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PubMed1982

Relative halothane accumulation in brain subcellular membranes in vitro.

The accumulation of halothane in brain homogenates was compared with halothane accumulation in brain during inhalation at anesthetic and subanesthetic levels. Anesthesia is achieved at a tissue concentration well below the halothane solubility in brain tissue. Analysis of halothane in the particulate solids of brain homogenate and in purified subcellular membranes indicates that a membrane constituent (presumably the lipids) acts as an ideal solvent in which halothane is fully miscible. Therefore, membranes offer a local microenvironment in which halothane accumulation deviates from Henry's law. Specifically, we observe that even slight increases of halothane in a saline medium result in a relatively large increase in the concentration of halothane in subcellular membranes suspended in the medium, eventually leading to solvation of the membrane in halothane. This observation offers a ready explanation for the high degree of positive correlation between MAC and lipid solubility and the small difference between anesthetic and lethal concentrations of halothane during inhalation. The rate of halothane increase in myelin exceeded the rate in other brain subcellular membranes, indicating that a major site of halothane localization is within this subcellular membrane.

Animals↗

PubMed1994

Subcellular distribution of carbonic anhydrase and Na+,K(+)-ATPase in the brain of the hyt/hyt hypothyroid mice.

Activities of carbonic anhydrase and Na+,K(+)-ATPase in tissue homogenates and in subcellular fractions from different brain regions were studied in inherited primary hypothyroid (hyt/hyt) mice. The body weight, the weight of different brain regions, and the plasma thyroxine and triiodothyronine levels of hyt/hyt mice were significantly lower than those of the age-matched hyt/+ controls. In tissue homogenates of cerebral cortex, brain stem and cerebellum of hypothyroid mice, the activity of carbonic anhydrase (units/mg protein) was 59.2, 57.6, and 43.2%, and the activity of Na+,K(+)-ATPase (nmol Pi/mg protein/min) was 73.7, 74.4 and 68.7%, respectively, of that in corresponding regions of euthyroid littermates. The decrease in enzyme activity in tissue homogenates was also reflected in different subcellular fractions. In cerebral cortex and brain stem, carbonic anhydrase activity in cytosol, myelin and mitochondrial fractions of hypothyroid mice was about 45-50% of that in euthyroid mice, while in cerebellum the carbonic anhydrase activity in these subcellular fractions of hyt/hyt mice was only 33-38% of that in hyt/+ controls. Na+,K(+)-ATPase activity in myelin fraction of different brain regions of hyt/hyt mice was about 34-42% of that in hyt/+ mice, while in mitochondria, synaptosome and microsome fractions were about 44-52, 46-53, and 66-68%, respectively of controls. These data indicate that the activity of both carbonic anhydrase and Na+,K(+)-ATPase was affected more in the myelin than other subcellular fractions and more in the cerebellum than cerebral cortex and brain stem by deficiency of thyroid hormones.(ABSTRACT TRUNCATED AT 250 WORDS)

Aging↗

PubMed1993

Distribution of ubiquinone and ubiquinol homologues in rat tissues and subcellular fractions.

The oxidized (UQox) and reduced (UQred) forms of ubiquinone (UQ) homologues in rat tissues and subcellular fractions were analyzed to elucidate their distribution and physiological role. UQ-9 and UQ-10 were detected in all tissues studied, and UQ-9 was the predominant homologue. The total amount of UQox-10 and UQred-10 was 20-50% that of UQox-9 and UQred-9. The levels of these homologues were highest in heart with lesser amounts occurring in kidney, liver and other organs. In liver and blood plasma, the UQred homologue amounted to 70-80% of the total UQ (UQox + UQred = t-UQ). UQred was less than 30% of t-UQ in other tissues and blood cells. t-UQ was much higher in leukocytes and platelets in blood than in erythrocytes. In erythrocytes, t-UQ was exclusively located in the cell membranes. UQox and UQred were also found in all subcellular fractions isolated from liver and kidney in about the same ratio as UQred/t-UQ was present in the whole organ. The levels of UQox and UQred per mg protein in subcellular fractions from liver were highest in mitochondria, with lesser amounts present in plasma membranes, lysosomes, Golgi complex, nuclei, microsomes and cytosol. In the mitochondria, the outer membranes were richer in t-UQ than the inner membranes. In the Golgi complex, the light and intermediate fractions were rich in t-UQ when compared to the heavy fraction. The possible physiological role of UQox and UQred in tissues and subcellular fractions is discussed.

