Biomedical subjects

A Alcázar

Publications and source records attributed to A Alcázar.

At least 19 recordsLinked to original sources

PubMed2006

Anti-heat shock protein 90beta antibodies are detected in patients with multiple sclerosis during remission.

Antibodies against heat shock protein 90beta (Hsp90beta) recognize the antigen on the surface of oligodendrocyte precursor cells and induce a decrease in the oligodendrocyte population of cell cultures. This study investigated the presence of anti-Hsp90beta antibodies in patients with multiple sclerosis (MS) during remission. Anti-Hsp90beta antibodies were detected in the cerebrospinal fluid (CSF) of patients with a specific and sensitive western blot procedure using Hsp90beta from cell membrane fraction. The number of patients with anti-Hsp90beta antibodies detected in the CSF was significantly greater in MS patients than in control patients with other neurological diseases, which included patients with other inflammatory neurological diseases. CSF anti-Hsp90beta antibody levels were significantly higher in MS patients than in control patients. The presence of anti-Hsp90beta antibodies in the CSF of MS patients during remission could suggest a potential pathogenic role for these autoantibodies in MS.

Adult↗

PubMed2006

Detection of anti-heat shock protein 90 beta (Hsp90beta) antibodies in cerebrospinal fluid.

Antibodies against heat shock protein 90 beta (Hsp90beta) recognize the antigen on the cell surface of the oligodendrocyte precursor cells and cause a decrease of oligodendrocyte population in cell cultures. These antibodies have been found in patients with multiple sclerosis (MS). This report describes an original and sensitive method to detect anti-Hsp90beta antibodies in cerebrospinal fluid (CSF) using a western blot procedure. We have developed the method for autoantibody detection using Hsp90beta from cell membrane fraction instead of commercial Hsp90beta as antigen. The presence of anti-Hsp90beta antibodies in CSF of MS patients may play a pathogenic role in MS, and a large-scale study is needed to establish a possible diagnostic value of these antibodies in MS patients.

Animals↗

PubMed2004

[Sedation in digestive endoscopy. Results of a hospital survey in Catalonia (Spain)].

INTRODUCTION: The need for sedation is increasing in digestive endoscopy units (DEU). There are no data on the use of sedation in DEU in Catalonia (Spain). OBJECTIVE: To evaluate the use of sedation in DEU in Catalonia. MATERIAL AND METHOD: A questionnaire on the practice of sedation was designed and sent to the heads of medical and nursing staff of the DEU of 63 public and private hospitals in Catalonia. Two mailings were sent with an interval of three months between each. The questionnaire included 62 items on the characteristics of the hospital and the DEU, number of explorations, frequency of sedation use, drugs employed, participation of an anesthesiologist, use of monitoring, and complications. RESULTS: Forty-four DEU (70%) corresponding to 31 public hospitals and 13 private hospitals completed the questionnaire. Evaluation of sedation patterns was based on 105,904 explorations performed in the various DEU (56,453 gastroscopies, 47,278 colonoscopies and 2,173 endoscopic retrograde cholangiopancreatographies (ERCP) in 2001. Sedation, sedation-analgesia or anesthesia was used in 17% of gastroscopies, 61% of colonoscopies and 100% of ERCP. Sedation was administered by an anesthesiologist in 7% of gastroscopies, 25% of colonoscopies and 38% of ERCP. Anesthesiologist administration was more frequent in private than in public centers (gastroscopies: 25% vs. 2%; colonoscopies: 57% vs. 9%, p < 0.001). No deaths associated with the use of sedation were reported. Eighty-nine percent of the DEU complied with standard recommendations for the practice of sedation. CONCLUSIONS: In Catalonia, the use of sedation is highly variable, depending on the endoscopic procedure and the DEU. Use of sedation in infrequent in gastroscopy, fairly widespread in colonoscopy and routine in ERCP. Anesthesiologist administration is significantly more frequent in private hospitals. Most DEU follow standard sedation practices.

Anesthesia↗

PubMed2003

Caspase inhibitors protect against neuronal apoptosis induced by cerebrospinal fluid from multiple sclerosis patients.

