👤 Noelia Garcia

The octadecadienoic conjugated linoleic acid (CLA) isomer with trans-11 and cis-13 double bonds (trans-11,cis-13 CLA) has been described in ruminant milk. For now, this specific CLA is suspected to derive exclusively from ruminal biohydrogenation of dietary α-linolenic acid. However, in rodents, the fatty acid desaturase 3 (FADS3) gene was recently shown to code for an enzyme able to catalyze the unexpected Δ13-desaturation of vaccenic acid, producing a Δ11,13-CLA with all the structural characteristics of the trans-11,cis-13 isomer, although no commercial standard exists for complete conclusive identification. Because the FADS3 gene has already been reported in bovine animals, we hypothesized in the present study that an alternative direct FADS3-catalyzed Δ13-desaturation of vaccenic acid in mammary tissue may therefore co-exist with α-linolenic acid biohydrogenation to explain the final ruminant milk trans-11,cis-13 CLA presence. Here, we first confirm that the FADS3 gene is present in ruminant mammal genomic sequence databases. Second, we demonstrate that the Δ11,13-CLA found in milk fat and the highly probable trans-11,cis-13 CLA isomer produced by rodent FADS3 possess exactly the same structural characteristics. Then, we show that bovine mammary MAC-T and BME-UV epithelial cells express both FADS3 and stearoyl-CoA desaturase 1 (SCD1) mRNA and are able to synthesize both the suspected trans-11,cis-13 CLA and cis-9,trans-11CLA (rumenic acid) isomers when incubated with vaccenic acid. Finally, the concomitant presence of the suspected trans-11,cis-13 CLA isomer with FADS3 mRNA was shown in goat mammary tissue, whereas both were conversely very low or even absent in goat liver. Therefore, this study provides several lines of evidence that, by analogy with rumenic acid, trans-11,cis-13 CLA may originate both from ruminal biohydrogenation and from direct FADS3-catalyzed Δ13-desaturation of vaccenic acid in mammary tissue. Show less

In pediatric acute leukemias, reciprocal chromosomal translocations frequently cause gene fusions involving the lysine (K)-specific methyltransferase 2A gene (KMT2A, also known as MLL). Specific KMT2A fusion partners are associated with the disease phenotype (lymphoblastic vs. myeloid), and the type of KMT2A rearrangement also has prognostic implications. However, the KMT2A partner gene cannot always be identified by banding karyotyping. We sought to identify such partner genes in 13 cases of childhood leukemia with uninformative karyotypes by combining molecular techniques, including multicolor banding FISH, reverse-transcriptase PCR, and long-distance inverse PCR. Of the KMT2A fusion partner genes, MLLT3 was present in five patients, all with acute lymphoblastic leukemia, MLLT1 in two patients, and MLLT10, MLLT4, MLLT11, and AFF1 in one patient each. Reciprocal reading by long-distance inverse PCR also disclosed KMT2A fusions with PITPNA in one patient, with LOC100132273 in another patient, and with DNA sequences not compatible with any gene in three patients. The most common KMT2A breakpoint region was intron/exon 9 (3/8 patients), followed by intron/exon 11 and 10. Finally, multicolor banding revealed breakpoints in other chromosomes whose biological and prognostic implications remain to be determined. We conclude that the combination of molecular techniques used in this study can efficiently identify KMT2A fusion partners in complex pediatric acute leukemia karyotypes. Copyright © 2016 John Wiley & Sons, Ltd. Show less

AF10, a DOT1L cofactor, is required for H3K79 methylation and cooperates with DOT1L in leukemogenesis. However, the molecular mechanism by which AF10 regulates DOT1L-mediated H3K79 methylation is not clear. Here we report that AF10 contains a "reader" domain that couples unmodified H3K27 recognition to H3K79 methylation. An AF10 region consisting of a PHD finger-Zn knuckle-PHD finger (PZP) folds into a single module that recognizes amino acids 22-27 of H3, and this interaction is abrogated by H3K27 modification. Structural studies reveal that H3 binding triggers rearrangement of the PZP module to form an H3(22-27)-accommodating channel and that the unmodified H3K27 side chain is encased in a compact hydrogen-bond acceptor-lined cage. In cells, PZP recognition of H3 is required for H3K79 dimethylation, expression of DOT1L-target genes, and proliferation of DOT1L-addicted leukemic cells. Together, our results uncover a pivotal role for H3K27-via readout by the AF10 PZP domain-in regulating the cancer-associated enzyme DOT1L. Show less

