Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Colocalization analysis”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 91 records · Page 5Linked to original sources
PubMed1989

A peptide histidine isoleucine/peptide histidine methionine-like peptide in the rabbit retina: colocalization with vasoactive intestinal peptide, synaptic relationships and activation of adenylate cyclase activity.

Antisera against peptide histidine isoleucine and peptide histidine methionine were found to label a subpopulation of amacrine and displaced amacrine cells in the rabbit retina with processes ramifying in sublaminas 1, 3 and 5 of the inner plexiform layer. Preadsorption controls demonstrated that this immunoreactivity was specific for a peptide histidine isoleucine- or peptide histidine methionine-like (peptide histidine isoleucine/peptide histidine methionine-like) peptide, and was not caused by cross-reactivity of the peptide histidine isoleucine or peptide histidine methionine antibodies with vasoactive intestinal peptide vasoactive intestinal peptide. In double-label studies, vasoactive intestinal peptide and peptide histidine isoleucine/peptide histidine methionine-like immunoreactivity were colocalized in the same population of retinal neurons. Electron microscopic analysis revealed that the peptide histidine isoleucine/peptide histidine methionine-labelled cells interacted with processes of bipolar cells, amacrine cells and ganglion cells. Peptide histidine methionine and peptide histidine isoleucine were slightly less potent than vasoactive intestinal peptide in stimulating adenylate cyclase activity in the rabbit retina, while the related peptides secretin, glucagon, and the C-terminal vasoactive intestinal peptide fragment, vasoactive intestinal peptide (10-28), showed little or no stimulatory activity. Stimulation of adenylate cyclase by high concentrations of vasoactive intestinal peptide and peptide histidine methionine were non-additive. These results suggest that a peptide histidine isoleucine/peptide histidine methionine-like peptide may function as a neuroactive peptide in the mammalian retina, and that this peptide appears to be cosynthesized and colocalized with vasoactive intestinal peptide and to mimic the activity of vasoactive intestinal peptide through interaction with vasoactive intestinal peptide receptor-adenylate cyclase complexes.

Adenylyl Cyclases↗
PubMed1991

Colocalization of neurotransmitters analyzed by in situ hybridization.

In situ hybridization and Northern blot analysis has been used to analyse in some detail the localization and regulation of the messenger molecules adrenaline, noradrenaline and neuropeptide tyrosine (NPY) within cells of the sympathetic nervous system and the adrenal medulla. In the rat adrenal gland, a novel NPY containing population of ganglion cells was found. Synthetic oligonucleotide probes complementary to mRNA coding for the catecholamine synthesizing enzymes phenylethanolamine N-methyltransferase (PNMT), tyrosine hydroxylase (TH) and NPY were used to analyse the regulation of these genes following administration of the catecholamine depleting drug reserpine. Twenty-four hours after a single dose of reserpine, a differential regulation of PNMT, TH and NPY was found. Thus, a dramatic decrease in PNMT mRNA was observed in the adrenal medulla. In contrast, mRNA for both TH and NPY exhibited an increase. Different regulatory mechanisms may thus operate for these three compounds coexisting in chromaffin cells of the adrenal medulla. The regulation of enzymes and peptides was also studied in human sympathetic ganglia. After brief electrical preganglionic stimulation of thoracic ganglia in humans, in situ hybridization was performed with synthetic oligonucleotide probes complementary to TH, dopamine beta-hydroxylase (DBH) and NPY mRNA respectively. A several fold increase in all three mRNAs was found in the principal ganglion cells. The results point to a very rapid regulation of genes involved in signal transmission in the sympathetic nervous system of humans. The results also suggest a novel way to define neuronal projections by visualizing increases in mRNA levels following electrical stimulation.

Adrenal Glands↗
PubMed1991

Amiloride-sensitive sodium channel is linked to the cytoskeleton in renal epithelial cells.