Neuronal apoptosis has recently been implicated in multiple sclerosis (MS). Apoptotic cell death of neurons is induced in cultures exposed to cerebrospinal fluid (CSF) from MS patients. Since caspases are essential in the regulation of apoptosis, direct evidence was sought linking caspases to CSF-induced neuronal death. Caspase activity was measured in cell extracts from MS CSF-treated cultured neurons by the cleavage of caspase-1 and caspase-3 substrates. Caspase-3 activity, but not caspase-1, was induced in neuronal cultures in response to MS CSF treatment. This caspase-3 activity was inhibited in vitro by Ac-YVAD-cmk and Ac-DEVD-cmk caspase inhibitors. Treatment of MS CSF-incubated neuronal cells with these caspase inhibitors completely preserved neuronal survival and largely attenuated DNA fragmentation detected in situ. These findings show that neuronal cells are rescued from MS CSF-induced death by caspase inhibitors and suggest ways to treat MS.

Adolescent↗

PubMed2001

Possible mechanisms involved in the down-regulation of translation during transient global ischaemia in the rat brain.

The striking correlation between neuronal vulnerability and down-regulation of translation suggests that this cellular process plays a critical part in the cascade of pathogenetic events leading to ischaemic cell death. There is compelling evidence supporting the idea that inhibition of translation is exerted at the polypeptide chain initiation step, and the present study explores the possible mechanism/s implicated. Incomplete forebrain ischaemia (30 min) was induced in rats by using the four-vessel occlusion model. Eukaryotic initiation factor (eIF)2, eIF4E and eIF4E-binding protein (4E-BP1) phosphorylation levels, eIF4F complex formation, as well as eIF2B and ribosomal protein S6 kinase (p70(S6K)) activities, were determined in different subcellular fractions from the cortex and the hippocampus [the CA1-subfield and the remaining hippocampus (RH)], at several post-ischaemic times. Increased phosphorylation of the alpha subunit of eIF2 (eIF2 alpha) and eIF2B inhibition paralleled the inhibition of translation in the hippocampus, but they normalized to control values, including the CA1-subfield, after 4--6 h of reperfusion. eIF4E and 4E-BP1 were significantly dephosphorylated during ischaemia and total eIF4E levels decreased during reperfusion both in the cortex and hippocampus, with values normalizing after 4 h of reperfusion only in the cortex. Conversely, p70(S6K) activity, which was inhibited in both regions during ischaemia, recovered to control values earlier in the hippocampus than in the cortex. eIF4F complex formation diminished both in the cortex and the hippocampus during ischaemia and reperfusion, and it was lower in the CA1-subfield than in the RH, roughly paralleling the observed decrease in eIF4E and eIF4G levels. Our findings are consistent with a potential role for eIF4E, 4E-BP1 and eIF4G in the down-regulation of translation during ischaemia. eIF2 alpha, eIF2B, eIF4G and p70(S6K) are positively implicated in the translational inhibition induced at early reperfusion, whereas eIF4F complex formation is likely to contribute to the persistent inhibition of translation observed at longer reperfusion times.

Animals↗

PubMed2000

Two different signal transduction pathways are implicated in the regulation of initiation factor 2B activity in insulin-like growth factor-1-stimulated neuronal cells.

Eukaryotic initiation factor eIF-2B plays an important role in translation regulation and has been suggested to be implicated in the increased protein synthesis promoted in response to growth factors. We have used primary cultured neurons to delineate the signaling pathways by which insulin-like growth factor-1 (IGF-1), which plays a critical role in the survival of neuronal cells, promotes eIF-2B and protein synthesis activation. Treatment of cortical neurons with IGF-1 (100 ng/ml) for 30 min stimulates [(3)H]methionine incorporation, and a parallel increase in eIF-2B activity was observed. Wortmannin and LY294002 reversed both effects, indicating that phosphatidylinositol 3-kinase mediates IGF-1-induced protein synthesis and eIF-2B activation. IGF-1 induced glycogen synthase kinase-3 (GSK-3) inactivation in a phosphatidylinositol 3-kinase-dependent fashion because it is inhibited by wortmannin and LY294002. By using GSK-3 immunoprecipitated from untreated and IGF-1-treated cells, we demonstrate the phosphorylation of eIF-2B coincident with its inactivation. The treatment of cortical neurons with IGF-1 also promoted the activation of mitogen-activated protein kinase (MAPK). The MAPK-activating kinase (MEK) inhibitor PD98059 inhibited MAPK activation and reversed IGF-1-induced protein synthesis and eIF-2B activation. These findings suggest that IGF-1-induced eIF-2B activation on neurons is promoted through phosphatidylinositol 3-kinase and GSK-3 kinase, and we report an IGF-1-induced MEK/MAPK activation pathway implicated in eIF-2B activation.