Coffee, a major dietary source of caffeine, is among the most widely consumed beverages in the world and has received considerable attention regarding health risks and benefits. We conducted a genome-wide (GW) meta-analysis of predominately regular-type coffee consumption (cups per day) among up to 91,462 coffee consumers of European ancestry with top single-nucleotide polymorphisms (SNPs) followed-up in ~30 062 and 7964 coffee consumers of European and African-American ancestry, respectively. Studies from both stages were combined in a trans-ethnic meta-analysis. Confirmed loci were examined for putative functional and biological relevance. Eight loci, including six novel loci, met GW significance (log10Bayes factor (BF)>5.64) with per-allele effect sizes of 0.03-0.14 cups per day. Six are located in or near genes potentially involved in pharmacokinetics (ABCG2, AHR, POR and CYP1A2) and pharmacodynamics (BDNF and SLC6A4) of caffeine. Two map to GCKR and MLXIPL genes related to metabolic traits but lacking known roles in coffee consumption. Enhancer and promoter histone marks populate the regions of many confirmed loci and several potential regulatory SNPs are highly correlated with the lead SNP of each. SNP alleles near GCKR, MLXIPL, BDNF and CYP1A2 that were associated with higher coffee consumption have previously been associated with smoking initiation, higher adiposity and fasting insulin and glucose but lower blood pressure and favorable lipid, inflammatory and liver enzyme profiles (P<5 × 10(-8)).Our genetic findings among European and African-American adults reinforce the role of caffeine in mediating habitual coffee consumption and may point to molecular mechanisms underlying inter-individual variability in pharmacological and health effects of coffee. Show less

Forced vital capacity (FVC), a spirometric measure of pulmonary function, reflects lung volume and is used to diagnose and monitor lung diseases. We performed genome-wide association study meta-analysis of FVC in 52,253 individuals from 26 studies and followed up the top associations in 32,917 additional individuals of European ancestry. We found six new regions associated at genome-wide significance (P < 5 × 10(-8)) with FVC in or near EFEMP1, BMP6, MIR129-2-HSD17B12, PRDM11, WWOX and KCNJ2. Two loci previously associated with spirometric measures (GSTCD and PTCH1) were related to FVC. Newly implicated regions were followed up in samples from African-American, Korean, Chinese and Hispanic individuals. We detected transcripts for all six newly implicated genes in human lung tissue. The new loci may inform mechanisms involved in lung development and the pathogenesis of restrictive lung disease. Show less

RNA trans-splicing has been explored as a therapeutic option for a variety of genetic diseases, but not for cardiac genetic disease. Hypertrophic cardiomyopathy (HCM) is an autosomal-dominant disease, characterized by left ventricular hypertrophy (LVH) and diastolic dysfunction. MYBPC3, encoding cardiac myosin-binding protein C (cMyBP-C) is frequently mutated. We evaluated the 5'-trans-splicing strategy in a mouse model of HCM carrying a Mybpc3 mutation. 5'-trans-splicing was induced between two independently transcribed molecules, the mutant endogenous Mypbc3 pre-mRNA and an engineered pre-trans-splicing molecule (PTM) carrying a FLAG-tagged wild-type (WT) Mybpc3 cDNA sequence. PTMs were packaged into adeno-associated virus (AAV) for transduction of cultured cardiac myocytes and the heart in vivo. Full-length repaired Mybpc3 mRNA represented up to 66% of total Mybpc3 transcripts in cardiac myocytes and 0.14% in the heart. Repaired cMyBP-C protein was detected by immunoprecipitation in cells and in vivo and exhibited correct incorporation into the sarcomere in cardiac myocytes. This study provides (i) the first evidence of successful 5'-trans-splicing in vivo and (ii) proof-of-concept of mRNA repair in the most prevalent cardiac genetic disease. Since current therapeutic options for HCM only alleviate symptoms, these findings open new horizons for causal therapy of the severe forms of the disease.Molecular Therapy-Nucleic Acids (2013) 2, e102; doi:10.1038/mtna.2013.31; published online 2 July 2013. Show less