Animals↗

PubMed1990

Conversion of alkylacetylglycerol to platelet-activating factor in HL-60 cells and subcellular localization of the mediator.

A human promyelocytic leukemia (HL-60) cell line was used to investigate the conversion of 1-alkyl-2-acetyl-sn-glycerol (alkylacetyl-G) to platelet-activating factor (PAF; 1-alkyl-2-acetyl-sn-glycero-3-phosphocholine) by intact cells and in subcellular fractions in order to examine the fate of PAF synthesized de novo. Lipid extracts obtained from undifferentiated HL-60 cells incubated with [3H]alkylacetyl-G contained 2-4% of the label as [3H]PAF; several related metabolites were also detected. The yield of [3H]PAF could be dramatically increased by pretreating the cells with either oleic acid, an activator of CTP:phosphocholine cytidylyltransferase, or phenylmethylsulfonyl fluoride, an inhibitor of PAF acetylhydrolase. These results, together with a kinetic study of [3H]alkylacetyl-G metabolism, indicate the sequential participation of a cholinephosphotransferase for the conversion of [3H]-alkylacetyl-G to PAF and acetylhydrolase and transacylase activities in the remodeling pathway that metabolize the newly formed [3H]PAF to 1-[3H]alkyl-2-acyl(long chain)-sn-glycero-3-phosphocholine. The dithiothreitol-insensitive cholinephosphotransferase activity capable of converting alkylacetyl-G to PAF was localized in subcellular fractions that contain CDP-choline:1,2-dioleoyl-sn-glycerol cholinephosphotransferase (dithiothreitol-sensitive), as well as marker enzyme activities for the endoplasmic reticulum and Golgi membranes. Subcellular localization analyses also indicated that the majority of newly formed [3H]PAF and a large portion of its deacetylated metabolite were associated with the plasma membrane-containing fractions, whereas most of the 1-[3H]alkyl-2-acyl(long chain)-sn-glycero-3- phosphocholine was present in the intracellular organelles. Incubations of HL-60 cells with exogenous [3H]PAF produced a similar subcellular distribution of metabolites. Very little (less than 10%) of the [3H]PAF produced from [3H]alkylacetyl-G was released from intact cells under a variety of incubation conditions but 50% of the de novo-derived mediator was recovered in the medium of cells that were permeabilized with saponin. Our results indicate that PAF is rapidly translocated from its intracellular site of enzymatic synthesis to the plasma membrane where it is apparently sequestered in a pool that is not accessible to extracellular acceptors in contact with intact cells.

Cell Line↗

PubMed1981

Optimal assay and subcellular location of phosphatidylglycerol synthesis in lung.

Synthesis of phosphatidylglycerol from CDPdiacylglycerol and glycerol 3-phosphate by membranous subcellular fractions of rat lung and liver was optimal when assayed in the presence of bovine serum albumin and Triton X-100. Specific activities of glycerolphosphate phosphatidyltransferase in all membranous subcellular fractions of lung were several times higher than the corresponding fractions from liver. Distribution of this enzyme in subcellular fractions of lung or liver closely parallel the activity of the mitochondrial enzymes monoamine oxidase and succinate cytochrome c reductase. The phosphatidylglycerol-synthesizing activity in microsomes of both lung and liver was a minor fraction of total tissue activity and could be interpreted as due either to contamination with outer mitochondrial membrane or to a small amount of activity innate to microsomes. These results suggest that phosphatidylglycerol, which is believed to be a component of pulmonary surfactant, is synthesized by lung at a rapid rate relative to liver and that the subcellular distribution of its synthesis is similar in both tissues, with mitochondria as the major site.

