly penetrant tumor suppressor, nor is it a factor in Neu-mediated tumorigenesis in this model.

The Neuregulin gene family encodes EGF-containing ligands which mediate their effects by binding to the ERBB receptor tyrosine kinases, a signalling network with important roles in both mammary gland development and breast cancer. Neuregulin3 (NRG3), a ligand for ERBB4, promotes early mammary morphogenesis and acts during specification of the mammary placode, an aggregate of epithelial cells that forms during mid-embryogenesis. Recent studies have shown that NRG3 can alter the cell fate of other epidermal progenitor populations when NRG3 is mis-expressed throughout the basal layer of the developing epidermis with the K14 promoter. Here evidence for a key function for NRG3 in promoting early mammary morphogenesis and the implication for the role of NRG3 in breast cancer and establishment of the mammary lineage are discussed.

Epidermal growth factor (EGF) superfamily of peptide growth factors (EGF-GFs) plays a role in male germ cell development, but the precise function is yet to be defined. The present study shows that EGF-GFs stimulate spermatogonial proliferation in vitro. The EGF-GF ligands, EGF, transforming growth factor-alpha and betacellulin all stimulated DNA synthesis in microdissected stage I segments of rat testis seminiferous tubules in vitro, as revealed by 3H-thymidine incorporation and 5-bromo-2'-deoxyuridine (BrdU) labeling. A fourfold increase over control of BrdU labeled cells, identified as spermatogonia, was seen after treatment with EGF. RT-PCR analysis revealed that the EGF receptors erbB1, erbB2, erbB3 and erbB4 were expressed at all stages of the spermatogenic wave, whereas differential expression was found in isolated Leydig, Sertoli and peritubular cells. The results show that EGF-GFs is spermatogonial growth factor(s) in vitro, although we have not discriminated between a direct action and an indirect effect via somatic cells. We suggest that EGF-GFs is involved in the paracrine control of spermatogenesis in vivo.

BACKGROUND

The vast majority of human genes (>70%) are alternatively spliced. Although alternative pre-mRNA processing is modified in multiple tumors, alternative hyper-splicing signatures specific to particular tumor types are still lacking. Here, we report the use of Affymetrix Human Exon Arrays to spot hyper-splicing events characteristic of myasthenia gravis (MG)-thymoma, thymic tumors which develop in patients with MG and discriminate them from colon cancer changes.

RESULTS

We combined GO term to parent threshold-based and threshold-independent ad-hoc functional statistics with in-depth analysis of key modified transcripts to highlight various exon-specific changes. These denote alternative splicing in MG-thymoma tumors compared to healthy human thymus and to in-house and Affymetrix datasets from colon cancer and healthy tissues. By using both global and specific, term-to-parent Gene Ontology (GO) statistical comparisons, our functional integrative ad-hoc method allowed the detection of disease-relevant splicing events.

CONCLUSIONS

Hyper-spliced transcripts spanned several categories, including the tumorogenic ERBB4 tyrosine kinase receptor and the connective tissue growth factor CTGF, as well as the immune function-related histocompatibility gene HLA-DRB1 and interleukin (IL)19, two muscle-specific collagens and one myosin heavy chain gene; intriguingly, a putative new exon was discovered in the MG-involved acetylcholinesterase ACHE gene. Corresponding changes in spliceosome composition were indicated by co-decreases in the splicing factors ASF/SF(2) and SC35. Parallel tumor-associated changes occurred in colon cancer as well, but the majority of the apparent hyper-splicing events were particular to MG-thymoma and could be validated by Fluorescent In-Situ Hybridization (FISH), Reverse Transcription-Polymerase Chain Reaction (RT-PCR) and mass spectrometry (MS) followed by peptide sequencing. Our findings demonstrate a particular alternative hyper-splicing signature for transcripts over-expressed in MG-thymoma, supporting the hypothesis that alternative hyper-splicing contributes to shaping the biological functions of these and other specialized tumors and opening new venues for the development of diagnosis and treatment approaches.