Exon skipping mediated by antisense oligoribonucleotides (AON) is a promising therapeutic approach for genetic disorders, but has not yet been evaluated for cardiac diseases. We investigated the feasibility and efficacy of viral-mediated AON transfer in a Mybpc3-targeted knock-in (KI) mouse model of hypertrophic cardiomyopathy (HCM). KI mice carry a homozygous G>A transition in exon 6, which results in three different aberrant mRNAs. We identified an alternative variant (Var-4) deleted of exons 5-6 in wild-type and KI mice. To enhance its expression and suppress aberrant mRNAs we designed AON-5 and AON-6 that mask splicing enhancer motifs in exons 5 and 6. AONs were inserted into modified U7 small nuclear RNA and packaged in adeno-associated virus (AAV-U7-AON-5+6). Transduction of cardiac myocytes or systemic administration of AAV-U7-AON-5+6 increased Var-4 mRNA/protein levels and reduced aberrant mRNAs. Injection of newborn KI mice abolished cardiac dysfunction and prevented left ventricular hypertrophy. Although the therapeutic effect was transient and therefore requires optimization to be maintained over an extended period, this proof-of-concept study paves the way towards a causal therapy of HCM. Show less

Most genome-wide association studies are confined to middle-aged populations. It is unclear whether associations between single nucleotide polymorphisms (SNPs) and obesity persist in old age. We aimed to relate 10 body mass index (BMI)-associated SNPs to weight, BMI, % fat, visceral and subcutaneous adipose tissue in Health ABC and AGES-Reykjavik comprising 4,846 individuals of European Ancestry, and 1,139 African Americans over age 65. SNPs were scaled using effect estimates from candidate SNPs. In Health ABC, a SNP near GNPDA2 was modestly associated with weight and SAT area (p = .008, p = .001). Risk score (sum of scaled SNPs) was associated with weight, BMI, and SAT area (p < .0001 for all), but neither GNPDA2 nor risk score was associated with weight, BMI, visceral adippose tissue, subcutaneous adipose tissue, or % fat in AGES-Reykjavik. In African Americans, a SNP near SEC16B was weakly associated with weight (p = .04). In this sample of older adults, no BMI-associated SNPs were associated with weight or adiposity. Show less

Histone post-translational modifications (PTMs) constitute a branch of epigenetic mechanisms that can control the expression of eukaryotic genes in a heritable manner. Recent studies have identified several PTM-binding proteins containing diverse specialized domains whose recognition of specific PTM sites leads to gene activation or repression. Here, we present a high-throughput proteogenomic platform designed to characterize the nucleosomal make-up of chromatin enriched with a set of histone PTM binding proteins known as histone PTM readers. We support our findings with gene expression data correlating to PTM distribution. We isolated human mononucleosomes bound by the bromodomain-containing proteins Brd2, Brd3 and Brd4, and by the chromodomain-containing heterochromatin proteins HP1β and HP1α. Histone PTMs were quantified by mass spectrometry (ChIP-qMS), and their associated DNAs were mapped using deep sequencing. Our results reveal that Brd- and HP1-bound nucleosomes are enriched in histone PTMs consistent with actively transcribed euchromatin and silent heterochromatin, respectively. Data collected using RNA-Seq show that Brd-bound sites correlate with highly expressed genes. In particular, Brd3 and Brd4 are most enriched on nucleosomes located within HOX gene clusters, whose expression is reduced upon Brd4 depletion by short hairpin RNA. Proteogenomic mapping of histone PTM readers, alongside the characterization of their local chromatin environments and transcriptional information, should prove useful for determining how histone PTMs are bound by these readers and how they contribute to distinct transcriptional states. Show less

