M J Radeke

CSU Mentor, Long Beach, California, United States

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Publications (45)257.61 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: Measles virus (MV) lacking expression of C protein (C(KO)) is a potent activator of the double-stranded RNA (dsRNA)-dependent protein kinase (PKR), whereas the isogenic parental virus expressing C protein is not. Here we demonstrate that significant amounts of dsRNA accumulate during C(KO) mutant infection, but not following parental virus infection. dsRNA accumulated during late stages of infection and localized with virus replication sites containing N and P proteins. PKR autophosphorylation and stress granule formation correlated with the timing of dsRNA appearance. Phospho-PKR localized to dsRNA-containing structures as revealed by immunofluorescence. Production of dsRNA was sensitive to cycloheximide but resistant to actinomycin D, suggesting that the dsRNA is a viral product. Quantitative PCR analyses revealed reduced viral RNA synthesis and a steepened transcription gradient in C(KO) compared to parental virus-infected cells. The observed alterations were further reflected in lower viral protein expression levels and reduced C(KO) virus infectious yield. RNA deep sequencing confirmed the viral RNA expression profile differences seen by qPCR between C(KO) mutant and the parental viruses. After one subsequent passage of the C(KO) virus, defective interfering RNA (DI-RNA) with duplex structure were obtained that were not seen with parental virus. We conclude that in the absence of C protein, the amount of PKR activator RNA including DI-RNA is increased, thereby triggering innate immune responses leading to impaired MV growth.
    Journal of Virology 10/2013; · 5.08 Impact Factor
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    ABSTRACT: Please see related commentary: http://www.biomedcentral.com/1741-7015/10/21/abstract Age-related macular degeneration (AMD) is a leading cause of blindness that affects the central region of the retinal pigmented epithelium (RPE), choroid, and neural retina. Initially characterized by an accumulation of sub-RPE deposits, AMD leads to progressive retinal degeneration, and in advanced cases, irreversible vision loss. Although genetic analysis, animal models, and cell culture systems have yielded important insights into AMD, the molecular pathways underlying AMD's onset and progression remain poorly delineated. We sought to better understand the molecular underpinnings of this devastating disease by performing the first comparative transcriptome analysis of AMD and normal human donor eyes. RPE-choroid and retina tissue samples were obtained from a common cohort of 31 normal, 26 AMD, and 11 potential pre-AMD human donor eyes. Transcriptome profiles were generated for macular and extramacular regions, and statistical and bioinformatic methods were employed to identify disease-associated gene signatures and functionally enriched protein association networks. Selected genes of high significance were validated using an independent donor cohort. We identified over 50 annotated genes enriched in cell-mediated immune responses that are globally over-expressed in RPE-choroid AMD phenotypes. Using a machine learning model and a second donor cohort, we show that the top 20 global genes are predictive of AMD clinical diagnosis. We also discovered functionally enriched gene sets in the RPE-choroid that delineate the advanced AMD phenotypes, neovascular AMD and geographic atrophy. Moreover, we identified a graded increase of transcript levels in the retina related to wound response, complement cascade, and neurogenesis that strongly correlates with decreased levels of phototransduction transcripts and increased AMD severity. Based on our findings, we assembled protein-protein interactomes that highlight functional networks likely to be involved in AMD pathogenesis. We discovered new global biomarkers and gene expression signatures of AMD. These results are consistent with a model whereby cell-based inflammatory responses represent a central feature of AMD etiology, and depending on genetics, environment, or stochastic factors, may give rise to the advanced AMD phenotypes characterized by angiogenesis and/or cell death. Genes regulating these immunological activities, along with numerous other genes identified here, represent promising new targets for AMD-directed therapeutics and diagnostics.