The intracellular domain of ErbB4 receptor tyrosine kinase is known to translocate to the nucleus of cells where it can regulate p53 transcriptional activity. The purpose of this study was to examine whether ErbB4 can localize to the nucleus of adult rat ventricular myocytes (ARVM), and regulate p53 in these cells. We demonstrate that ErbB4 does locate to the nucleus of cardiac myocytes as a full-length protein, although nuclear location occurs as a full-length protein that does not require Protein Kinase C or γ-secretase activity. Consistent with this we found that only the non-cleavable JM-b isoform of ErbB4 is expressed in ARVM. Doxorubicin was used to examine ErbB4 role in regulation of a DNA damage response in ARVM. Doxorubicin induced p53 and p21 was suppressed by treatment with AG1478, an EGFR and ErbB4 kinase inhibitor, or suppression of ErbB4 expression with small interfering RNA. Thus ErbB4 localizes to the nucleus as a full-length protein, and plays a role in the DNA damage response induced by doxorubicin in cardiac myocytes.

OBJECTIVE

ErbB receptor proteins are transmembrane tyrosine kinase receptors; when they are activated by interaction with ligands, they generate diverse cellular responses, especially during lesion development and progression to cancer. In this study the expression of ErbB receptors and TGF-alpha were investigated using an experimental cirrhosis rat model giving rise to hepatocellular neoplasms, similar to human liver diseases.

METHODS

Fifty three male rats received intraperitoneal injection of diethylnitrosamine (DEN, 50 mg/kg), weekly for 18 weeks. Until the eighth week, two rats were sacrificed every two weeks and from the tenth to the eighteenth week, five rats were sacrificed weekly. Grossly, dyschromatic and dysmorphic nodules were counted and categorized into three groups: N1/N2/N3: 3 mm < or = x < 5 mm/5 mm < or = x < 10 mm/x>> or = 10 mm in diameter. All nodules were examined, histologically. Antibodies for GSTp, TGF-alpha, EGF-R, ErbB2, ErbB3 and ErbB4 were used for immunohistochemistry.

RESULTS

The onset of cirrhoses was noted from the twelfth week. Preneoplastic foci, hepatocellular adenomas (HCA) and hepatocellular carcinomas (HCC) were noted from the second, eleventh and fifteenth week, respectively. The nodules (N1/N2/N3: 397/258/64) included regenerating nodule; RN (N1/N2/N3: 72.3%/15.9%/0%), HCA (N1/N2/N3: 27.2%/82.2%/7.6%) and HCC (N1/N2/N3: 0.5%/ 1.9%/92.4%). EGF-R was expressed in 12.5% of RN, 64.7% HCA and 75.2% HCC. TGF-alpha was expressed in 92.4% of RN, 91.3% HCA and 93.2% HCC. Sixty eight percent of TGF-alpha expressing nodules showed concurrent EGF-R expression. ErbB2 was expressed in 83.6% of RN, 72.9% HCA and 88.7% HCC. ErbB4 was expressed in 95.2% of RN, 86.3% HCA and 62.5% HCC.

CONCLUSIONS

Increased expression of EGF-R and decreased expression of ErbB4, might be related with tumor progression during DEN-induced hepatocarcinogenesis.

Enhanced NRG1-ERBB4 signaling is a risk pathway in schizophrenia, and corresponding mouse models display several endophenotypes of the disease. Nonetheless, pathway-directed treatment strategies with clinically applicable compounds have not been identified. Here, we applied a cell-based assay using the split TEV technology to screen a library of clinically applicable compounds to identify modulators of NRG1-ERBB4 signaling for repurposing. We recovered spironolactone, known as antagonist of corticosteroids, as an inhibitor of the ERBB4 receptor and tested it in pharmacological and biochemical assays to assess secondary compound actions. Transgenic mice overexpressing Nrg1 type III display cortical Erbb4 hyperphosphorylation, a condition observed in postmortem brains from schizophrenia patients. Spironolactone treatment reverted hyperphosphorylation of activated Erbb4 in these mice. In behavioral tests, spironolactone treatment of Nrg1 type III transgenic mice ameliorated schizophrenia-relevant behavioral endophenotypes, such as reduced sensorimotor gating, hyperactivity, and impaired working memory. Moreover, spironolactone increases spontaneous inhibitory postsynaptic currents in cortical slices supporting an ERBB4-mediated mode-of-action. Our findings suggest that spironolactone, a clinically safe drug, provides an opportunity for new treatment options for schizophrenia.