Sex hormone-binding globulin (SHBG) is a glycoprotein responsible for the transport and biologic availability of sex steroid hormones, primarily testosterone and estradiol. SHBG has been associated with chronic diseases including type 2 diabetes (T2D) and with hormone-sensitive cancers such as breast and prostate cancer. We performed a genome-wide association study (GWAS) meta-analysis of 21,791 individuals from 10 epidemiologic studies and validated these findings in 7,046 individuals in an additional six studies. We identified twelve genomic regions (SNPs) associated with circulating SHBG concentrations. Loci near the identified SNPs included SHBG (rs12150660, 17p13.1, p = 1.8 × 10(-106)), PRMT6 (rs17496332, 1p13.3, p = 1.4 × 10(-11)), GCKR (rs780093, 2p23.3, p = 2.2 × 10(-16)), ZBTB10 (rs440837, 8q21.13, p = 3.4 × 10(-09)), JMJD1C (rs7910927, 10q21.3, p = 6.1 × 10(-35)), SLCO1B1 (rs4149056, 12p12.1, p = 1.9 × 10(-08)), NR2F2 (rs8023580, 15q26.2, p = 8.3 × 10(-12)), ZNF652 (rs2411984, 17q21.32, p = 3.5 × 10(-14)), TDGF3 (rs1573036, Xq22.3, p = 4.1 × 10(-14)), LHCGR (rs10454142, 2p16.3, p = 1.3 × 10(-07)), BAIAP2L1 (rs3779195, 7q21.3, p = 2.7 × 10(-08)), and UGT2B15 (rs293428, 4q13.2, p = 5.5 × 10(-06)). These genes encompass multiple biologic pathways, including hepatic function, lipid metabolism, carbohydrate metabolism and T2D, androgen and estrogen receptor function, epigenetic effects, and the biology of sex steroid hormone-responsive cancers including breast and prostate cancer. We found evidence of sex-differentiated genetic influences on SHBG. In a sex-specific GWAS, the loci 4q13.2-UGT2B15 was significant in men only (men p = 2.5 × 10(-08), women p = 0.66, heterogeneity p = 0.003). Additionally, three loci showed strong sex-differentiated effects: 17p13.1-SHBG and Xq22.3-TDGF3 were stronger in men, whereas 8q21.12-ZBTB10 was stronger in women. Conditional analyses identified additional signals at the SHBG gene that together almost double the proportion of variance explained at the locus. Using an independent study of 1,129 individuals, all SNPs identified in the overall or sex-differentiated or conditional analyses explained ~15.6% and ~8.4% of the genetic variation of SHBG concentrations in men and women, respectively. The evidence for sex-differentiated effects and allelic heterogeneity highlight the importance of considering these features when estimating complex trait variance. Show less

Excessive dietary fructose intake may have an important role in the current epidemics of fatty liver, obesity and diabetes as its intake parallels the development of these syndromes and because it can induce features of metabolic syndrome. The effects of fructose to induce fatty liver, hypertriglyceridemia and insulin resistance, however, vary dramatically among individuals. The first step in fructose metabolism is mediated by fructokinase (KHK), which phosphorylates fructose to fructose-1-phosphate; intracellular uric acid is also generated as a consequence of the transient ATP depletion that occurs during this reaction. Here we show in human hepatocytes that uric acid up-regulates KHK expression thus leading to the amplification of the lipogenic effects of fructose. Inhibition of uric acid production markedly blocked fructose-induced triglyceride accumulation in hepatocytes in vitro and in vivo. The mechanism whereby uric acid stimulates KHK expression involves the activation of the transcription factor ChREBP, which, in turn, results in the transcriptional activation of KHK by binding to a specific sequence within its promoter. Since subjects sensitive to fructose often develop phenotypes associated with hyperuricemia, uric acid may be an underlying factor in sensitizing hepatocytes to fructose metabolism during the development of fatty liver. Show less

Although recovery after spinal cord injury (SCI) is rare in humans, recent literature indicates that some patients do recover sensorimotor function years after the trauma. This study seeks to elucidate the genetic underpinnings of SCI repair through the investigation of neurodegenerative and regenerative associated genes involved in the response to SCI during the chronic phase in adult rats. Intervention on the level of gene regulation focused on enhancing naturally attempting SCI regenerative genes has the potential to promote SCI repair. Our aim was to analyze gene expression characteristics of candidate genes involved in the neuro-degenerative and -regenerative processes following various animal models of SCI. We compiled data showing gene expression changes after SCI in adult rats and created a chronological time-line of candidate genes differentially expressed during the chronic phase of SCI. Compiled data showed that SCI induced a transient upregulation of endogenous neuro-regenerative genes not only within a few hours but also within a few days, weeks, and months after SCI. For example, gene controlling growth-associated protein-43 (GAP-43), brain-derived neurotrophic factor (BDNF), glial cell line-derived neurotrophic factor (GDNF), and others, showed significant changes in mRNA accumulation in SCI animals, from 48 hours to 12 weeks after SCI. Similarly, inhibitory genes, such as RhoA, LINGO-1, and others, were upregulated as late as 4 to 14 days after injury. This indicates that gene specific regulation changes, corresponding to repair and regenerative attempts, are naturally orchestrated over time after injury. These delayed changes after SCI give ample time for therapeutic gene modulation through upregulation or silencing of specific genes responsible for the synthesis of the corresponding biogenic proteins. By following the examination of differential gene regulation during the chronic phase, we have determined times, successions, co-activations, interferences, and dosages for potential therapeutic synchronized interventions. Finally, local cellular specificities and their neuropathophysiologies have been taken into account in the elaboration of the combination treatment strategy we propose. The interventions we propose suggest the delivery of exogenous therapeutic agents to upregulate or downregulate chosen genes or the expression of the downstream proteins to revert the post-traumatic stage of SCI during the chronic phase. The proposed combination and schedule of local cell-specific treatment should enhance intrinsic regenerative machinery and provide a promising strategy for treating patients sustaining chronic SCI. Show less