Animals↗

PubMed1991

Subcellular localization of rat gastric phospholipase A2.

In the present study, we have performed experiments to gain some insight into the subcellular localization and biochemical properties of gastric mucosal phospholipase A2. After classical subcellular fractionation of whole glandular stomach mucosa, we found that gastric phospholipase A2 was essentially enriched in the 105,000 x g pellet that contains microsomes and plasma membranes. Except for the cytosol, all the subcellular fractions exhibited similar phospholipase A2 activity (i.e., optimum of pH, calcium dependence, apparent Km and positional specificity). The high-speed pellet was further characterized by ultracentrifugation on a sucrose gradient. Data showed that the sedimentation profile of phospholipase A2 was quite similar to those of plasma membrane markers and more specifically to an apical membrane marker. These results, taken together, showed that a gastric phospholipase A2 is distributed among the various subcellular fractions (as a result of cross-contamination) together with the membrane fraction on which it is associated. It is proposed that this fraction is the apical plasma membrane which would be the main site of phospholipase A2 action for arachidonic acid release. Lysophospholipase showed the same sedimentation profile as phospholipase A2, whereas acyl CoA-lysophosphatidylcholine: acyltransferase mainly sedimented with heavy microsomes. The substrate specificity of the enzyme was assessed by endogenous hydrolysis of gastric mucosal phospholipids. We were able to show that the enzyme acts at nearly the same rate on two major gastric membrane phospholipids, namely phosphatidylcholine and phosphatidylethanolamine.

Animals↗

PubMed1993

Type II phospholipase A2 in human gestational tissues: subcellular distribution of placental immuno- and catalytic activity.

The aims of this study were to determine the subcellular distribution of Type II phospholipase A2 immunoactivity (irPLA2) and in vitro net PLA2 catalytic activity in human term placenta and to establish the efficacy of previously utilised homogenisation procedures with respect to the quantitative recovery of Type II PLA2 immunoreactive and in vitro net PLA2 catalytic activity. Type II PLA2 immunoactivity and PLA2 catalytic activity recovered in 900 x g supernates prepared from placental tissue (n = 3) homogenised in low ionic strength media (sucrose 0.32 M Hepes 20 mM; phosphate-buffered saline or phosphate-buffered saline containing 3 mM EGTA) was less than 10% of that recovered following homogenisation in high ionic strength medium (ammonium sulphate 10%, w/v). The subcellular distribution of Type II PLA2 immunoactivity and PLA2 catalytic activity was established by the differential centrifugation (10,000, 20,000 and 100,000 x g) of placental homogenates (n = 3). Although Type II PLA2 immunoactivity was equally distributed throughout the particulate subcellular fractions examined, PLA2 catalytic activity increased by comparison in 100,000 x g particulate material. This apparent dissociation between irType II PLA2 and catalytic activity may indicate the presence of other types of PLA2 in this fraction. The data obtained in this study indicate that previous studies which have utilised low ionic strength extractions of human gestational tissue to characterise PLA2 catalytic activity and subcellular distribution have largely excluded the contribution made by Type II PLA2. Consequently, much of the available published data on the role of PLA2 in human parturition is inadequate. A reappraisal of this enzyme's contribution to the biochemical events associated with human pregnancy and labour is required.

Female↗

PubMed1995

Subcellular localisation and induction of NADH-sensitive acetyl-CoA hydrolase and propionyl-CoA hydrolase activities in rat liver under lipogenic conditions after treatment with sulfur-substituted fatty acids.