Cellular transformation occurs only in cells that express both ErbB1 and ErbB4 receptors, but not in cells expressing only one or the other of these receptors. However, when both receptors are coexpressed and ligand-stimulated, they interact with virtually the same adaptor/effector proteins as when expressed singly. To reveal the underlying regulatory mechanism of the kinase/phosphatase network in ErbB homo- and heterodimer receptor signaling, extracellular signal-regulated kinase (ERK) and Akt activities were evaluated in the presence of several enzyme inhibitors in ligand-induced cells expressing ErbB1 (E1), ErbB4 (E4), and ErbB1/ErbB4 (E1/4) receptor. The PP2A inhibitor okadaic acid showed receptor-specific inhibitory profiles for ERK and Akt activities. Moreover, B-Raf isolated only from E1/4 cells could induce in vitro phosphorylation for MEK; this B-Raf kinase activity was abolished by pretreatment of the cells with okadaic acid. Our study further showed that the E1/4 cell-specific B-Raf activity was stimulated by PLC gamma and subsequent Rap1 activation. The present study suggests that B-Raf kinase, which was specifically activated in the cells coexpressing ErbB1 and ErbB4 receptors, elevates total ERK activity within the cell and, therefore, can induce cellular transformation.

Neuregulin, previously known as ARIA, is a signaling protein involved in cell survival, synaptic plasticity, cell communication and differentiation. Neuregulin has also been described as a potent inducer of acetylcholine receptor transcription in muscle and although both neuregulin and acetylcholine have been individually described to have neuroprotective roles, their relationship in the cholinergic anti-inflammatory pathway of the brain has not been examined. Using three cell lines, BV-2, EOC-20 and RAW 264.7, we investigated the role that neuregulin signaling through the Erb family of tyrosine kinases may play in the anti-inflammatory process mediated by the α7 nicotinic acetylcholine receptors. Here we show that ErbB4 is expressed in all of our cell lines and is phosphorylated upon treatment with neuregulin. Neuregulin treatment further increases the expression of α7 nicotinic acetylcholine receptors in the microglial lines tested. Given the central role of α7 nicotinic acetylcholine receptors in regulating system inflammation we analyzed the expression of several pro-inflammatory cytokines in our system. Using ELISAs for TNF-α and IL-6 we show that treatment with NRG can produce a nearly a 33% decrease in the levels of tumor necrosis factor-α secreted by activated microglia and a nearly 88% decrease in IL-6. Given these results we propose a neuroprotective role for neuregulin wherein it modulates the expression of TNF-α and thus inflammation in the CNS via the upregulation of α7 nicotinic acetylcholine receptor expression in microglia in vitro. We suggest that the disregulation of neuregulin expression may be pivotal in neurological disorders characterized by inflammation.

Betacellulin (BTC), a new member of the EGF family, has been reported to be a potent mitogen for rat vascular smooth muscle cells (SMCs). BTC mRNA is known to be expressed in several human organs. However, the localization of BTC in human vascular tissues has not yet been clarified. We investigated whether or not BTC protein is involved in the pathogenesis of human atherosclerosis. Recombinant human BTC showed a mitogenic activity on cultured human aortic SMCs by measuring [3H]thymidine incorporation. The immunohistochemical localization of BTC, SMCs, macrophages, EGF receptors and ErbB4 was examined in autopsied human aortas. BTC was detected in both intimal and medial SMCs of the aortic wall. The percentage of BTC-positive medial SMCs in early types of atherosclerotic lesions decreased with age, but in adult, it was significantly higher in advanced types than in early types of atherosclerotic lesions. BTC-positive SMCs were predominantly localized in the medial side of the intima. Furthermore, numerous BTC-positive SMCs and macrophages were observed around the core lesion of atherosclerotic plaques. Receptors for BTC, EGF receptor and ErbB4, were expressed on SMCs, suggesting that BTC is associated with EGF receptor family-mediated signaling. BTC is produced in human aortic tissue and might play important roles in atherogenesis.

Recent human genetic studies and postmortem brain examinations of schizophrenia patients strongly indicate that dysregulation of NRG1 and ErbB4 may be important pathogenic factors of schizophrenia. However, this hypothesis has not been validated and fully investigated in animal models of schizophrenia. In this study we quantitatively examined NRG1 and ErbB4 protein expressions by immunohistochemistry and Western blot in the brain of a rat schizophrenia model induced by chronic administration of MK-801 (a noncompetitive NMDA receptor antagonist). Our data showed that NRG1beta and ErbB4 expressions were significantly increased in the rat prefrontal cortex and hippocampus but in different subregions. These findings suggest that altered expressions of NRG1 and ErbB4 might be attributed to the schizophrenia. Further study in the role and mechanism of NRG1 and ErbB4 may lead to better understanding of the pathophysiology for this disorder.