Amiloride-sensitive sodium channels are localized to the microvillar domain of apical membranes in sodium-transporting renal epithelial cells. To elucidate the elements that maintain sodium channel distribution at the apical membrane, we searched for specific proteins associating with the channel. Triton X-100 extraction of A6 epithelial cells reveals that sodium channels are associated with detergent-insoluble and assembled cytoskeleton. Indirect immunofluorescence and confocal microscopy show that sodium channels are segregated to the apical microvillar membrane and colocalize with ankyrin, fodrin, and actin. We document by immunoblot analysis that ankyrin and fodrin remain associated with sodium channels after isolation and purification from bovine renal papillae. 125I-labeled ankyrine can be precipitated by anti-sodium-channel antibodies only in the presence of purified bovine sodium-channel complex. Direct binding of 125I-labeled ankyrin shows ankyrin binds to the 150-kDa subunit of the channel. Fluorescence photobleach lateral-diffusion measurements indicate sodium channels are severely restricted in their lateral mobility. We conclude that ankyrin links the amiloride-sensitive sodium channel to the underlying cytoskeleton and this association may sequester sodium channels at apical microvilli and maintain their polarized distribution in renal epithelial cells.

Amiloride↗
PubMed1992

Distinct sites of production and deposition of the putative cell death marker clusterin in the human thymus.

Clusterin is a multifunctional protein endowed with cell-aggregating, complement-inhibitory, and lipid-binding properties. Since several studies have demonstrated highly increased clusterin gene expression in epithelial and nervous tissues regressing as a consequence of tissue involution and apoptotic cell death, clusterin is also considered as a specific marker of dying cells. To determine whether clusterin expression is also upregulated during thymocyte death occurring during the negative selection process we analyzed the cellular distribution of clusterin mRNA and protein by in situ hybridization and immunocytochemistry in the human thymus. We observed that the expression of clusterin mRNA was confined to cells present in the thymic medulla, concentrated mainly around Hassal's bodies. Immunostaining of adjacent sections with antikeratin Ab revealed that cells containing clusterin mRNA were predominantly epithelial. By contrast no clusterin mRNA was found in thymocytes by in situ hybridization and Northern blot analysis of total RNA from purified thymocyte populations. Clusterin protein colocalized with the membrane attack complex of complement and vitronectin in the center of the largest Hassal's bodies, but was not detectable by immunocytochemistry in or at the surface of epithelial cells. Our results demonstrate that clusterin gene expression does not take place in apoptotic thymocytes, and therefore that clusterin synthesis by the dying cell is probably not a prerequisite to its death. However, synthesis of clusterin by medullary epithelial cells may be related to their terminal differentiation, and, furthermore, its presence in Hassal's bodies raises the possibility that the secreted protein is involved in the disposal of cell debris resulting from thymocyte apoptosis.

Apoptosis↗
PubMed1989

Expression of rat neurofilament proteins NF-L and NF-M in transfected non-neuronal cells.

Two cDNA clones fully encoding the rat neurofilament proteins NF-L and NF-M were subcloned into eukaryotic expression vectors behind the strong constitutive viral promoters from SV40 and Rous sarcoma viruses. Transient transfection of L tk- and Cos cell lines with these expression constructs resulted in cells expressing the neurofilament proteins in an intermediate filament-type pattern. Additionally, a putative juxtanuclear organizing center or region was observed in the transfected cells, most noticeable shortly after the transfection procedure. Stable transfections were performed on mouse L tk- and Swiss 3T6 cells using NF-L and NF-M constructs bearing an SV40 early promoter driven neomycin selectable marker. Although G418-resistant clones were recovered with both the NF-L and the NF-M constructs, only clones expressing immunofluorescently stainable amounts of NF-M were detected and established. Immunoelectron microscopic analysis revealed NF-M and vimentin proteins to be colocalized on the same intermediate filaments.

Amino Acid Sequence↗
PubMed1992

The small GTP-binding protein rab6p is distributed from medial Golgi to the trans-Golgi network as determined by a confocal microscopic approach.

