scholarly journals Deriving accurate molecular indicators of protein synthesis through Raman-based sparse classification

2021 ◽  
Author(s):  
N. Pavillon ◽  
N. I. Smith
Keyword(s):  
Protein Synthesis ◽  
Linear Models ◽  
Linear Method ◽  
Classification Performance ◽  
Experimental Conditions ◽  
Protein Levels ◽  
Spectral Coefficients ◽  
Rna Accumulation ◽  
Molecular Indicators

AbstractRaman spectroscopy has the ability to retrieve molecular information from live biological samples non-invasively through optical means. Coupled with machine learning, it is possible to use the large amount of information contained in a Raman spectrum to create models that can predict the state of new samples based on statistical analysis from previous measurements. Furthermore, in case of linear models, the separation coefficients can be used to interpret which bands are contributing to the discrimination between experimental conditions, which correspond here to single-cell measurements of macrophages under in vitro immune stimulation. We here evaluate a typical linear method using discriminant analysis and PCA, and compare it to regularized logistic regression (Lasso). We find that the use of PCA is not beneficial to the classification performance. Furthermore, the Lasso approach yields sparse separation vectors, since it suppresses spectral coefficients which do not improve classification, making interpretation easier. To further evaluate the approach, we apply the Lasso technique to a well-defined case where protein synthesis is inhibited, and show that the separating features are consistent with RNA accumulation and protein levels depletion. Surprisingly, when Raman features are selected purely in terms of their classification power (Lasso), the selected coefficients are contained in side bands, while typical strong Raman peaks are not present in the discrimination vector. We propose that this occurs because large Raman bands are representative of a wide variety of cellular molecules and are therefore less suited for accurate classification.

In vitro Na+, K+-ATPase (EC 3.6.1.3)-dependent respiration and protein synthesis in skeletal muscle of pigs fed at three dietary protein levels

1989 ◽  
Vol 61 (3) ◽  
pp. 453-465 ◽  
Author(s):  
O. Adeola ◽  
L. G. Young ◽  
B. W. Mcbride ◽  
R. O. Ball
Keyword(s):  
Protein Synthesis ◽  
Dietary Protein ◽  
Protein Concentration ◽  
Live Weight ◽  
Synthesis Rate ◽  
Sartorius Muscle ◽  
Close Link ◽  
Protein Levels ◽  
Daily Gain

1. Eighteen pigs were offered diets containing 130, 170 or 210 g protein/kg with three barrows and three gilts per diet from 20 to 60 kg live weight. Oxygen consumption, Na1, K1-ATPase (EC 3·6·1· 3)-dependent and -independent respiration and protein synthesis were measured in vitro in intercostal and sartorius muscle preparations from these pigs.2. Increasing dietary protein concentration increased (P < 0·01) daily gain and dissectible muscle in carcass.3. O2 consumption and Na+, K+-ATPase-dependent respiration of the intercostal and sartorius muscles increased linearly (P < 0·01) with increase in dietary protein concentration. The requirement for the support of the transport of Na+ and K+ across the cell membrane in these muscles, on average, accounted for 22–25% of the O2 consumption.4. Synthesis rate (mg/g per d) of protein in the sartorius muscle increased (P < 0·05) from 3·05 to 5·07 and increased (P < 0·1) from 2·57 to 4.06 in the intercostal muscle as dietary protein increased from 130 to 210 g/kg diet.5. Regression of Na+, K+-ATPase-dependent respiration against protein synthesis in each of intercostal and sartorius muscles showed a linear relation, an attestation of a close link between productive processes and auxiliary energy expenditure.

Influence of steroid hormones on the incorporation of amino acids in uterine and cervical tissue of pregnant women

1987 ◽  
Vol 115 (4) ◽  
pp. 537-543
Author(s):  
Ingrid Wiqvist ◽  
Anders Linde
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Pregnant Women ◽  
Tissue Concentration ◽  
Uterine Tissue ◽  
Protein Synthesis Inhibitors ◽  
Experimental Conditions ◽  
Pregnant Uterus ◽  
High Concentrations

Abstract. The influence of steroids on protein synthesis in cervical and uterine tissue obtained from early and term pregnant women was studied by measuring the incorporation of labelled amino acids into total protein. It was found that oestradiol-17β and progesterone significantly reduced the incorporation of [3H]proline. Androstenedione and cortisol had no significant effect on the incorporation of [3H]proline even at high concentrations. The protein synthesis inhibitors puromycin and cycloheximide blocked the incorporation of [3H]proline to 80–85%. However, there was no further reduction in the incorporation in the presence of oestradiol. Oestradiol was found to reduce the incorporation of [14C]glycine but not that of [3H]serine. The results indicate that oestradiol and progesterone reduce protein synthesis in human cervical and uterine tissue and that this reduction, at least partially, involves collagen synthesis. Oestradiol and progesterone were equipotent under in vitro experimental conditions. The tissue concentration of progesterone in the pregnant uterus is, however, much higher than that of oestradiol. It seems therefore probable that progesterone rather than oestradiol restricts unopposed synthesis of proteins, presumably mainly collagen.