The effects of sulfur-substituted fatty acid analogues on the subcellular distribution and activities of acetyl-CoA and propionyl-CoA hydrolases in rats fed a high carbohydrate diet were studied. Among subcellular fractions of liver homogenates from rats fed a high carbohydrate diet (20%), the acetyl-CoA and propionyl-CoA hydrolase activities are found in the mitochondrial, peroxisome-enriched and cytosolic fractions. We have shown that the subcellular distribution of acetyl-CoA hydrolase appears to be different from the distribution propionyl-CoA hydrolase activity. Thus, the highest specific activity of acetyl-CoA hydrolase was found in the mitochondrial fraction, whereas the highest specific activity of propionyl-CoA hydrolase was found in the peroxisome-enriched fraction. Rats treated with sulfur-substituted fatty acids, i.e., 3-thiadicarboxylic acid (400 mg/day per kg body weight), showed a significant increase in acetyl-CoA hydrolase activity where the peroxisomal and cytosolic hydrolases were increased 3.9- and 2.7-fold, respectively, compared to palmitic acid treated rats. Similar results were obtained with tetradecylthioacetic acid treated rats. Propionyl-CoA hydrolase activities, in rats treated with these two peroxisome proliferating fatty acid analogues showed increased activity mainly in the mitochondrial and the cytosolic subcellular fractions. Acetyl-CoA hydrolase activity was sensitive to NADH, whereas no stimulation of the propionyl-CoA hydrolase activity was observed in the presence of NADH. The hepatic amounts of acetyl-CoA, propionyl-CoA, and free CoASH were elevated after sulfur-substituted fatty acid treatment. Sulfur-substituted fatty acids also elevated the specific acetyl-CoA hydrolase activity in the mitochondrial fraction and the propionyl-CoA hydrolase activity in the light-mitochondrial fraction. These results, therefore, suggest that acetyl-CoA hydrolase and propionyl-CoA hydrolase are two distinct proteins and that these two enzymes have a multiorganelle localisation.

Acetyl Coenzyme A↗

PubMed1993

Oxidation of alpha-tocopherol in subcellular fractions from rat brain and its possible involvement in nerve function.

The turnover rate of vitamin E is slow in nerve tissue. Therefore, we have developed in vitro techniques to study the biochemical reactions of this nutrient in brain. Subcellular fractions were isolated from the cerebral hemispheres of 4-month-old, male, Fisher 344 rats. Aliquots of fractions (500 micrograms protein) were suspended in 50 mM phosphate buffer at pH 7.4 and incubated at room temperature (20-22 degrees) or 37 degrees for 2 hr in the presence or absence of the following oxidizing agents: 1 mM tertiary butyl hydroperoxide, 10 microM linoleic acid hydroperoxide, 0.5 to 50 mM 2,2'-azobis (2-amidinopropane) dihydrochloride (ABAPH) or 0.1 to 2 mM 2,2'-azobis (2,4-dimethyl) valeronitrile (ABDVN). The latter two compounds generate free radicals upon heating. After oxidation, the subcellular fractions were sedimented, saponified and assayed for tocopherol by liquid chromatography. Linoleic acid hydroperoxide was the most potent oxidizing agent, suggesting that endogenous fatty acid peroxides (e.g. eicosanoid intermediates) are very powerful oxidizing agents. Vitamin E may play an important role in providing antioxidant protection for membranes against excessive oxidation induced by these peroxides. Tocopherol in mitochondria and microsomes was much more susceptible to oxidation than synaptosomal tocopherol. The possible reasons for this observation are: (a) mitochondria and microsomes may contain less of the other reducing agents such as sulfhydryl compounds than synaptosomes, and/or (b) the electron transport structures in the former two subcellular fractions may be facilitating oxidation of tocopherol induced by free radicals. A portion of tocopherol remained unoxidized in all subcellular fractions even at high concentrations of ABAPH, suggesting that tocopherol exists in labile and nonlabile biochemical compartments or complexes.

Amidines↗

PubMed1985

Changes in the subcellular distribution of calmodulin-kinase II during brain development.