    Genome Medicine 02/2012; 4(2):16. · 4.94 Impact Factor
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    ABSTRACT: We introduce a human retinal pigmented epithelial (RPE) cell-culture model that mimics several key aspects of early stage age-related macular degeneration (AMD). These include accumulation of sub-RPE deposits that contain molecular constituents of human drusen, and activation of complement leading to formation of deposit-associated terminal complement complexes. Abundant sub-RPE deposits that are rich in apolipoprotein E (APOE), a prominent drusen constituent, are formed by RPE cells grown on porous supports. Exposure to human serum results in selective, deposit-associated accumulation of additional known drusen components, including vitronectin, clusterin, and serum amyloid P, thus suggesting that specific protein-protein interactions contribute to the accretion of plasma proteins during drusen formation. Serum exposure also leads to complement activation, as evidenced by the generation of C5b-9 immunoreactive terminal complement complexes in association with APOE-containing deposits. Ultrastructural analyses reveal two morphologically distinct forms of deposits: One consisting of membrane-bounded multivesicular material, and the other of nonmembrane-bounded particle conglomerates. Collectively, these results suggest that drusen formation involves the accumulation of sub-RPE material rich in APOE, a prominent biosynthetic product of the RPE, which interacts with a select group of drusen-associated plasma proteins. Activation of the complement cascade appears to be mediated via the classical pathway by the binding of C1q to ligands in APOE-rich deposits, triggering direct activation of complement by C1q, deposition of terminal complement complexes and inflammatory sequelae. This model system will facilitate the analysis of molecular and cellular aspects of AMD pathogenesis, and the testing of new therapeutic agents for its treatment.
    Proceedings of the National Academy of Sciences 11/2011; 108(45):18277-82. · 9.81 Impact Factor
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    ABSTRACT: Vitamin D has been shown to have anti-angiogenic properties and to play a protective role in several types of cancer, including breast, prostate and cutaneous melanoma. Similarly, vitamin D levels have been shown to be protective for risk of a number of conditions, including cardiovascular disease and chronic kidney disease, as well as numerous autoimmune disorders such as multiple sclerosis, inflammatory bowel diseases and type 1 diabetes mellitus. A study performed by Parekh et al. was the first to suggest a role for vitamin D in age-related macular degeneration (AMD) and showed a correlation between reduced serum vitamin D levels and risk for early AMD. Based on this study and the protective role of vitamin D in diseases with similar pathophysiology to AMD, we examined the role of vitamin D in a family-based cohort of 481 sibling pairs. Using extremely phenotypically discordant sibling pairs, initially we evaluated the association of neovascular AMD and vitamin D/sunlight-related epidemiological factors. After controlling for established AMD risk factors, including polymorphisms of the genes encoding complement factor H (CFH) and age-related maculopathy susceptibility 2/HtrA serine peptidase (ARMS2/HTRA1), and smoking history, we found that ultraviolet irradiance was protective for the development of neovascular AMD (p = 0.001). Although evaluation of serum vitamin D levels (25-hydroxyvitamin D [25(OH)D]) was higher in unaffected individuals than in their affected siblings, this finding did not reach statistical significance. Based on the relationship between ultraviolet irradiance and vitamin D production, we employed a candidate gene approach for evaluating common variation in key vitamin D pathway genes (the genes encoding the vitamin D receptor [VDR]; cytochrome P450, family 27, subfamily B, polypeptide 1 [CYP27B1]; cytochrome P450, family 24, subfamily A, polypeptide 1 [CYP24A1]; and CYP27A1) in this same family-based cohort. Initial findings were then validated and replicated in the extended family cohort, an unrelated case-control cohort from central Greece and a prospective nested case-control population from the Nurse's Health Study and Health Professionals Follow-Up Studies, which included patients with all subtypes of AMD for a total of 2,528 individuals. Single point variants in CYP24A1 (the gene encoding the catabolising enzyme of the vitamin D pathway) were demonstrated to influence AMD risk after controlling for smoking history, sex and age in all populations, both separately and, more importantly, in a meta-analysis. This is the first report demonstrating a genetic association between vitamin D metabolism and AMD risk. These findings were also supplemented with expression data from human donor eyes and human retinal cell lines. These data not only extend previous biological studies in the AMD field, but further emphasise common antecedents between several disorders with an inflammatory/immunogenic component such as cardiovascular disease, cancer and AMD.
    Human genomics 10/2011; 5(6):538-68.