Amino Acid Sequence↗

PubMed2000

Axonal damage induced by cerebrospinal fluid from patients with relapsing-remitting multiple sclerosis.

The importance of axonal damage in multiple sclerosis (MS) has been recently stressed in proton magnetic resonance spectroscopy and pathological studies, but the exact mechanism producing this damage is unknown. The aim of our study was to ascertain whether soluble mediators present in the cerebrospinal fluid (CSF) of patients with relapsing-remitting MS could induce neuron injury in culture. Different biochemical and cytochemical parameters were determined in primary embryonal rat neuron cultures following 8 days of exposure to CSF. Cytotoxic activity was evaluated with a blue formazan production colorimetric assay. Morphological and immunocytochemical studies performed with antibodies against beta-tubulin revealed neuritic fragmentation, axonal damage and cellular shrinkage indicating apoptosis. Detection of apoptosis was carried out using the fluorescent DNA-binding dye Hoechst 33342, as well as by a Terminal deoxynucleotidyl transferase-mediated dUTP Nick End-Labeling assay. We observed that soluble factors in CSF from patients with "aggressive" MS i.e, those with poor recovery after relapses, induced neurite breakdown and neuronal apoptosis in cultures. Neuron injury is not related with blood-brain barrier dysfunction nor with IgG index. Interestingly, CSF from patients with "non-aggressive" MS i.e., relapsing-remitting patients with a good recovery after relapses, did not induce any damage. In conclusion, we report that CSF from patients with aggressive MS bears soluble mediators that induce axonal damage and apoptosis of neurons in culture. These mediators can be present during the first attack of the disease, and the neuronal damage caused could be related to the functional deficit of these MS patients.

Adolescent↗

PubMed1999

Resistance of initiation factor 2 (eIF-2alpha) kinases to staurosporine: an approach for assaying the kinases in crude extracts.

We studied the effect of staurosporine on two well characterised mammalian eIF-2alpha kinases, the heme-regulated translational inhibitor (HRI), and interferon-inducible double-stranded RNA-activated protein kinase (PKR). Both pure eIF-2 and a synthetic peptide used to measure the activity of purified or immunoprecipitated enzymes (sequence ILLSELSRRRIRAI) were phosphorylated with purified enzymes and crude preparations of tissues or cells in the presence of the inhibitor. In the presence of 0.25 microM staurosporine (a concentration which completely inhibits a wide range of Ser/Thr protein kinases), the phosphorylation of eIF-2alpha by HRI and PKR was not inhibited. The lack of response of eIF-2alpha kinases to staurosporine allowed us to measure PKR activity in salt washed postmicrosomal supernatants without previous purification of the enzyme. In the presence of poly(I):poly(C), the PKR activator, we detected both an increased phosphorylation of eIF-2alpha and an increment in the autophosphorylation of PKR. We also confirmed an induction of PKR in cultured neuronal cells after treatment with interferon. The results obtained following phosphorylation of the synthetic peptide with crude extracts are less conclusive. Although its phosphorylation is specific because it displaces eIF-2 phosphorylation, and the presence of staurosporine prevents its phosphorylation by other serine/threonine kinases, it is a rather poor substrate for PKR.

Animals↗

PubMed1998

Induction of apoptosis by cerebrospinal fluid from patients with primary-progressive multiple sclerosis in cultured neurons.

We have studied the noxious effect of cerebrospinal fluids (CSF) from patients with primary-progressive multiple sclerosis (MS) on cultured neurons. Cells were exposed to CSF for 8 days and the possible neuronal damage was determined. Morphological studies with phase-contrast microscopy showed cellular shrinkage indicating apoptosis. CSF-induced apoptosis as evidenced by the fluorescent DNA-binding dye Hoechst 33342, as well as by the TUNEL-reaction, was only present in primary-progressive MS patients with a worsening disease. This neuron injury did not correlate with blood-brain barrier dysfunction nor with intrathecal IgG synthesis. On the contrary, CSF from either stable primary-progressive or other non-inflammatory neurological diseases, did not induce any culture damage. Undetectable or low similar tumor necrosis factor-alpha (TNF-alpha) levels (range to 8.7 pg/ml) were found in the CSFs tested regardless they damage cultures or not. These results suggest that soluble factors, other than TNF-alpha, molecules transudated from blood or IgG, present in the CSF of active primary-progressive patients with MS induce neuronal apoptosis.