Subcellular fractions prepared from rodent forebrain at different postnatal ages were examined for calmodulin-binding proteins using [125I]calmodulin and a gel overlay technique. Synaptic junction (SJ) fractions from newborn brain, which display purity comparable to adult SJ fractions, contain low but detectable amounts of 60 and 50 kdalton calmodulin-binding polypeptides; the latter being the major postsynaptic density protein. These polypeptides have recently been shown to be the calmodulin-binding protein subunits of calmodulin-dependent protein kinase II (CaM-kinase II). CaM-kinase II polypeptides represented the predominent calmodulin-binding proteins in nearly every subcellular fraction examined, regardless of postnatal age. Large increases were observed in the CaM-kinase II content of every subcellular fraction throughout postnatal development. During development, a striking shift in the subcellular distribution of CaM-kinase Ii was observed. Over 4 times as much CaM-kinase II was cytosolic relative to particulate in newborn brain while this ratio was completely reversed in adult brain. Large age-dependent increases in particulate-associated CaM-kinase II were observed in highly purified synaptic plasma membrane (5-fold) and SJ (14-fold) fractions. The CaM-kinase II content of SJ fractions increased approximately 70% between days 24 and 90, a period in development that follows the most active stages of synapse formation in situ. In adult brain, approximately 60% of CaM-kinase II in crude synaptosomal fractions (P2-INT) was recovered in SJ fractions. The CaM-kinase II in SPM fractions from all developmental ages resists solubilization in Triton X-100 and greater than 90% is recovered in SJ fractions. These studies indicate that during brain development the accumulation of SJ-associated CaM-kinase II represents an important process in the molecular and enzymatic maturation of CNS postsynaptic structures.

Age Factors↗

PubMed1995

Expression and subcellular distribution of phosphoenolpyruvate carboxykinase in primary cultures of rabbit kidney proximal tubule cells: comparative study with renal and hepatic PEPCK in vivo.

The behaviour of the phosphoenolpyruvate carboxykinase (PEPCK) in rabbit proximal tubule cells in primary culture was investigated and compared with renal and hepatic PEPCK in vivo. The enzyme activity decreased rapidly in rabbit proximal tubule cells developed in hormonally defined medium supplemented with glucose and insulin. In this condition, the cytosolic form disappears with time. Without glucose and insulin, the subcellular location of PEPCK is similar to the location observed in proximal tubule freshly isolated and in renal cortex, with approx. 50% of mitochondrial form and approx. 50% of cytosolic form. However, the levels of mRNA that encode the cytosolic PEPCK are not detectable in cell cultures, whatever the medium composition. Treatment with dibutyryl cAMP caused a 14-fold induction of PEPCK mRNA in 6 h. This result indicates that the transcription of cytosolic PEPCK can be induced in cell cultures. Lactate or pyruvate additions did not modify the levels of PEPCK mRNA whereas specific activity increased rapidly, suggesting an activation of an inactive form in cell cultures. Moreover, lactate induced increased specific activity of the sole mitochondrial form while pyruvate induced increased specific activities of both mitochondrial and cytosolic form. Thus, subcellular location of PEPCK in rabbit proximal tubule cells appears to be modulated by the available substrate in culture medium. This observation parallels the changes observed in vivo since a modification of subcellular location of this enzyme was seen between fed and fasted rabbit, when subcellular distribution remains similar between fed and starved rats. Moreover, in the fasted liver of rabbit, a decrease of the mitochondrial PEPCK specific activity is seen concomitant with an increase in cytosolic PEPCK activity. These results point out the relative contributions of the cytosolic and mitochondrial PEPCK to rabbit gluconeogenesis.

Animals↗

PubMed1982

Subcellular distribution of weak acids and bases in adrenal medulla.

1. Studies of the subcellular distribution of two acids and three bases in adrenal medulla support the hypothesis that the charge on a drug molecule is an important factor determining its subcellular distribution in this tissue. 2. Acidic drugs carrying a negative charge do not bind well to adrenomedullary subcellular particles and accumulate predominantly in cell supernatant. 3. Basic drugs tend to bind more extensively to adrenomedullary intracellular organelles and distribute more uniformly over all subcellular fractions.