A key role in the regulation of membrane traffic is played by the rab proteins, members of a family of ras-related small GTP-binding proteins. This family comprises at least 25 identified members, the intracellular localization of only a few of which has been investigated. rab6p has been shown to be distributed along the exocytic pathway in association with the medial and trans regions of the Golgi apparatus. A confocal laser scanning microscopic (CLSM) approach coupled with image analysis was used to compare the localization of rab6p with selected reference Golgi markers by double immunofluorescence on culture cell lines. CLSM analysis shows that, under a set of well-defined conditions, one can investigate the possible colocalization of known markers of Golgi compartments and orientate a couple of labeled Golgi antigens with regard to the polarity of the Golgi apparatus. Thus, having validated the CLSM analysis, the localization of rab6p was studied and compared with some of these markers and the VSV-G protein in VSV (vesicular stomatitis virus)-infected cells blocked at 20 degrees C. rab6p is shown to be associated in all the cell lines used with the last cisternae of the Golgi apparatus and particularly with the trans-Golgi network (TGN), the site of protein sorting at the exit of the Golgi apparatus. These results were supported by an electron microscopic study using double-immunolabeled cryosections: rab6p was found in some flat cisternae of the Golgi stack and colocalized with the VSV-G protein in the TGN. Our results show that the small GTP-binding protein rab6p is distributed from medial Golgi to TGN along the exocytic pathway.

Biological Transport↗
PubMed2026

Integrative cross-tissue transcriptome-wide association and metabolomic analysis reveals novel genetic risk loci for aortic aneurysm.

BACKGROUND: Aortic aneurysm (AA) is a life-threatening cardiovascular condition with a strong genetic component, however, its molecular mechanisms remain poorly understood. Although genome-wide association studies (GWAS) have identified numerous risk loci, most prior studies have investigated genetic and metabolic factors separately, leaving the causal pathways from genetic variants to disease largely unexplored. METHODS: We established an integrative framework combining cross-tissue transcriptome-wide association studies (TWAS) with metabolomic mediation analysis. First, we integrated GWAS data from FinnGen R12 with multi-tissue expression quantitative trait loci (eQTL) data from Genotype-Tissue Expression Project (GTEx) V8, then performed cross-tissue TWAS using the Unified Test for MOlecular SignaTures (UTMOST) and single-tissue validation with the Functional Summary-based Imputation (FUSION) to prioritize susceptibility genes. Second, we applied Mendelian randomization (MR), colocalization, and Fine-mapping Of CaUsal gene Sets (FOCUS) to assess causality and identify high-confidence genes. Third, we performed metabolite mediation analysis to uncover metabolic pathways linking genetic variants to disease risk. Finally, we validated key findings in mouse models of thoracic aortic aneurysm (TAA) and abdominal aortic aneurysm (AAA) using Quantitative Real-Time Reverse Transcription Polymerase Chain Reaction (RT-qPCR) and Western blotting. RESULTS: We identified multiple novel susceptibility genes for AA and its subtypes. Key genes included ADH family members (ADH1A, ADH1B, ADH4, ADH6) and ZNF827, which showed cross-subtype associations with strong colocalization evidence in vascular tissues. Metabolite mediation analysis revealed significant pathways involving N-acetylphenylalanine and methionine sulfoxide. Functional enrichment revealed distinct biological mechanisms: AA and AAA were primarily associated with metabolic pathways, whereas TAA-related genes were enriched in developmental and contractile processes. PheWAS indicated no significant off-target associations. Critically, experimental validation in mouse models confirmed significant upregulation of ZNF827 in TAA and ADH6 in AAA at both mRNA and protein levels, corroborating the genetic predictions. CONCLUSION: This integrated cross-omics analysis identifies novel genetic loci and, crucially, uncovers specific nutrient-related metabolic pathways that mediate genetic risk. These findings provide a mechanistic basis for future nutritional and metabolic intervention studies in AA and its subtypes.

MAGMA↗
PubMed1992

Distribution of protein gene product 9.5 (PGP 9.5) in the vertebrate retina: evidence that immunoreactivity is restricted to mammalian horizontal and ganglion cells.