The R820W mutation in the MYBPC3 gene has been associated with the development of hypertrophic cardiomyopathy (HCM) in rag-doll cats, but had not been described in humans. To describe the phenotype associated with the R820W mutation identified in a human family. The R820W was identified by direct sequencing of the MYBPC3 gene in a 47 year old woman with HCM and left ventricular non-compaction (LVNC). Clinical and genetic studies of the R820W mutation were performed in her family. The index patient was homozygous for the mutation and had no additional mutations in the main sarcomeric genes (MYH7, TNNT2, TNNI3, and TPM1). She had HCM with LVNC and normal systolic function. One brother had died suddenly at age 43 years. Another brother diagnosed of LVNC with severe systolic dysfunction and a cardiac arrest was also homozygous for the mutation. One heterozygous 31 year old sister, and three heterozygous sons of the index (ages 14, 20 and 23 years old) were clinically unaffected. The father of the index was apparently healthy and her mother had atrial fibrillation and an electrocardiographic diagnosis of left ventricular hypertrophy at age 86 years. The R820W mutation in the MYBPC3 gene, previously associated with HCM in rag-doll cats, causes both HCM and LVNC in homozygous human carriers, with mild or null clinical expression in heterozygous carriers. Show less

To identify loci for age at menarche, we performed a meta-analysis of 32 genome-wide association studies in 87,802 women of European descent, with replication in up to 14,731 women. In addition to the known loci at LIN28B (P = 5.4 × 10⁻⁶⁰) and 9q31.2 (P = 2.2 × 10⁻³³), we identified 30 new menarche loci (all P < 5 × 10⁻⁸) and found suggestive evidence for a further 10 loci (P < 1.9 × 10⁻⁶). The new loci included four previously associated with body mass index (in or near FTO, SEC16B, TRA2B and TMEM18), three in or near other genes implicated in energy homeostasis (BSX, CRTC1 and MCHR2) and three in or near genes implicated in hormonal regulation (INHBA, PCSK2 and RXRG). Ingenuity and gene-set enrichment pathway analyses identified coenzyme A and fatty acid biosynthesis as biological processes related to menarche timing. Show less

Induction of apoptosis by TNF has recently been shown to implicate proteases from lysosomal origin, the cathepsins. Here, we investigated the role in apoptosis of palmitoyl protein thioesterase 1 (PPT1), another lysosomal enzyme that depalmitoylates proteins. We show that transformed fibroblasts derived from patients with the infantile form of neuronal ceroid lipofuscinosis (INCL), a neurodegenerative disease due to deficient activity of PPT1, are partially resistant to TNF-induced cell death (57-75% cell viability vs. 15-30% for control fibroblasts). TNF-initiated proteolytic cleavage of caspase-8, Bid and caspase-3, as well as cytochrome c release was strongly attenuated in INCL fibroblasts as compared to control cells. Noteworthy, activation of p42/p44 mitogen-activated protein kinase and of transcription factor NF-kappaB by TNF, and induction of cell death by staurosporine or chemotherapeutic drugs in INCL cells were unaffected by PPT1 deficiency. Resistance to TNF-induced apoptosis was also observed in embryonic fibroblasts derived from Ppt1/Cln1-deficient mice but not from mice with a targeted deletion of Cln3 or Cln5. Finally, reconstitution of PPT1 activity in mutant cells was accompanied by resensitization to TNF-induced caspase activation and toxicity. These observations emphasize for the first time the role of PPT1 and, likely, protein depalmitoylation in the regulation of TNF-induced apoptosis. Show less