Acids↗

PubMed1990

Subcellular and regional distribution of casein kinase II and initiation factor 2 activities during rat brain development.

The possible relationship between the subcellular and regional distribution of the activities of initiation factor 2 and casein kinase II, responsible for the phosphorylation of the beta subunit of the factor, has been studied during postnatal rat brain development. Both activities have been measured in four brain regions: diencephalon, hemispheres, cerebellum and brain stem, and in two subcellular fractions: postmicrosomal supernatant and the protein fraction associated with ribosomes, or crude initiation factors fraction. The specific activity of both the factor and the protein kinase is much higher in the protein fraction associated with ribosomes than in the soluble fraction and slightly higher in the hemispheres than in the other three regions. Changes in the activity of both proteins are in parallel with development, the activities increase in the postmicrosomal supernatant and decrease in the fraction associated with ribosomes from suckling (5-day-old) to adult (60-day-old) animals. The total activity of the factor and its kinase, calculated by summation of the activities of both subcellular fractions, does not change during development, and the distribution of activities between the two subcellular fractions observed during brain development, appears as an attractive regulation mechanism for the function of both proteins.

Aging↗

PubMed2006

Prediction of protein subcellular location using hydrophobic patterns of amino acid sequence.

The function of eukaryotic protein is closely correlated with its subcellular location. The number of newly found protein sequences entering into data banks is rapidly increasing with the success of human genome project. It is highly desirable to predict a protein subcellular automatically from its amino acid sequence. In this paper, amino acid hydrophobic patterns and average power-spectral density (APSD) are introduced to define pseudo amino acid composition. The covariant-discriminant predictor is used to predict subcellular location. Immune-genetic algorithm (IGA) is used to find the fittest weight factors which are very important in this method. As such, high success rates are obtained by both self-consistency test (86%) and jackknife test (73%). More than 80% predictive accuracy is achieved in independent dataset test. The results demonstrate that the proposed method is practical. And, the method illuminates that the protein subcellular location can be predicted from its surface physio-chemical characteristic of protein folding.

Algorithms↗

PubMed2005

Subcellular positioning of F plasmid mediated by dynamic localization of SopA and SopB.

SopA, SopB proteins and the cis-acting sopC DNA region of F plasmid are essential for partitioning of the plasmid, ensuring proper subcellular positioning of the plasmid DNA molecules. We have analyzed by immunofluorescence microscopy the subcellular localization of SopA and SopB. The majority of SopB molecules formed foci, which localized frequently with F plasmid DNA molecules. The foci increased in number in proportion to the cell length. Interestingly, beside the foci formation, SopB formed a spiral structure that was dependent on SopA, which also formed a spiral structure, independent of the presence of SopB, and these two structures partially overlapped. On the basis of these results and previous biochemical studies together with our simulations, we propose a theoretical model named "the reaction-diffusion partitioning model", using reaction-diffusion equations that explain the dynamic subcellular localization of SopA and SopB proteins and the subcellular positioning of F plasmid. We hypothesized that sister copies of plasmid DNA compete with each other for sites at which SopB multimer is at the optimum concentration. The plasmid incompatibility mediated by the Sop system might be explained clearly by this hypothesis.

Animals↗

PubMed1998

Heterogeneous forms of adenotin-1 of different subcellular localization.