Using light microscopic immunocytochemistry, we have studied the distribution of protein gene product 9.5 (PGP 9.5), a neuron-specific protein first extracted from human brain (Doran et al., '83:J. Neurochem. 40:1542-1547), in the vertebrate retina. Retinas were obtained from frog, chicken, rat, rabbit, cow, cat, dog, and human. No immunoreactivity was observed in frog and only a faint staining was present in chicken. In mammalian retinas, a strong positive reaction was restricted to horizontal and ganglion cells, with minor interspecies variations. Immunostaining was present throughout the cell body and the dendritic tree in horizontal cells. At the level of retinal ganglion cells, immunolabel was particularly abundant in cell bodies and axons forming the optic nerve. Only the main dendrites were stained, the remainder of the dendritic tree giving rise to a diffuse punctate reaction in the inner plexiform layer. In rats, displaced amacrine cells, which are known to contribute largely (40-50%) to the total neuronal population within the ganglion cell layer (Perry, '81: Neuroscience 6:931-944) were not immunoreactive, as demonstrated from (i) analysis of the morphology, cell size and cell density of immunoreactive neurons in wholemounts; (ii) colocalization of retrograde label and PGP 9.5 immunoreactivity in about 80% of ganglion cells after injection of peroxidase into the optic nerve; and (iii) reduction of immunoreactivity in the inner plexiform and ganglion cell layers following optic nerve transection. Western blot analysis of extracts from rabbit retinas indicated that the immunoreactive species is PGP 9.5 or a closely related molecule. Recent studies have demonstrated that PGP 9.5 is a ubiquitin carboxyl-terminal hydrolase (Wilkinson et al., '89:Science 246:670-673). The present results, therefore, suggest that differences in the ubiquitination process exist between retinal neurons.

Animals↗
PubMed2026

Tonsillar expression quantitative trait loci verify and expand genetic contributors to childhood atopic diseases.

BACKGROUND: The spectrum of causal variants, mechanisms, and immunologic gene networks that influence pediatric atopic traits is not completely understood. Human genetic variation associated with transcript abundance (expression quantitative trait loci [eQTLs]) can help to advance our understanding, yet prior work has focused on profiling immune cell populations collected from peripheral blood primarily in adult populations, leaving tissue-resident lymphocytes collected from children uncharacterized. OBJECTIVE: We sought to characterize gene expression of 4 populations of tonsil-derived immune cell types collected from pediatric patients. METHODS: We collected naive B, germinal center B, naive T, and T follicular helper cells from the discarded tonsils of 103 children across development (age range 1-19). Following genotyping and RNA sequencing of samples, we performed differential expression and eQTL analysis, then statistically linked eQTL signals to relevant atopic traits via colocalization. RESULTS: We found differentially expressed genes across cell types and identified 13,393 expression genes (eGenes) (1,793 eGenes not previously reported in similar datasets) influenced by 27,603 eQTLs (5,199 eQTLs not previously reported). We linked eQTLs to associations identified in pediatric and adult asthma and atopy traits, nominating 78 eGenes including TRAF3, ZBTB10, and JAZF1 in disease-relevant cell types. CONCLUSIONS: Our freely available resource exemplifies the importance of discovery in native tissues and across human development.

Expression quantitative trait locus↗
PubMed1991

Expression of intercellular adhesion molecule-1 (ICAM-1) in the skin of patients with systemic scleroderma.

The expression and tissue distribution of intercellular adhesion molecule-1 (ICAM-1) in skin biopsies from 12 patients with systemic (SSc) and localized (LS) scleroderma was studied and compared to the biopsies from patients with lupus erythematosus (LE) and normal individuals. In normal human skin ICAM-1 expression was restricted to the vascular endothelium, infiltrating mononuclear cells (MNC), and to few individual keratinocytes. In the inflammatory stage of SSc, however, the expression of ICAM-1 was dramatically increased at the site of MNC infiltrates and could also be detected on fibroblast-like cells lying well apart from these infiltrates in the deep dermis. In contrast, in LS ICAM-1 was expressed mainly at the sites of MNC infiltrates. In LE ICAM-1 expression was confined to the keratinocytes, endothelial cells, and mononuclear cells in the upper parts of the dermis. Analysis of serial tissue sections from patients with SSc demonstrated also colocalization of staining of ICAM-1 around blood vessels with LFA-1-positive lymphocytes. Increased expression of ICAM-1 in the dermis of patients with SSc may represent an important mechanism by which MNC become localized and retained at a site of connective tissue inflammation, leading to the activation of fibroblasts.

Adult↗
PubMed1991

Colocalization of N-CAM and N-cadherin in avian skeletal myoblasts.