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    ABSTRACT: ROBO1 is a strong candidate gene for age-related macular degeneration (AMD) based upon its location under a linkage peak on chromosome 3p12, its expression pattern, and its purported function in a pathway that includes RORA, a gene previously associated with risk for neovascular AMD. Previously, we observed that expression of ROBO1 and RORA is down-regulated among wet AMD cases, as compared to their unaffected siblings. Thus, we hypothesized that contribution of association signals in ROBO1, and interaction between these two genes may be important for both wet and dry AMD. We evaluated association of 19 single nucleotide polymorphisms (SNPs) in ROBO1 with wet and dry stages of AMD in a sibling cohort and a Greek case-control cohort containing 491 wet AMD cases, 174 dry AMD cases and 411 controls. Association signals and interaction results were replicated in an independent prospective cohort (1070 controls, 164 wet AMD cases, 293 dry AMD cases). The most significantly associated ROBO1 SNPs were rs1387665 under an additive model (meta P = 0.028) for wet AMD and rs9309833 under a recessive model (meta P = 6 × 10(-4)) for dry AMD. Further analyses revealed interaction between ROBO1 rs9309833 and RORA rs8034864 for both wet and dry AMD (interaction P<0.05). These studies were further supported by whole transcriptome expression profile studies from 66 human donor eyes and chromatin immunoprecipitation assays from mouse retinas. These findings suggest that distinct ROBO1 variants may influence the risk of wet and dry AMD, and the effects of ROBO1 on AMD risk may be modulated by RORA variants.
    PLoS ONE 01/2011; 6(10):e25775. · 3.53 Impact Factor
  • Journal of Vision - J VISION. 01/2010; 8(17):47-47.
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    ABSTRACT: During the past ten years, dramatic advances have been made in unraveling the biological bases of age-related macular degeneration (AMD), the most common cause of irreversible blindness in western populations. In that timeframe, two distinct lines of evidence emerged which implicated chronic local inflammation and activation of the complement cascade in AMD pathogenesis. First, a number of complement system proteins, complement activators, and complement regulatory proteins were identified as molecular constituents of drusen, the hallmark extracellular deposits associated with early AMD. Subsequently, genetic studies revealed highly significant statistical associations between AMD and variants of several complement pathway-associated genes including: Complement factor H (CFH), complement factor H-related 1 and 3 (CFHR1 and CFHR3), complement factor B (CFB), complement component 2 (C2), and complement component 3 (C3). In this article, we revisit our original hypothesis that chronic local inflammatory and immune-mediated events at the level of Bruch's membrane play critical roles in drusen biogenesis and, by extension, in the pathobiology of AMD. Secondly, we report the results of a new screening for additional AMD-associated polymorphisms in a battery of 63 complement-related genes. Third, we identify and characterize the local complement system in the RPE-choroid complex - thus adding a new dimension of biological complexity to the role of the complement system in ocular aging and AMD. Finally, we evaluate the most salient, recent evidence that bears directly on the role of complement in AMD pathogenesis and progression. Collectively, these recent findings strongly re-affirm the importance of the complement system in AMD. They lay the groundwork for further studies that may lead to the identification of a transcriptional disease signature of AMD, and hasten the development of new therapeutic approaches that will restore the complement-modulating activity that appears to be compromised in genetically susceptible individuals.
    Progress in Retinal and Eye Research 12/2009; 29(2):95-112. · 9.44 Impact Factor
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    ABSTRACT: The animal egg is a unique quiescent cell, prepackaged with maternal mRNAs and proteins that have functions in early development. Rapid, transient signaling at fertilization alters egg physiology, resulting in Ca(2+) release from the endoplasmic reticulum (ER) and cytoplasmic alkalinization. These events trigger the zygote developmental program through initiation of DNA synthesis and entry into mitosis. Post-translational modifications of maternal proteins are responsible for the spatio-temporal regulation that orchestrates egg activation. We used functional proteomics to identify the candidate maternal proteins involved in egg activation and early development. As the first step of this analysis, we present the data on the baseline maternal proteome, in particular, on proteins exhibiting changes in abundance and in phosphorylation state upon egg activation. We identify 94 proteins that were stable, reproducibly displayed a shift in isoelectric point, or changed in relative abundance at specific times after activation. The identities of these proteins were determined by quadrupole time-of-flight tandem mass spectrometry. The set of the most dynamic proteins appear to be enriched in intermediary metabolism proteins, cytoskeletal proteins, gamete associated proteins and proteins that have Ca(2+) mediated activities.