scholarly journals MAP2 is differentially phosphorylated in schizophrenia, altering its function

2021 ◽  
Author(s):  
M. J. Grubisha ◽  
X. Sun ◽  
M. L. MacDonald ◽  
M. Garver ◽  
Z. Sun ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Disease Risk ◽  
Protein Translation ◽  
Synaptic Proteins ◽  
Synaptic Protein ◽  
Spine Density ◽  
Neuronal Structure ◽  
Protein Levels ◽  
And Function

AbstractSchizophrenia (Sz) is a highly polygenic disorder, with common, rare, and structural variants each contributing only a small fraction of overall disease risk. Thus, there is a need to identify downstream points of convergence that can be targeted with therapeutics. Reduction of microtubule-associated protein 2 (MAP2) immunoreactivity (MAP2-IR) is present in individuals with Sz, despite no change in MAP2 protein levels. MAP2 is phosphorylated downstream of multiple receptors and kinases identified as Sz risk genes, altering its immunoreactivity and function. Using an unbiased phosphoproteomics approach, we quantified 18 MAP2 phosphopeptides, 9 of which were significantly altered in Sz subjects. Network analysis grouped MAP2 phosphopeptides into three modules, each with a distinct relationship to dendritic spine loss, synaptic protein levels, and clinical function in Sz subjects. We then investigated the most hyperphosphorylated site in Sz, phosphoserine1782 (pS1782). Computational modeling predicted phosphorylation of S1782 reduces binding of MAP2 to microtubules, which was confirmed experimentally. We generated a transgenic mouse containing a phosphomimetic mutation at S1782 (S1782E) and found reductions in basilar dendritic length and complexity along with reduced spine density. Because only a limited number of MAP2 interacting proteins have been previously identified, we combined co-immunoprecipitation with mass spectrometry to characterize the MAP2 interactome in mouse brain. The MAP2 interactome was enriched for proteins involved in protein translation. These associations were shown to be functional as overexpression of wild type and phosphomimetic MAP2 reduced protein synthesis in vitro. Finally, we found that Sz subjects with low MAP2-IR had reductions in the levels of synaptic proteins relative to nonpsychiatric control (NPC) subjects and to Sz subjects with normal and MAP2-IR, and this same pattern was recapitulated in S1782E mice. These findings suggest a new conceptual framework for Sz—that a large proportion of individuals have a “MAP2opathy”—in which MAP function is altered by phosphorylation, leading to impairments of neuronal structure, synaptic protein synthesis, and function.

scholarly journals MAP2 is Differentially Phosphorylated in Schizophrenia, Altering its Function

2019 ◽  
Author(s):  
MJ Grubisha ◽  
X Sun ◽  
ML MacDonald ◽  
M Garver ◽  
Z Sun ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Disease Risk ◽  
Protein Translation ◽  
Synaptic Proteins ◽  
Synaptic Protein ◽  
Spine Density ◽  
Neuronal Structure ◽  
Protein Levels ◽  
And Function

AbstractSchizophrenia (Sz) is a highly polygenic disorder, with common, rare, and structural variants each contributing only a small fraction of overall disease risk. Thus, there is a need to identify downstream points of convergence that can be targeted with therapeutics. Reduction of Microtubule-associated Protein 2 (MAP2) immunoreactivity (MAP2-IR) is present in individuals with Sz, despite no change in MAP2 protein levels. MAP2 is phosphorylated downstream of multiple receptors and kinases identified as Sz risk genes, altering its immunoreactivity and function. Using an unbiased phosphoproteomics approach we quantified 18 MAP2 phosphopeptides, 9 of which were significantly altered in Sz subjects. Network analysis grouped MAP2 phosphopeptides into 3 modules, each with a distinct relationship to dendritic spine loss, synaptic protein levels, and clinical function in Sz subjects. We then investigated the most hyperphosphorylated site in Sz, phosphoserine1782 (pS1782). Computational modeling predicted phosphorylation of S1782 reduces binding of MAP2 to microtubules, which was confirmed experimentally. We generated a transgenic mouse containing a phosphomimetic mutation at S1782 (S1782E) and found reductions in basilar dendritic length and complexity along with reduced spine density. Because only a limited number of MAP2 interacting proteins have been previously identified, we combined co-immunoprecipitation with mass spectrometry to characterize the MAP2 interactome in mouse brain. The MAP2 interactome was enriched for proteins involved in protein translation. These associations were shown to be functional as overexpression of wildtype and phosphomimetic MAP2 reduced protein synthesis in vitro. Finally, we found that Sz subjects with low MAP2-IR had reductions in the levels of synaptic proteins relative to nonpsychiatric control (NPC) subjects and to Sz subjects with normal and MAP2-IR, and this same pattern was recapitulated in S1782E mice. These findings suggest a new conceptual framework for Sz - that a large proportion of individuals have a “MAP2opathy” - in which MAP function is altered by phosphorylation, leading to impairments of neuronal structure, synaptic protein synthesis, and function.