The localization of the low-affinity adenosine binding protein adenotin-1 with respect to distribution in rat organs and subcellular compartments was investigated. Adenotin-1 was characterized by 5'-N-ethylcarboxamido[2,8-3H]adenosine ([3H]NECA) binding and Western blotting. Cytosolic as well as membrane fractions of all tissues contained adenotin-1. Highest levels of membrane-bound adenotin-1 were found in the liver (liver > kidney approximately spleen approximately lung > forebrain approximately cerebellum > fat heart - striated muscle), whereas highest levels of cytosolic adenotin-1 were detected in spleen, liver, lung and fat. Subcellular fractions from rat liver were prepared by differential and density gradient centrifugation. Like the homologous proteins endoplasmin or gp96, adenotin-1 is enriched in the endoplasmic reticulum. Cytosolic and membrane-bound adenotin-1 species are pharmacologically distinct, because in the liver particulate fraction adenotin-1 showed a more rapid binding kinetics, a twofold lower affinity for [3H]NECA (KD 227 nM vs. 105 nM) and a sevenfold higher affinity for 2-chloroadenosine than the cytosolic protein (Ki 1.48 microM vs. 9.25 microM). In rat liver cytosol, two different binding sites were found, which differed in [3H]NECA binding kinetics and displayed a hundredfold difference in their affinity for 2-chloro-5'-N-methylcarboxamidoadenosine (Ki 45.8 nM vs. 4.76 microM). The presence of adenotin-1 in subcellular fractions, as determined by radioligand binding, was confirmed by Western blotting. Adenotin-1 was detected as a 98-kDa band in all rat liver subcellular fractions, which agrees with the molecular mass determined for the purified protein. In the cytosol, a 65-kDa hand was labeled more intensely than the 98-kDa band. This additional band probably represents the pharmacologically distinct species of adenotin-1 found in the cytosol.

Adenosine-5'-(N-ethylcarboxamide)↗

PubMed2002

Artificial neural network model for predicting protein subcellular location.

The function of a protein is closely correlated to its subcellular location. Is it possible to utilize a bioinformatics method to predict the protein subcellular location? To explore this problem, proteins are classified into 12 groups (Protein Eng. 12 (1999) 107-118) according to their subcellular location: (1) chloroplast, (2) cytoplasm, (3) cytoskeleton, (4) endoplasmic reticulum, (5) extracellular, (6) Golgi apparatus, (7) lysosome, (8) mitochondria, (9) nucleus, (10) peroxisome, (11) plasma membrane and (12) vacuole. In this paper, the neural network method was proposed to predict the subcellular location of a protein according to its amino acid composition. Results obtained through self-consistency, cross-validation and independent dataset tests are quite high. Accordingly, the present method can serve as a complement tool for the existing prediction methods in this area.

Algorithms↗

PubMed1999

Cellular and subcellular localization of a newly identified member of the protein 4.1 family, brain 4.1, in the cerebellum of adult and postnatally developing rats.

For obtaining a deeper insight into the properties of a newly characterized member of the protein 4.1 family, brain 4.1, the cellular and subcellular localization was investigated in the cerebellar cortex of adult and postnatally developing rats. Fluorescent immunohistochemical observations showed that brain 4.1 localized predominantly to glomeruli in the granular layer and throughout the molecular layer in adult rat cerebellar cortex. Analysis of subcellular localization of brain 4.1 by immuno-electron microscopy further demonstrated that presynaptic terminals of mossy fibers and parallel fibers, cytoplasm of granule cells and cytoplasm and/or processes of glial cells contained brain 4.1 while postsynaptic regions of the dendrites of granule cells and Purkinje cells, axons and myelin sheaths did not. Thus, one of the major subcellular destination of brain 4.1 was presynaptic terminal in the cerebellum. This was further supported by the fact that the immunostaining pattern of brain 4.1 in the cerebellum changed in a similar way to that of a synaptic terminal marker, synaptophysin during the postnatal development. Immunoblot analysis also demonstrated that contents of brain 4.1 isoforms varied in parallel with the changes of the immunostaining pattern. Biochemical analysis confirmed the presence of brain 4.1 at synaptic terminals, but there was no obvious correlation between each isoform and its subcellular localization. These results suggested that brain 4.1 is involved in the formation and maintenance of synapse as a membrane skeletal component at presynaptic terminals in the cerebellum.

Aging↗