    Developmental Biology 02/2008; 313(2):630-47. · 3.87 Impact Factor
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    ABSTRACT: The discoveries of gene variants associated with macular diseases have provided valuable insight into their molecular mechanisms, but they have not clarified why the macula is particularly vulnerable to degenerative disease. Its predisposition may be attributable to specialized structural features and/or functional properties of the underlying macular RPE/choroid. To examine the molecular basis for the macula's disease susceptibility, we compared the gene expression profile of the human RPE/choroid in the macula with the profile in the extramacular region using DNA microarrays. Seventy-five candidate genes with differences in macular:extramacular expression levels were identified by microarray analysis, of which 29 were selected for further analysis. Quantitative PCR confirmed that 21 showed statistically significant differences in expression. Five genes were expressed at higher levels in the macula. Two showed significant changes in the macular:extramacular expression ratio; another two exhibited changes in absolute expression level, as a function of age or AMD. Several of the differentially expressed genes have potential relevance to AMD pathobiology. One is an RPE cell growth factor (TFPI2), five are extracellular matrix components (DCN, MYOC, OGN, SMOC2, TFPI2), and six are related to inflammation (CCL19, CCL26, CXCL14, SLIT2) and/or angiogenesis (CXCL14, SLIT2, TFPI2, WFDC1). The identification of regional differences in gene expression in the RPE/choroid is a first step in clarifying the macula's propensity for degeneration. These findings lay the groundwork for further studies into the roles of the corresponding gene products in the normal, aged, and diseased macula.
    Experimental Eye Research 10/2007; 85(3):366-80. · 3.03 Impact Factor
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    ABSTRACT: Polymorphisms in the complement factor H gene (CFH) are associated with a significantly increased risk for, or protection against, the development of age-related macular degeneration (AMD). The most documented risk-conferring single-nucleotide polymorphism results in a tyrosine-to-histidine substitution at position 402 (Y402H) of the CFH protein. In this work, we examined the ocular distributions and relative abundance of CFH, several CFH-binding proteins, and abundant serum proteins in the retinal pigmented epithelium (RPE), Bruch's membrane, and choroid (RPE-choroid) in CFH homozygotes possessing either the "at-risk" 402HH or "normal" 402YY variants. Although CFH immunoreactivity is high in the choroid and in drusen, no differences in CFH-labeling patterns between genotypes are apparent. In contrast, at-risk individuals have significantly higher levels of the CFH-binding protein, C-reactive protein (CRP), in the choroidal stroma. Immunoblots confirm that at-risk individuals have approximately 2.5-fold higher levels of CRP in the RPE-choroid; no significant differences in the levels of CFH or other serum proteins are detected. Similarly, we find no differences in CFH transcription levels in the RPE-choroid nor evidence for local ocular CRP transcription. Increased levels of CRP in the choroid may reflect a state of chronic inflammation that is a by-product of attenuated CFH complement-inhibitory activity in those who possess the CFH at-risk allele. Because the CRP-binding site in CFH lies within the domain containing the Y402H polymorphism, it is also possible that the AMD risk-conferring allele alters the binding properties of CFH, thereby leading to choroidal CRP deposition, contributing to AMD pathogenesis.
    Proceedings of the National Academy of Sciences 12/2006; 103(46):17456-61. · 9.81 Impact Factor
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    ABSTRACT: Variants in the complement factor H gene (CFH) are associated with age-related macular degeneration (AMD). CFH and five CFH-related genes (CFHR1-5) lie within the regulators of complement activation (RCA) locus on chromosome 1q32. Aims and Methods. In this study, the structural and evolutionary relationships between these genes and AMD was refined using a combined genetic, molecular and immunohistochemical approach. We identify and characterize a large, common deletion that encompasses both the CFHR1 and CFHR3 genes. CFHR1, an abundant serum protein, is absent in subjects homozygous for the deletion. Genotyping analyses of AMD cases and controls from two cohorts demonstrates that deletion homozygotes comprise 1.1% of cases and 5.7% of the controls (chi-square=32.8; P= 1.6 E-09). CFHR1 and CFHR3 transcripts are abundant in liver, but undetectable in the ocular retinal pigmented epithelium/choroid complex. AMD-associated CFH/CFHR1/CFHR3 haplotypes are widespread in human populations. The absence of CFHR1 and/or CFHR3 may account for the protective effects conferred by some CFH haplotypes. Moreover, the high frequencies of the 402H allele and the delCFHR1/CFHR3 alleles in African populations suggest an ancient origin for these alleles. The considerable diversity accumulated at this locus may be due to selection, which is consistent with an important role for the CFHR genes in innate immunity.