The cell-cell adhesion molecules, N-CAM and N-cadherin, have been shown previously to mediate myoblast interaction during cell fusion accompanying skeletal myogenesis. To study the localization of both molecules in fusion-competent myoblasts, we used antigen-specific primary antibodies and a double-labeling preembedding immuno-electron microscopy technique. Ultrastructural observations and quantitative analysis of the results reveal that N-CAM and N-cadherin frequently colocalize in clusters on the myoblast plasma membrane. The data provide morphological evidence that the two adhesion glycoproteins cooperate in mediating myoblast interaction during myoblast fusion.

Animals↗
PubMed2025

Integration of multiple omics reveals key targets and cellular mechanisms for intervention in sarcopenia.

BACKGROUND: Sarcopenia, an age-related syndrome characterized by progressive loss of muscle mass, strength, and function, presents a significant global health burden with limited therapeutic interventions. This study integrates genomic causality, multi-tissue omics, and cellular mediation analyses to identify and prioritize mechanistically grounded therapeutic targets. METHODS: A multi-tiered analytical framework was applied, beginning with two-sample Mendelian randomization (MR) to infer causal relationships between 4907 plasma proteins (cis-pQTLs from 35,559 individuals) and sarcopenia traits in Pan-UK Biobank participants. Bayesian colocalization and transcriptomic validation in human sarcopenia muscle biopsies were employed to prioritize targets. Cellular mediation analysis quantified contributions of immune and stromal cell subtypes to protein-trait pathways using transcriptomic deconvolution. RESULTS: MR identified 1237 plasma proteins causally associated with sarcopenia traits, with six targets (HGFAC, GATM, HMOX2, F2, LMAN2L, HPGDS) validated through colocalization, transcriptomic expression, and sarcopenia-related dysregulation. Cellular mediation revealed immune mechanisms underlying HGFAC's effects, with CD4+ regulatory T cells mediating 3.49 % of its impact on sarcopenia traits. Prothrombin exhibited muscle-protective effects independent of coagulation. CONCLUSION: This study establishes a causal map linking plasma proteins to sarcopenia through immune-stromal interactions. The integration of MR, multi-omics validation, and cellular mediation prioritizes six proteins as actionable targets, supporting repurposing of thrombin inhibitors and development of immunometabolic therapies. The framework bridges genomic causality with cellular pathophysiology, advancing precision strategies for age-related muscle decline.

Humans↗
PubMed2026

Genetic Determinants of Pulmonary Artery Size in over 50,000 Subjects with and without COPD.

RATIONALE: Pulmonary artery (PA) enlargement is a non-invasive imaging biomarker associated with pulmonary hypertension and mortality in COPD; however, its genetic determinants remain incompletely understood. OBJECTIVES: To characterize the genetic architecture of PA size across COPD-enriched and population-based cohorts. METHODS: We performed genome-wide association analyses of PA diameter using whole-genome sequencing in COPDGene (n=9,418) and ECLIPSE (n=1,859), and imputed-genotype data from the UK Biobank (n=37,073). We replicated lead variants in the Framingham Heart Study (FHS; n=3,289), incorporated all four studies into a joint meta-analysis, and identified independent signals through conditional analyses. Candidate effector genes were prioritized using coding variant annotation, colocalization, and integrative regulatory evidence. MEASUREMENTS AND MAIN RESULTS: We identified 44 independent genome-wide significant PA diameter signals within 39 loci, including 8 variants replicated in FHS, novel associations near FRMD4B, SLC20A2, BORCS7-ASMT, and KCNRG, and 5 signals in conditional analysis including multiple signals at ANO1. Genetic effects were concordant across imaging modalities and cohorts of differing COPD burden. Effector-gene prioritization nominated ABCC8, PDGFD, HMCN1, CCNE1, and TBX20, implicating pathways in vascular remodeling, developmental regulation, smooth muscle and endothelial function, ion-channel signaling, and extracellular matrix organization. Colocalization with pulse pressure GWAS demonstrated substantial shared causal variation between pulmonary and systemic vascular biology. CONCLUSIONS: In this largest genetic study of pulmonary vascular imaging to date, PA diameter exhibits a polygenic architecture consistent across imaging modalities and cohorts of differing COPD burden. The prioritized effector genes bridge rare-variant pulmonary hypertension biology with common-variant systemic vascular biology.