Animals↗

PubMed1998

Increased activity of eukaryotic initiation factor 2B in PC12 cells in response to differentiation by nerve growth factor.

Translational rates, and activities and levels of initiation factors 2 and 2B were assessed in rat pheochromocytoma cells upon nerve growth factor (NGF) treatment. Two or 5 days of exposure to NGF caused significant quantitative increases in protein synthesis rate that are deemed necessary for neuronal differentiation. Changes in initiation factor 2 activity, as measured by its capacity to form a ternary complex, occur parallel to the observed changes in protein synthesis. Nevertheless, neither the intracellular levels of the initiation factor 2 nor the degree of phosphorylation of its alpha subunit can justify this increased activity. Interestingly, initiation factor 2B activity increases parallel to the neurite outgrowth, being significantly higher after 5 days of exposure to NGF, and could be responsible for the elevated rate of protein synthesis. No significant changes in the levels of eukaryotic initiation factor 2B, as determined with two different antibodies against the gamma and epsilon subunits of the factor, were observed, implying that the increased activity should be regulated by factors other than its cellular concentration. Our results support the hypothesis that initiation factor 2B may play a role in the biochemical events controlling the differentiative growth factor-induced signaling pathway in these cells.

Animals↗

PubMed1998

The intraischemic and early reperfusion changes of protein synthesis in the rat brain. eIF-2 alpha kinase activity and role of initiation factors eIF-2 alpha and eIF-4E.

Rats were subjected to the standard four-vessel occlusion model of transient cerebral ischemia (vertebral and carotid arteries). The effects of normothermic ischemia (37 degrees C) followed or not by 30-minute reperfusion, as well as 30-minute postdecapitative ischemia, on translational rates were examined. Protein synthesis rate, as measured in a cell-free system, was significantly inhibited in ischemic rats, and the extent of inhibition strongly depended on duration and temperature, and less on the model of ischemia used. The ability of reinitiation in vitro (by using aurintricarboxylic acid) decreased after ischemia, suggesting a failure in the synthetic machinery at the initiation level. Eukaryotic initiation factor 2 (eIF-2) presented almost basal activity and levels after 30-minute normothermic ischemia, and the amount of phosphorylated eIF-2 alpha in these samples, as well as in sham-control samples, was undetectable. The decrease in the levels of phosphorylated initiation factor 4E (eIF-4E) after 30-minute ischemia (from 32% to 16%) could explain, at least partially, the impairment of initiation during transient cerebral ischemia. After reperfusion, eIF-4E phosphorylation was almost completely restored to basal levels (29%), whereas the level of phosphorylated eIF-2 alpha was higher (13%) than in controls and ischemic samples (both less than 2%). eIF-2 alpha kinase activity in vitro as measured by phosphorylation of endogenous eIF-2 in the presence of ATP/Mg2+, was higher in ischemic samples (8%) than in controls (4%). It seems probable that the failure of the kinase in phosphorylating eIF-2 in vivo during ischemia is due to the depletion of ATP stores. The levels of the double-stranded activated eIF-2 alpha kinase were slightly higher in ischemic animals than in controls. Our results suggest that the modulation of eIF-4E phosphorylation could be implicated in the regulation of translation during ischemia. On the contrary, phosphorylation of eIF-2 alpha, by an eIF-2 alpha kinase already activated during ischemia, represents a plausible mechanism for explaining the inhibition of translation during reperfusion.

Animals↗

PubMed1997

Calcium mobilization by ryanodine promotes the phosphorylation of initiation factor 2alpha subunit and inhibits protein synthesis in cultured neurons.

Protein synthesis plays an important role in the viability and function of the cell. There is evidence indicating that Ca2+ may be a physiological regulator of the translational process. In the present study, the effect of agents that increase intracellular calcium levels by different mechanisms, as well as repercussion on the rate of protein synthesis, including phosphorylation of initiation factor 2alpha subunit, and double-stranded RNA-dependent eIF-2alpha kinase (PKR) activity were analyzed. Glutamate (100 microM) and K+ (60 mM), which increase intracellular calcium levels (the former mostly by the influx of extracellular calcium via voltage-sensitive calcium channels, and the latter by receptor-operated calcium channels), and carbachol (1 mM), as well as glutamate, which mobilizes intracellular calcium from the endoplasmic reticulum via activation of inositol 1,4,5-trisphosphate receptor, did not modify any of the analyzed parameters. Nevertheless, 100 nM ryanodine, which increases intracellular calcium concentration by activating the ryanodine receptor, promoted a significant decrease in the rate of protein synthesis and increased both initiation factor 2alpha subunit phosphorylation and PKR activity. From our results, we can conclude that inhibition of protein synthesis is dependent on the mobilization of intracellular calcium from internal stores. Moreover, they strongly suggest that this inhibition is only promoted when calcium is increased via ryanodine receptor, and possibly by activation of PKR activity.