Independence of the androgen-induced mRNA synthesis on exogenous glucose in the ventral prostate of the rat

1982 ◽  
Vol 101 (3) ◽  
pp. 472-480
Author(s):  
Anja Kuosa
Keyword(s):  
Protein Synthesis ◽  
Ventral Prostate ◽  
Experimental Conditions ◽  
Hormonal Stimulation ◽  
Electrophoretic Patterns ◽  
Exogenous Glucose ◽  
Polysomal Mrna ◽  
Mrna Content ◽  
Hormonal Activation

Abstract. Poly (A)-containing RNA species were isolated from prostatic polysomes after administration of testosterone to castrated rats. After exposure of prostatic tissue to radiolabelled uridine in vitro for 1 h the radioactivity of polysomal poly (A)-RNA increased at 6–12 h after testosterone treatment, which coincides with the hormonal activation of protein synthesis under the same experimental conditions. The increased radioactivity of polysomal mRNA was not due to changes in the uptake and phosphorylation or radiolabelled uridine or in the endogenous content of uridine nucleotides representing thus an increased mRNA content. Electrophoretic patterns of double-labelled polysomal mRNA failed to show detectable amounts of androgenspecific species. The larger part of increase in mRNA content thus consists of mRNAs common to prostatic cells of both testosterone-treated and non-treated castrated animals. The hormonal stimulation of polysomal poly (A)-RNA accumulation was not dependent upon exogenous glucose which is in contrast to protein synthesis. However, glucose was necessary for the hormonal activation of the phosphorylation of [3H]uridine leading ultimately to increased labelling of all RNA species. It is concluded that increased accumulation of polysomal mRNA may be necessary, but not sufficient for the hormonal activation of protein synthesis.

Observations of Frozen-Hydrated Microtubules

1990 ◽  
Vol 48 (1) ◽  
pp. 504-505
Author(s):  
D. Chrétien ◽  
D. Job ◽  
R.H. Wade
Keyword(s):  
Fourier Transforms ◽  
Structural Complexity ◽  
Image Contrast ◽  
Phase Behaviour ◽  
Experimental Conditions ◽  
Thin Sections ◽  
Surface Lattice ◽  
Cilia And Flagella ◽  
Outstanding Question

Microtubules are filamentary structures found in the cytoplasm of eukaryotic cells, where, together with actin and intermediate filaments, they form the components of the cytoskeleton. They have many functions and show various levels of structural complexity as witnessed by the singlet, doublet and triplet structures involved in the architecture of centrioles, basal bodies, cilia and flagella. The accepted microtubule model consists of a 25 nm diameter hollow tube with a wall made up of 13 paraxial protofilaments (pf). Each pf is a string of aligned tubulin dimers. Some results have suggested that the pfs follow a superhelix. To understand how microtubules function in the cell an accurate model of the surface lattice is one of the requirements. For example the 9x2 architecture of the axoneme will depend on the organisation of its component microtubules. We should also note that microtubules with different numbers of pfs have been observed in thin sections of cellular and of in-vitro material. An outstanding question is how does the surface lattice adjust to these different pf numbers?We have been using cryo-electron microscopy of frozen-hydrated samples to study in-vitro assembled microtubules. The experimental conditions are described in detail in this reference. The results obtained in conjunction with thin sections of similar specimens and with axoneme outer doublet fragments have already allowed us to characterise the image contrast of 13, 14 and 15 pf microtubules on the basis of the measured image widths, of the the image contrast symmetry and of the amplitude and phase behaviour along the equator in the computed Fourier transforms. The contrast variations along individual microtubule images can be interpreted in terms of the geometry of the microtubule surface lattice. We can extend these results and make some reasonable predictions about the probable surface lattices in the case of other pf numbers, see Table 1. Figure 1 shows observed images with which these predictions can be compared.