Central abdominal fat is a strong risk factor for diabetes and cardiovascular disease. To identify common variants influencing central abdominal fat, we conducted a two-stage genome-wide association analysis for waist circumference (WC). In total, three loci reached genome-wide significance. In stage 1, 31,373 individuals of Caucasian descent from eight cohort studies confirmed the role of FTO and MC4R and identified one novel locus associated with WC in the neurexin 3 gene [NRXN3 (rs10146997, p = 6.4x10(-7))]. The association with NRXN3 was confirmed in stage 2 by combining stage 1 results with those from 38,641 participants in the GIANT consortium (p = 0.009 in GIANT only, p = 5.3x10(-8) for combined analysis, n = 70,014). Mean WC increase per copy of the G allele was 0.0498 z-score units (0.65 cm). This SNP was also associated with body mass index (BMI) [p = 7.4x10(-6), 0.024 z-score units (0.10 kg/m(2)) per copy of the G allele] and the risk of obesity (odds ratio 1.13, 95% CI 1.07-1.19; p = 3.2x10(-5) per copy of the G allele). The NRXN3 gene has been previously implicated in addiction and reward behavior, lending further evidence that common forms of obesity may be a central nervous system-mediated disorder. Our findings establish that common variants in NRXN3 are associated with WC, BMI, and obesity. Show less

The Wnt/beta-catenin signaling pathway is critical in both cellular proliferation and organismal development. However, how the beta-catenin degradation complex is inhibited upon Wnt activation remains unclear. Using a directed RNAi screen we find that protein phosphatase 1 (PP1), a ubiquitous serine/threonine phosphatase, is a novel potent positive physiologic regulator of the Wnt/beta-catenin signaling pathway. PP1 expression synergistically activates, and inhibition of PP1 inhibits, Wnt/beta-catenin signaling in Drosophila and mammalian cells as well as in Xenopus embryos. The data suggest that PP1 controls Wnt signaling through interaction with, and regulated dephosphorylation of, axin. Inhibition of PP1 leads to enhanced phosphorylation of specific sites on axin by casein kinase I. Axin phosphorylation markedly enhances the binding of glycogen synthase kinase 3, leading to a more active beta-catenin destruction complex. Wnt-regulated changes in axin phosphorylation, mediated by PP1, may therefore determine beta-catenin transcriptional activity. Specific inhibition of PP1 in this pathway may offer therapeutic approaches to disorders with increased beta-catenin signaling. Show less

A major obstacle for successful axon regeneration in the adult central nervous system (CNS) arises from inhibitory molecules in CNS myelin, which signal through a common receptor complex on neurons consisting of the ligand-binding Nogo-66 receptor (NgR) and two transmembrane coreceptors, p75 and LINGO-1. However, p75 expression is only detectable in subpopulations of mature neurons, raising the question of how these inhibitory signals are transduced in neurons lacking p75. In this study, we demonstrate that TROY (also known as TAJ), a TNF receptor family member selectively expressed in the adult nervous system, can form a functional receptor complex with NgR and LINGO-1 to mediate cellular responses to myelin inhibitors. Also, both overexpressing a dominant-negative TROY or presence of a soluble TROY protein can efficiently block neuronal response to myelin inhibitors. Our results implicate TROY in mediating myelin inhibition, offering new insights into the molecular mechanisms of regeneration failure in the adult nervous system. Show less

Neuregulins regulate the expression of ligand- and voltage-gated channels in neurons and skeletal muscle by the activation of their cognate tyrosine kinase receptors, ErbB 1-4. The subcellular distribution and mechanisms that regulate the localization of ErbB receptors are unknown. We have found that ErbB receptors are present in brain subcellular fractions enriched for postsynaptic densities (PSD). The ErbB-4 receptor is unique among the ErbB proteins because its C-terminal tail (T-V-V) conforms to a sequence that binds to a protein motif known as the PDZ domain. Using the yeast two-hybrid system, we found that the C-terminal region of ErbB-4 interacts with the three related membrane-associated guanylate kinases (MAGUKs) PSD-95/SAP90, PSD-93/chapsyn-110, and SAP 102, which harbor three PDZ domains, as well as with beta(2)-syntrophin, which has a single PDZ domain. As with N-methyl-D-aspartate (NMDA) receptors, ErbB4 interacts with the first two PDZ domains of PSD-95. Using coimmunoprecipitation assays, we confirmed the direct interactions between ErbB-4 and PSD-95 in transfected heterologous cells, as well as in vivo, where both proteins are coimmunoprecipitated from brain lysates. Moreover, evidence for colocalization of these proteins was also observed by immunofluorescence in cultured hippocampal neurons. ErbB-4 colocalizes with PSD-95 and NMDA receptors at a subset of excitatory synapses apposed to synaptophysin-positive presynaptic terminals. The capacity of ErbB receptors to interact with PDZ-domain proteins at cell junctions is conserved from invertebrates to mammals. As discussed, the interactions found between receptor tyrosine kinases and MAGUKs at neuronal synapses may have important implications for activity-dependent plasticity. Show less