Animals↗

PubMed1996

Changes in the phosphorylation of eukaryotic initiation factor 2 alpha, initiation factor 2B activity and translational rates in primary neuronal cultures under different physiological growing conditions.

Phosphorylation of the alpha-subunit of eukaryotic initiation factor 2 (eIF-2) is one of the best known mechanisms regulating protein synthesis in a wide range of eukaryotic cells, from yeast to human. To determine whether this mechanism operates in primary neuronal cells, we have cultured primary neuronal cells for 7 days under two optimal growing conditions, complete medium (containing 15% serum) and serum-free medium, and determined the protein synthesis rate, eukaryotic initiation 2 and 2B (eIF-2B) activities, as well as the level of phosphorylation of eIF-2. Cells cultured in serum-free medium exhibited a lower rate of protein synthesis (75%), concomitant to a decreased eIF-2 activity (71%), and slightly higher eIF-2(alpha P) levels (from 10 to 16% of total eIF-2) with respect to cells cultured in complete media. eIF-2B activity, as measured at saturating eIF-2. GDP concentrations (assay independent on the presence of eIF-2(alpha P)) was similar under the two culture conditions. When neurons cultured in serum-free medium are exposed to complete medium for only 24 h, there is a clear decrease in the phosphorylation of eIF-2 alpha (16-3%). This decrease correlates in time with an increase in the protein synthesis rate (154%), as well as eIF-2 activity (236%). The increased levels of eIF-2(alpha P), a competitive inhibitor of eIF-2B in the guanine-exchange reaction, are responsible for the decreased eIF-2B activity found in the neurons cultured in serum-free medium. Additionally, eIF-2(alpha P) is accountable for the lower effect of exogenous eIF-2B in ternary complex formation from preformed eIF-2. GDP in the serum-free media. These changes in phosphorylation of eIF-2 alpha in normal mammalian cells in response to changes in the extracellular medium are reported here for the first time.

Animals↗

PubMed1995

Phosphorylation of initiation factor 2 alpha subunit and apoptosis in Ca2+ ionophore-treated cultured neuronal cells.

The initial step of protein synthesis is regulated by the eukaryotic initiation factor 2 (eIF-2) whose phosphorylation in the alpha subunit by specific kinases, as double-stranded RNA-dependent protein kinase (PKR), produces an inhibition of the translational rates. Besides, intracellular Ca2+ mobilization has been associated with an increase in the PKR activity. Our results show, in primary neuronal cultures, that the treatment with the Ca2+ ionophore A23187 induces an increase in the phosphorylation of the alpha subunit of eIF-2 (eIF-2 alpha) and inhibition of protein synthesis. Those biochemical changes run parallel to the appearance of specific apoptosis characteristics such as cell shrinkage, segmentation of chromatin into small round bodies, and cleavage of DNA into 180 bp multimers. These results indicate that one of the targets during the process of apoptosis induced by the rise in the intracellular Ca2+, could be eIF-2 factor.

Animals↗

PubMed1995

Purification and characterization of guanine nucleotide-exchange factor, eIF-2B, and p37 calmodulin-binding protein from calf brain.

Eukaryotic initiation factor 2B, or guanine nucleotide-exchange factor, has been purified for the first time from the brain by a novel procedure that allows the purification of initiation factor 2 as well and uses a salt wash postmicrosomal supernatant as starting material. The procedure includes a three-part chromatographic step in heparin-Sepharose and in SP-5PW and diethylaminoethyl-5PW ion-exchange high-performance chromatographies. The purification of the factors was followed by measuring activity in the guanine nucleotide-exchange assay and the capacity of initiation factor 2 to form a ternary complex with the initiation form of methionyl-tRNA and GTP. The method yields guanine nucleotide-exchange factor (75%) and highly purified initiation factor 2 (> 95%), which are separated in the last step. The exchange factor from the brain is a multimeric protein with five subunits of molecular masses of 82, 65, 52, 42, and 30 kDa; it stimulates ternary complex formation in the presence of GDP, and this activity is inhibited by N-ethylmaleimide. A 37-kDa protein that copurifies with initiation factors is characterized in this study as a new calmodulin-binding protein (p37); it is highly phosphorylated by casein kinase activities and can comigrate with the alpha subunit of initiation factor 2 under standard sodium dodecyl sulfate electrophoresis conditions.