Growth factors and estrogen receptor (ER) signaling cooperate to play essential roles in cell proliferation, differentiation and tumor progression in mouse reproductive organs. Treatment of neonatal mice with diethylstilbestrol (DES) induces an estrogen-independent persistent proliferation and cornification of the vaginal epithelium, which results in cancerous lesions later in life. However, the mechanisms of the estrogen-dependent and -independent pathways essentially remain unknown. We characterized the expression of epidermal growth factor (EGF)-like growth factors (EGF, transforming growth factor alpha (TGF-alpha), heparin-binding EGF-like growth factor (HB-EGF), betacellulin (BTC), amphiregulin (APR), epiregulin (EPR) and neuregulin (NRG) 1) and erbB receptors (EGF receptor (EGFR), erbB2/neu, erbB3 and erbB4) in the vaginae of mice treated either neonatally (0-4 day) or as adults (55-59 day) with estrogens. EGFR and erbB2 were activated in the vaginal epithelium of mice by estrogen treatment. This activation was also encountered in vaginae from neonatally DES-exposed mice, along with the expression of EGF, TGF-alpha, HB-EGF, BTC, APR, EPR and NRG1. Immunohistochemical analysis indicated that erbB2 was primarily expressed in vaginal epithelium. Finally, we found that serine 118 and 167 located in the AF-1 domain of ERalpha were phosphorylated in these vaginae. AG825, AG1478 or ICI 182,780 administration blocked proliferation of vaginal epithelium induced by neonatal DES exposure. Thus, signal transduction via EGFR and erbB2 could be related to the estrogen-induced vaginal changes and persistent erbBs phosphorylation and sustained expression of EGF-like growth factors, leading to ERalpha activation that may result in cancerous lesions in vaginae from neonatally DES-exposed mice later in life.

OBJECTIVE

The molecular events in the malignant progression of colon adenoma after loss of adenomatous polyposis coli (APC) are not fully understood. KITENIN (KAI1 C-terminal interacting tetraspanin) increases the invasiveness of colorectal cancer cells, and we identified a novel EGFR-independent oncogenic signal of EGF that works under coexpressed KITENIN and ErbB4. Here we tested whether elevated KITENIN and ErbB4 contribute to further progression of intestinal adenoma following APC loss.

RESULTS

The intestinal tissues of villin-KITENIN transgenic mice in which villin-driven KITENIN expression induces increased c-Jun expression exhibit mild epithelial cell proliferation but no epithelial lineage changes compared with those of nontransgenic mice. Among the four ErbB4 isoforms, JM-a/CYT-2 and JM-b/CYT-2 exhibited the highest AP-1 activity when cells coexpressing KITENIN and each isoform were stimulated by EGF. Interestingly, predominant overexpression of the ErB4-CYT-2 mRNA as well as increased EGFR expression were observed in intestinal adenoma of APC(min/+) mice, which makes the microenvironment of activated EGF signaling. When we crossed villin-KITENIN mice with APC(min/+) mice, intestinal tumor tissues in the crossed mice showed the characteristics of early-stage invading adenocarcinoma. In patients with colorectal cancer, ErbB4-CYT-2 mRNA expression was significantly greater in tumor tissues than in normal adjacent tissues, but no significant differences in tumor tissue expression were found between different colorectal cancer stages. Furthermore, the mRNA expression of KITENIN and that of ErbB4-CYT-2 were positively correlated in human colorectal cancer tissue.

CONCLUSIONS

Elevated coexpression of KITENIN and ErbB4-CYT-2 promotes the transition of colon adenoma to adenocarcinoma within an APC loss-associated tumor microenvironment.