    Annals of Medicine 02/2006; 38(8):592-604. · 4.73 Impact Factor
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    ABSTRACT: Growth associated protein 43 (GAP 43) is involved in synapse formation and it is expressed in the retina in a very specific pattern. Although GAP 43 is downregulated at the time of synapse formation, it can be re-expressed following injury such as axotomy or ischemia. Because of this we sought to characterize the expression of GAP 43 after retinal detachment (RD). Immunoblot, immunocytochemical and quantitative polymerase chain reaction (QPCR) techniques were used to assess the level of GAP 43 expression after experimental RD. GAP 43 was localized to three sublaminae of the inner plexiform layer of the normal retina. GAP 43 became upregulated in a subset of retinal ganglion cells following at least 7 days of RD. By immunoblot GAP 43 could be detected by 3 days. QPCR shows the upregulation of GAP 43 message by 6hr of detachment. To further characterize changes in ganglion cells, we used an antibody to neurofilament 70 and 200kDa (NF) proteins. Anti-NF labels horizontal cells, ganglion cell dendrites in the inner plexiform layer, and ganglion cell axons (fasicles) in the normal retina. Following detachment it is upregulated in horizontal cells and ganglion cells. When detached retina was double labelled with anti-GAP 43 and anti-NF, some cells were labelled with both markers, while others labelled with only one. We have previously shown that second order neurons respond to detachment; here we show that third order neurons are responding as well. Cellular remodelling of this type in response to detachment may explain the slow recovery of vision that often occurs after reattachment, or those changes that are often assumed to be permanent.
    Experimental Eye Research 04/2003; 76(3):333-42. · 3.03 Impact Factor
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    ABSTRACT: Age-related macular degeneration (AMD) is a leading cause of irreversible vision loss in older individuals worldwide. The disease is characterized by abnormal extracellular deposits, known as drusen, that accumulate along the basal surface of the retinal pigmented epithelium. Although drusen deposition is common in older individuals, large numbers of drusen and/or extensive areas of confluent drusen represent a significant risk factor for AMD. Widespread drusen deposition is associated with retinal pigmented epithelial cell dysfunction and degeneration of the photoreceptor cells of the neural retina. Recent studies have shown that drusen contain a variety of immunomodulatory molecules, suggesting that the process of drusen formation involves local inflammatory events, including activation of the complement cascade. Similar observations in Alzheimer's disease (AD) have lead to the hypothesis that chronic localized inflammation is an important element of AD pathogenesis, with significant neurodegenerative consequences. Accordingly, the amyloid beta (A beta) peptide, a major constituent of neuritic plaques in AD, has been implicated as a primary activator of complement in AD. Here we show that A beta is associated with a substructural vesicular component within drusen. A beta colocalizes with activated complement components in these "amyloid vesicles," thereby identifying them as potential primary sites of complement activation. Thus, A beta deposition could be an important component of the local inflammatory events that contribute to atrophy of the retinal pigmented epithelium, drusen biogenesis, and the pathogenesis of AMD.
    Proceedings of the National Academy of Sciences 10/2002; 99(18):11830-5. · 9.81 Impact Factor
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    ABSTRACT: Drusen are abnormal extracellular matrix deposits characteristic of age-related macular degeneration (AMD), a leading cause of blindness in the aging human population. The mechanisms underlying drusen formation are not well characterized. The purpose of this study was to examine the expression of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) in drusen, and in the surrounding cells and tissue. To assess the extent of MMP and TIMP expression by retinal pigment epithelial (RPE) cells, cDNA arrays were screened with probes generated from cultured human RPE cells. The distribution of MMP-1, -2 and -3 and TIMP-1, -2, -3 and -4 was determined using immunohistochemistry in human RPE choroid from donor eyes with and without a clinical history of AMD. Gelatinase activity was assessed in unfixed frozen sections using in situ zymography. In cultured RPE cells, expression of 10 MMP and all four known TIMP mRNAs was detected. MMP immunoreactivity was widespread in the RPE choroid, but was absent from the interior of drusen. TIMP-3, but not other TIMPs, was detected in the drusen interior. Likewise, metal ion dependent gelatinase activity could be detected in RPE choroid, but not in drusen. These results show that, while metalloproteinase activity is widespread throughout the RPE choroid, drusen are cold spots for proteolysis. The data lead to the speculation that high TIMP-3 concentrations within drusen could inhibit MMPs and as a result slow the proteolytic degradation of these deposits.