Amino Acid Sequence↗

PubMed1994

Phosphorylation of the alpha subunit of initiation factor 2 correlates with the inhibition of translation following transient cerebral ischaemia in the rat.

Rats were subjected to the standard four-vessel occlusion model of cerebral transient ischaemia (vertebral and carotid arteries) for 15 and 30 min. After a 30 min recirculation period, protein synthesis rate, initiation factor 2 (eIF-2) and guanine nucleotide exchange factor (GEF) activities, and the level of phosphorylation of the alpha subunit of eIF-2 (eIF-2 alpha) were determined in the neocortex region of the brain from sham-operated controls and ischaemic animals. Following reversible cerebral ischaemia, the protein synthesis rate, as measured in a cell-free system, was significantly inhibited (70%) in the ischaemic animals. eIF-2 activity, as measured by its ability to form a ternary complex, also decrease parallel to the decrease in protein synthesis. As eIF-2 activity was assayed in the presence of Mg2+ and GTP-regenerating capacity, the decrease in ternary-complex formation indicated the possible impairment of GEF activity. Since phosphorylated eIF-2 [eIF-2(alpha P)] is a powerful inhibitor of GEF, the levels of phosphorylated eIF-2 alpha were determined, and an increase from 7% phosphorylation in sham control rats to 20% phosphorylation in 15 min and 29% phosphorylation in 30 min in ischaemic rats was observed, providing evidence for a tight correlation of phosphorylation of eIF-2 alpha and inhibition of protein synthesis. Moreover, GEF activity measured in the GDP-exchange assay was in fact inhibited in the ischaemic animals, proving that protein synthesis is impaired by the presence of eIF-2(alpha P), which blocks eIF-2 recycling.

Animals↗

PubMed1994

Protein synthesis in the developing rat liver: participation of initiation factors eIF-2 and eIF-2B.

We studied the decline in protein synthesis in the developing liver in suckling rats (4 to 10 days) and adult rats (2 mo). The rate of protein synthesis was measured with a cell-free system and compared with the activity of two initiation factors, eukaryotic initiation factor-2 and eukaryotic initiation factor-2B, and with casein kinase II, which phosphorylates both factors in vitro. The specific activity of the three parameters decreased in adult rats compared with suckling rats and in parallel to the rate of protein synthesis. Quantification of eukaryotic initiation factor-2 in the ribosomal salt wash and in the postmicrosomal supernatant showed that both the specific activity and the levels of eukaryotic initiation factor-2, are much higher in the ribosomal salt wash fractions than in postmicrosomal supernatants, but no differences were found between the two age groups. The eukaryotic initiation factor-2/ribosome ratio was higher in adult rats than in suckling rats, and this parameter seems to be inversely proportional to the rate of protein synthesis. The phosphorylation state of eukaryotic initiation factor-2 alpha, as determined by isoelectric focusing followed by protein immunoblotting, revealed very low and equal levels of phosphorylation in the two animal groups. The lack of changes in eukaryotic initiation factor-2 levels and phosphorylation status suggests that a decrease in the activity or levels of eukaryotic initiation factor-2B could be responsible for the decrease in eukaryotic initiation factor-2 activity and account for, at least in part, the differences observed in the rates of protein synthesis.

Aging↗

PubMed1993

Translational initiation factor eIF-2 subcellular levels and phosphorylation status in the developing rat brain.

We have quantified the levels of the alpha subunit of initiation factor 2 (eIF-2) in the postmicrosomal supernatant and the ribosomal salt wash fractions from suckling and adult rat brain. The levels of eIF-2 in the ribosomal salt wash decrease in adult with respect to that present in suckling rat brain, but the total amount remains fairly constant, and a very close parallelism exists between the eIF-2 associated with ribosomes and RNA levels in the microsomal fraction in the two age groups. The phosphorylation state of eIF-2 alpha, as determined by isoelectric focusing followed by protein immunoblotting, in the same subcellular fractions, did not reveal the presence of the phosphorylated form in any of the fractions studied. These results suggest that phosphorylation of the alpha subunit is not implied in the regulation of protein synthesis initiation during brain development, and some other component regulates both the number of active ribosomes and eIF-2 levels in microsomes.

Aging↗