Neuregulin 1 (NRG1) is a trophic factor that is thought to have important roles in the regulating brain circuitry. Recent studies suggest that NRG1 regulates synaptic transmission, although the precise mechanisms remain unknown. Here we report that NRG1 influences glutamate uptake by increasing the protein level of excitatory amino acid carrier (EAAC1). Our data indicate that NRG1 induced the up-regulation of EAAC1 in primary cortical neurons with an increase in glutamate uptake. These in vitro results were corroborated in the prefrontal cortex (PFC) of mice given NRG1. The stimulatory effect of NRG1 was blocked by inhibition of the NRG1 receptor ErbB4. The suppressed expression of ErbB4 by siRNA led to a decrease in the expression of EAAC1. In addition, the ablation of ErbB4 in parvalbumin (PV)-positive neurons in PV-ErbB4(-/-) mice suppressed EAAC1 expression. Taken together, our results show that NRG1 signaling through ErbB4 modulates EAAC1. These findings link proposed effectors in schizophrenia: NRG1/ErbB4 signaling perturbation, EAAC1 deficit, and neurotransmission dysfunction.

Numerous genetic and functional studies implicate variants of Neuregulin-1 (NRG1) and its neuronal receptor ErbB4 in schizophrenia and many of its endophenotypes. Although the neurophysiological and behavioral phenotypes of NRG1 mutant mice have been investigated extensively, practically nothing is known about the function of NRG2, the closest NRG1 homolog. We found that NRG2 expression in the adult rodent brain does not overlap with NRG1 and is more extensive than originally reported, including expression in the striatum and medial prefrontal cortex (mPFC), and therefore generated NRG2 knockout mice (KO) to study its function. NRG2 KOs have higher extracellular dopamine levels in the dorsal striatum but lower levels in the mPFC; a pattern with similarities to dopamine dysbalance in schizophrenia. Like ErbB4 KO mice, NRG2 KOs performed abnormally in a battery of behavioral tasks relevant to psychiatric disorders. NRG2 KOs exhibit hyperactivity in a novelty-induced open field, deficits in prepulse inhibition, hypersensitivity to amphetamine, antisocial behaviors, reduced anxiety-like behavior in the elevated plus maze and deficits in the T-maze alteration reward test-a task dependent on hippocampal and mPFC function. Acute administration of clozapine rapidly increased extracellular dopamine levels in the mPFC and improved alternation T-maze performance. Similar to mice treated chronically with N-methyl-d-aspartate receptor (NMDAR) antagonists, we demonstrate that NMDAR synaptic currents in NRG2 KOs are augmented at hippocampal glutamatergic synapses and are more sensitive to ifenprodil, indicating an increased contribution of GluN2B-containing NMDARs. Our findings reveal a novel role for NRG2 in the modulation of behaviors with relevance to psychiatric disorders.

ErbB family members, ErbB1/EGFR/HER-1, ErbB2/HER-2, ErbB3/HER-3 and ErbB4/HER-4, have been implicated in breast cancer (BC) tumorigenicity. Recently, crucial roles for RANK/RANKL signaling in addition to key downstream factor NF-κB have been demonstrated in mammary tumorigenesis. Here, we present the hypothesis of a novel association between ErbB and RANK pathways in promoting BC. The proposed model alludes to the cross-talk that might occur between RANK and ErbB receptors. This interplay might regulate RANK signaling and consequently, modulate carcinogenesis, mainly in ErbB2 over-expressing BC cells. Thus, we highlight the significance of the RANK/RANKL axis as a putative therapeutic target in this malignancy, and furthermore, suggest that the combination of ErbB and RANK/RANKL inhibitors may have therapeutic benefit for certain BC patients.

Obesity reduces breastfeeding success and lactation performance in women. However, the mechanisms involved are not entirely understood. In the present study, female C57BL/6 mice were chronically exposed to a high-fat diet to induce obesity and subsequently exhibited impaired offspring viability (only 15% survival rate), milk production (33% reduction), mammopoiesis (one-third of the glandular area compared to control animals) and postpartum maternal behaviors (higher latency to retrieving and grouping the pups). Reproductive experience attenuated these defects. Diet-induced obese mice exhibited high basal pSTAT5 levels in the mammary tissue and hypothalamus, and an acute prolactin stimulus was unable to further increase pSTAT5 levels above basal levels. In contrast, genetically obese leptin-deficient females showed normal prolactin responsiveness. Additionally, we identified the expression of leptin receptors specifically in basal/myoepithelial cells of the mouse mammary gland. Finally, high-fat diet females exhibited altered mRNA levels of ERBB4 and NRG1, suggesting that obesity may involve disturbances to mammary gland paracrine circuits that are critical in the control of luminal progenitor function and lactation. In summary, our findings indicate that high leptin levels are a possible cause of the peripheral and central prolactin resistance observed in obese mice which leads to impaired lactation performance.