    Experimental Eye Research 02/2002; 74(1):141-54. · 3.03 Impact Factor
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    ABSTRACT: We have used X-ray fiber diffraction to probe the structure of fibers of tau and tau fragments. Fibers of fragments from the microtubule binding domain had a cross beta-structure that closely resembles that reported both for neurofibrillary tangles found in Alzheimer's disease brain and for fibrous lesions from other protein folding diseases. In contrast, fibers of full-length tau had a different, more complex structure. Despite major differences at the molecular level, all fiber types exhibited very similar morphology by electron microscopy. These results have a number of implications for understanding the etiology of Alzheimer's and other tauopathic diseases. The morphology of the peptide fibers suggests that the region in tau corresponding to the peptides plays a critical role in the nucleation of fiber assembly. The dramatically different structure of the full length tau fibers suggests that some region in tau has enough inherent structure to interfere with the formation of cross beta-fibers. Additionally, the similar appearance by electron microscopy of fibrils with varying molecular structure suggests that different molecular arrangements may exist in other samples of fibers formed from tau.
    Protein Science 01/2001; 9(12):2427-35. · 2.74 Impact Factor
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    S Ozaki, M J Radeke, D H Anderson
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    ABSTRACT: To determine the mechanism by which basic fibroblast growth factor (bFGF) exerts its neuroprotective effects on degenerating or injured photoreceptors. Confocal immunofluorescence microscopy was used to identify sites of bFGF and FGF receptor 1 (FGFR1) expression after focal injury or experimental retinal detachment in adult rats. FGFR1 expression was analyzed immunohistochemically and at the transcription level in single photoreceptor cells, after reverse transcription (RT), using the polymerase chain reaction (PCR). Real time quantitative RT-PCR was used to measure changes in FGFR1 mRNA levels in the retina in response to injury or detachment. Confocal immunofluorescence observations showed that FGFR1 immunoreactivity in the rat retina is concentrated primarily in the perinuclear cytoplasm of photoreceptor cell bodies. Reverse transcription of total RNA derived from dissociated rat photoreceptor cells, followed by amplification of FGFR1 cDNA using the PCR, verified the presence of FGFR1 transcripts in normal rat photoreceptor cells; in contrast, no evidence of bFGF transcription was detected. Collectively, these results provide compelling evidence for FGFR1 gene expression by rat photoreceptors in situ. Within hours after experimental retinal detachment or focal injury, there is a twofold increase in FGFR1 immunoreactivity in the outer nuclear layer that persists for at least 7 days; a similar increase in bFGF immunoreactivity in the interphotoreceptor matrix is also apparent. This increase in FGFR1 protein levels after detachment and injury also was confirmed by western blot analysis. Real time quantitative RT-PCR analyses revealed that a rapid upregulation of FGFR1 mRNA occurred within 12 hours after retinal injury/detachment, but then declined to near baseline levels by 24 hours. This body of evidence strongly suggests that the photoreceptor rescue effect elicited by retinal injury as well as by injection of exogenous bFGF is mediated, at least in part, by upregulation of the FGFR1 by the photoreceptor cells.
    Investigative Ophthalmology &amp Visual Science 03/2000; 41(2):568-79. · 3.44 Impact Factor
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    ABSTRACT: To investigate the role of neurotrophins in the initial formation of striatal patch versus matrix, the spatial and temporal expression of trkB receptors was examined using immunohistochemistry. Polyclonal antibodies, against the C-terminus or the tyrosine kinase domain, revealed trkB-immunoreactive cells and fibers localized to patches beginning on embryonic day 19 in the rat, which co-localized with patchy dopamine fibers, substance P-immunoreactive neurons and glutamate receptors. Patchy striatal trkB expression was maintained after lesioning the nigrostriatal dopamine system. The patchy trkB distribution persisted through postnatal day 14, then became more homogeneous at the same time that nigrostriatal afferents become homogeneous. Later in development, trkB immunoreactivity was most intense in a subpopulation of large striatal cells that were similar in size and frequency to those immunoreactive for choline acetyltransferase. The spatiotemporal expression of trkB receptor in phenotypically distinct striatal patches, as well as evidence that neurotrophins regulate expression of neuronal phenotypic markers during development, may indicate a convergence of neurotrophins and afferent innervation on to future patch cells that may regulate the establishment of striatal compartmentalization.