Pulmonary artery diameter↗
PubMed2026

Deciphering the Genetic Underpinnings of Liver Cirrhosis-Heart Failure Comorbidity Through Multi-Omics: CRIM1 as a Key Endothelial Mediator.

The co-occurrence of liver cirrhosis (LC) and heart failure (HF) poses considerable clinical challenges, yet the cellular and molecular determinants of this comorbidity remain poorly characterized. To address this, we developed an integrative multi-omics pipeline encompassing GWAS meta-analysis, gsMap-based spatial transcriptomic projection, GeneEnrich functional annotation, single-cell atlas construction, seismicGWAS and ECLIPSER cell-type scoring, eCAVIAR and fastenloc colocalization, hdWGCNA network inference, scTenifoldKnk in silico gene perturbation, and GCTA-COJO fine-mapping. Quality-controlled meta-analysis yielded 12,347,758 and 9,256,862 variant-level associations for LC and HF, respectively. Spatial projection confirmed preferential enrichment of disease signals within embryonic hepatic and cardiac compartments. Pathway analyses disclosed that LC-linked loci were concentrated in lipid metabolic programs, whereas HF-linked loci implicated mitochondrial bioenergetics and lysosomal degradation. At the cellular level, endothelial cells emerged as the dominant HF-associated population. Convergent evidence from five orthogonal algorithms pinpointed CRIM1 as the sole robustly supported shared gene, selectively enriched in HF endothelial cells; virtual perturbation further identified LCP1 and PTPRC as downstream regulatory nodes. Fine-mapping of the chromosome 2 locus harboring rs12476437 revealed multiple statistically independent signals in the vicinity of CRIM1. Collectively, these findings computationally prioritize the endothelial-CRIM1 axis as a previously unappreciated candidate mechanistic bridge between LC and HF requiring experimental validation.

Humans↗
PubMed2026

The causal relationships and potential pathways between birth weight and cardiovascular diseases: A human genomics study.

The causal relationships and potential pathways between birth weight (BW) and various cardiovascular diseases (CVDs) remain unclear, particularly when discriminating maternal and fetal contributions of BW to CVDs. Leveraging the genome-wide association studies (GWASs) of BW (N = 321,223) and a range of CVDs (ncases = 43,676-181,522), we performed a 2-sample Mendelian randomization (MR) analysis to estimate the causal effect of BW, fetal-specific BW, and maternal-specific BW on coronary artery disease (CAD), myocardial infarction (MI), heart failure (HF), atrial fibrillation (AF), and stroke. Furthermore, we applied a stepwise MR analysis approach to assess the potential involvement of childhood body mass index (CBMI) and age at menarche (AAM) in the causal pathways from BW to CVDs, while considering adult BMI. Finally, we performed colocalization analyses to justify the different biological mechanisms of maternal-specific and fetal-specific BW. The 2-sample MR analysis revealed that genetically predicted higher BW per standard deviation (SD) was associated with a decreased risk of CAD (odds ratio [OR] = 0.804, 95% confidence interval [CI]: 0.731-0.883), MI (OR = 0.720, 95% CI: 0.638-0.814), and stroke (OR = 0.900, 95% CI: 0.823-0.985), but an increased risk of AF (OR = 1.279, 95% CI: 1.160-1.410). Similar associations were observed for fetal-specific/maternal-specific BW. The stepwise MR analysis indicated that CBMI and AAM could serve as factors linking BW/fetal-specific BW and CVDs, albeit in different roles, by displaying an indirect causal effect through adult BMI. However, for maternal-specific BW, our results failed to support a causal effect on CBMI or AAM. Colocalization analyses supported the distinct biological mechanisms for maternal-specific and fetal-specific BW by showing different causal genes. The study suggested that both fetal genotype and intrauterine environmental exposure contribute to the causal associations. Additionally, AAM and CBMI may play a role in the pathways linking BW and CVDs, though the effect was only observed for fetal-specific BW.

Humans↗
PubMed1992

Scyllatoxin, a blocker of Ca(2+)-activated K+ channels: structure-function relationships and brain localization of the binding sites.