Increased Protein Synthesis by Human Platelets during Phagocytosis of Latex Particles in Vitro

1976 ◽  
Vol 35 (02) ◽  
pp. 350-357 ◽  
Author(s):  
Hana Bessler ◽  
Galila Agam ◽  
Meir Djaldetti
Keyword(s):  
Protein Synthesis ◽  
Fold Increase ◽  
Human Platelets ◽  
Polystyrene Latex ◽  
Latex Particles ◽  
The Third ◽  
Platelet Protein ◽  
Dose Dependent ◽  
Phagocytotic Activity

SummaryA three-fold increase of protein synthesis by human platelets during in vitro phagocytosis of polystyrene latex particles was detected. During the first two hours of incubation, the percentage of phagocytizing platelets and the number of latex particles per platelet increased; by the end of the third hour, the first parameter remained stable, while the number of latex particles per cell had decreased.Vincristine (20 μg/ml of cell suspension) inhibited platelet protein synthesis. This effect was both time- and dose-dependent. The drug also caused a decrease in the number of phagocytizing cells, as well as in their phagocytotic activity.

Superoxide, Xanthine Oxidase and Platelet Reactions: Further Studies on Mechanisms by which Oxidants Influence Platelets

1981 ◽  
Vol 45 (03) ◽  
pp. 290-293 ◽  
Author(s):  
Peter H Levine ◽  
Danielle G Sladdin ◽  
Norman I Krinsky
Keyword(s):  
Superoxide Dismutase ◽  
Cytochrome C ◽  
Xanthine Oxidase ◽  
Direct Effect ◽  
Platelet Function ◽  
Experimental Conditions ◽  
Release Reaction

SummaryIn the course of studying the effects on platelets of the oxidant species superoxide (O- 2), Of was generated by the interaction of xanthine oxidase plus xanthine. Surprisingly, gel-filtered platelets, when exposed to xanthine oxidase in the absence of xanthine substrate, were found to generate superoxide (O- 2), as determined by the reduction of added cytochrome c and by the inhibition of this reduction in the presence of superoxide dismutase.In addition to generating Of, the xanthine oxidase-treated platelets display both aggregation and evidence of the release reaction. This xanthine oxidase induced aggreagtion is not inhibited by the addition of either superoxide dismutase or cytochrome c, suggesting that it is due to either a further metabolite of O- 2, or that O- 2 itself exerts no important direct effect on platelet function under these experimental conditions. The ability of Of to modulate platelet reactions in vivo or in vitro remains in doubt, and xanthine oxidase is an unsuitable source of O- 2 in platelet studies because of its own effects on platelets.

Desmopressin (DDAVP) Enhances Platelet Adhesion to the Extracellular Matrix of Cultured Human Endothelial Cells through Increased Expression of Tissue Factor

1997 ◽  
Vol 77 (05) ◽  
pp. 0975-0980 ◽  
Author(s):  
Angel Gálvez ◽  
Goretti Gómez-Ortiz ◽  
Maribel Díaz-Ricart ◽  
Ginés Escolar ◽  
Rogelio González-Sarmiento ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Endothelial Cells ◽  
Tissue Factor ◽  
Platelet Adhesion ◽  
Umbilical Vein ◽  
Procoagulant Activity ◽  
Experimental Conditions ◽  
Chromogenic Assay

SummaryThe effect of desmopressin (DDAVP) on thrombogenicity, expression of tissue factor and procoagulant activity (PCA) of extracellular matrix (ECM) generated by human umbilical vein endothelial cells cultures (HUVEC), was studied under different experimental conditions. HUVEC were incubated with DDAVP (1, 5 and 30 ng/ml) and then detached from their ECM. The reactivity towards platelets of this ECM was tested in a perfusion system. Coverslips covered with DD A VP-treated ECMs were inserted in a parallel-plate chamber and exposed to normal blood anticoagulated with low molecular weight heparin (Fragmin®, 20 U/ml). Perfusions were run for 5 min at a shear rate of 800 s1. Deposition of platelets on ECMs was significantly increased with respect to control ECMs when DDAVP was used at 5 and 30 ng/ml (p <0.05 and p <0.01 respectively). The increase in platelet deposition was prevented by incubation of ECMs with an antibody against human tissue factor prior to perfusion. Immunofluorescence studies positively detected tissue factor antigen on DDAVP derived ECMs. A chromogenic assay performed under standardized conditions revealed a statistically significant increase in the procoagulant activity of the ECMs produced by ECs incubated with 30 ng/ml DDAVP (p <0.01 vs. control samples). Northern blot analysis revealed increased levels of tissue factor mRNA in extracts from ECs exposed to DDAVP. Our data indicate that DDAVP in vitro enhances platelet adhesion to the ECMs through increased expression of tissue factor. A similar increase in the expression of tissue factor might contribute to the in vivo hemostatic effect of DDAVP.

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