    Neuroscience 04/1999; 89(2):505-13. · 3.12 Impact Factor
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    ABSTRACT: The interaction between tubulin subunits and microtubule-associated proteins (MAPs) such as tau is fundamental for microtubule structure and function. Previous work has suggested that the "microtubule binding domain" of tau (composed of three or four imperfect 18-amino acid repeats, separated by 13- or 14-amino acid inter-repeat regions) can bind to the C-terminal ends of both alpha and beta tubulin monomers. Here, using covalent cross-linking strategies, we demonstrate that there are two distinct tau cross-linking sites (designated as "C-terminal" and "internal") on each alpha and beta tubulin monomer. The C-terminal tau cross-linking site is located within the 12 C-terminal amino acids of both alpha and beta tubulin, while the internal tau cross-linking site is located within the C-terminal one-third of alpha and beta tubulin but not within the last 12 amino acids. In addition, we show that tau cross-links to the C-terminal site via its repeat 1 and/or the R1-R2 inter-repeat. The cross-linking of tau to the internal site is mediated by some subset of its other repeat units. Integrating these and earlier data with the 3.7 A resolution model of the alphabeta tubulin dimer recently presented by E. Nogales et al. [(1998), Nature 391, 199-203], we propose a new model for the tau-microtubule interaction.
    Biochemistry 01/1999; 37(51):17692-703. · 3.38 Impact Factor
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    ABSTRACT: The neurotrophins brain-derived neurotrophic factor (BDNF) and NT-4/5 exert their trophic effects on the nervous system via signaling through trkB receptors. These receptors occur as splice variants of the trkB gene that encodes a full-length receptor containing the signal transducing tyrosine kinase domain as well as truncated forms lacking this domain. Because the importance of the trkB isoforms for development and maturation of the nervous system is unknown, we have examined the expression of trkB receptor isoforms during development of the rat forebrain using 1) a sensitive ribonuclease protection assay to distinguish full-length and truncated trkB transcripts, 2) western blot analysis to characterize developmental changes in trkB proteins, and 3) immunohistochemistry to determine the cellular localization of trkB receptors. In the rat forebrain, adult mRNA levels for full-length trkB are reached by birth, whereas truncated trkB message does not peak until postnatal days 10–15. Western blot analysis indicates that full-length trkB protein is the major form during early development, whereas truncated trkB protein predominates in all forebrain regions of late postnatal and adult rats. These data also suggest that the glycosylation state of these receptors changes during postnatal maturation. TrkB immunoreactivity is present predominately within neurons, where it is localized to axons, cell soma, and dendrites. Strong dendritic immunostaining is particularly evident in certain neuronal populations, such as pyramidal neurons in the hippocampus and in layer V of the neocortex. The dendritic localization of trkB receptors supports the hypothesis that dendrites, as well as axons, are important sites for neurotrophin actions in the central nervous system. © 1996 Wiley-Liss, Inc.
    The Journal of Comparative Neurology 12/1998; 374(1):21 - 40. · 3.66 Impact Factor
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    ABSTRACT: Nerve growth factor (NGF), which has long been considered to be a trophic factor for peripheral sensory and sympathetic neurons, has been found recently to influence cholinergic neurons in the basal forebrain and neostriatum. In the present study, we provide evidence that brainstem neurons in the perihypoglossal area that relay information from the inner ear and vestibular apparatus to the cerebellum and tectum are responsive to NGF. These neurons, which are located in the nucleus prepositus hypoglossi (NPH), spinal vestibular nucleus, cochlear complex, and gigantocellular and paragigantocellular nuclei of the reticular formation, express functional receptors for NGF and up-regulate the expression of trkA receptors after injection of NGF into targets. In addition, the developmental up-regulation of NGF in the cerebellum coincides with the differentiation of the perihypoglossal nuclei. These results suggest that neurons representing the principal brain relays for auditory and vestibular pathways and perihypoglossal neurons involved in gaze coordination are a novel group of central neurons (besides cholinergic neurons in the basal forebrain and neostriatum) that respond to NGF. J. Comp. Neurol. 383:123-134, 1997. © 1997 Wiley-Liss, Inc.
    The Journal of Comparative Neurology 12/1998; 383(2):123 - 134. · 3.66 Impact Factor

Publication Stats

3k Citations
257.61 Total Impact Points

Institutions

  • 2012
    • CSU Mentor
      Long Beach, California, United States
  • 1991–2012
    • University of California, Santa Barbara
      • • Department of Molecular, Cellular, and Developmental Biology
      • • Neuroscience Research Institute
      Santa Barbara, California, United States
  • 1997
    • Molecular and Cellular Biology Program
      Seattle, Washington, United States
  • 1990
    • Stanford Medicine
      • Department of Neurobiology
      Stanford, California, United States
  • 1987
    • Stanford University
      • Department of Neurobiology
      Stanford, CA, United States