Chemical modifications of scyllatoxin (leiurustoxin I) have shown that two arginines in the sequence, Arg6 and Arg13, are essential both for binding to the Ca(2+)-activated K+ channel protein and for the functional effect of the toxin. His31 is important both for the binding activity of the toxin and for the induction of contractions on taenia coli. However, although its iodination drastically decreases the toxin activity, it does not abolish it. Chemical modification of lysine residues or of Glu27 does not significantly alter toxin binding, but it drastically decreases potency with respect to contraction of taenia coli. The same observation has been made after chemical modification of the lysine residues. The brain distribution of scyllatoxin binding sites has been analyzed by quantitative autoradiographic analysis. It indicates that apamin (a bee venom toxin) binding sites are colocalized with scyllatoxin binding sites. The results are consonant with the presence of apamin/scyllatoxin binding sites associated with Ca(2+)-activated K+ channels. High-affinity binding sites for apamin can be associated with very-high-affinity (less than 70 pM), high-affinity (approximately 100-500 pM), or moderate-affinity (greater than 800 pM) binding sites for scyllatoxin.

Amino Acid Sequence↗
PubMed1990

Two novel peripheral membrane proteins, pasin 1 and pasin 2, associated with Na+,K(+)-ATPase in various cells and tissues.

Purification of pig kidney Na+,K(+)-ATPase at low concentrations of SDS (0.5%) allowed copurification of several peripheral membrane proteins. Some of these associated proteins were identified as components of the membrane cytoskeleton. Here we describe two novel globular proteins of of Mr 77,000 (pasin 1) and Mr 73,000 (pasin 2) which copurify and coimmunoprecipitate with Na+,K(+)-ATPase and can be stripped off Na+,K(+)-ATPase microsomes by 1 M KCl. Pasin 1 and pasin 2 were detected by immunoblot analysis in various cells and tissues including erythrocytes and platelets. Immunostaining revealed colocalization of pasin 1 and Na+,K(+)-ATPase along the basolateral cell surface of epithelial cells of kidney tubules and parotid striated ducts (titers of pasin 2 antibodies were too weak for immunocytochemistry). In erythrocytes, pasin 1 and pasin 2 are minor components bound to the cytoplasmic surface of the plasma membrane. Pasin 1 showed the same electrophoretic mobility as protein 4.1b. However, both proteins have different isoelectric points (pasin 1, pI 6; protein 4.1, pI 7), different chymotryptic fragments, and are immunologically unrelated. Short pieces of sequence obtained from pasin 1 and pasin 2 were not found in any other known protein sequence. The occurrence of pasin 1 and pasin 2 in diverse cells and tissues and their association with Na+,K(+)-ATPase suggests a general role of these proteins in Na+,K(+)-ATPase function.

Amino Acid Sequence↗
PubMed1990

Analysis of adenosine immunoreactivity, uptake, and release in purified cultures of developing chick embryo retinal neurons and photoreceptors.

We have investigated the presence of endogenous adenosine and of mechanisms for adenosine uptake and release in chick embryo retinal neurons and photoreceptors grown in purified cultures in the absence of glial cells. Simultaneous autoradiographic and immunocytochemical analysis showed that endogenous adenosine and the uptake mechanism for this nucleoside colocalize in practically all the photoreceptors, but only in approximately 20% of the neurons. Approximately 25% of the neurons showed either immunocytochemical labeling or autoradiographic labeling, while greater than 50% of the neurons were unlabeled with both techniques. [3H]Adenosine uptake was saturable and could be inhibited by nitrobenzylthioinosine and dipyridamole and by pretreatment of the [3H]adenosine with adenosine deaminase. Although these observations indicate that the uptake is specific for adenosine, only 35% of accumulated radioactivity was associated with adenosine, with the remaining 65% representing inosine, hypoxanthine, and nucleotides plus uric acid. Adenosine as well as several of its metabolites were released by the cells under basal as well as K(+)-stimulated conditions. Potassium-enhanced release was blocked by 10 mM CoCl2 or in Ca2(+)-free, Mg2(+)-rich solutions. The results indicate that retinal cells that synthesize, store, and release adenosine differentiate early during embryogenesis and are therefore consistent with a hypothetical role for adenosine in retinal development.

Adenosine↗