scholarly journals Synthesis and glycosylation of CD52, the major 'maturation-associated' antigen of rat spermatozoa, in the cauda epididymidis

Reproduction ◽  
2001 ◽  
pp. 435-446 ◽  
Author(s):  
P Derr ◽  
CH Yeung ◽  
TG Cooper ◽  
C Kirchhoff
Keyword(s):  
Blot Analysis ◽  
Cdna Sequence ◽  
Northern Blot Analysis ◽  
Lectin Binding ◽  
Transcriptional Level ◽  
Lectin Blot ◽  
Molecular Masses ◽  
Rat Epididymis ◽  
Sperm Membrane ◽  
Cauda Epididymidis

A western and lectin blot analysis was performed of the major 'maturation-associated' antigen of rat spermatozoa, which is the rat counterpart of human CD52. In the absence of a suitable antibody, direct study of this approximately 26 kDa antigen, named previously SMemG, had been difficult. In the present study, these problems were overcome by raising a polyclonal antibody against a chemosynthetic peptide predicted from the cDNA sequence of the antigen. The antibody bound to a glycoprotein of rat cauda epididymidal tissue and spermatozoa, this glycoprotein was cleaved by phosphatidylinositol-specific phospholipase C and, after deglycosylation, was reduced to approximately 6 kDa. Northern blot analysis confirmed that the CD52 mRNA was transcribed only post-testicularly, and antibody binding to testicular and sperm proteins of different molecular masses was shown to be nonspecific. Flow cytometry also indicated that the antigen was inserted into the sperm membrane during epididymal transit. Moreover, despite the presence of CD52 mRNA in all parts of the rat epididymis, only the 'long' mRNA molecules of the cauda region were efficiently translated and the antigen glycosylated, indicating that expression of rat CD52 is regulated on a post-transcriptional level. Lectin binding and deglycosylation studies supported the contention that there is extensive mucin-type O-glycosylation of rat CD52. In rats, there was no indication of complex N-linked carbohydrates similar to those described for human CD52.

scholarly journals Glycosylation Profile of the Transferrin Receptor in Gestational Iron Deficiency and Early-Onset Severe Preeclampsia

2019 ◽  
Vol 2019 ◽  
pp. 1-9 ◽  
Author(s):  
Alejandra María Gómez-Gutiérrez ◽  
Beatriz Elena Parra-Sosa ◽  
Julio Cesar Bueno-Sánchez
Keyword(s):  
Iron Deficiency ◽  
Western Blot ◽  
Blot Analysis ◽  
Transferrin Receptor ◽  
Lectin Binding ◽  
Hypoxia Inducible Factor ◽  
Severe Preeclampsia ◽  
Hypoxia Inducible Factor 1 ◽  
Lectin Blot ◽  
Low Expression

Objective. To examine the expression of hypoxia-inducible factor-1α(HIF-1α), TfR1, and TfR1-attached terminal monosaccharides in placentas of women with IDAP and severe preeclampsia.Methods. TfR1 and HIF-1αwere detected by western blot. Immunoadsorption of TfR1 was performed to characterize the terminal monosaccharides by specific lectin binding.Results. There was no difference in the expression of TfR1 and HIF-1αbetween groups. Lectin blot analysis pointed out an overexpression of galactoseβ1-4N-acetylglucosamine (Gal-GlcNAc) and mannose in severe preeclampsia.Conclusion. The increase in Gal-GlcNAc may be due to the increased presence of antennary structures and the mannose glycans of TfR1 may indicate the presence of misfolded or incomplete proteins. These findings may be associated with the low expression of placental TfR1 in women with preeclampsia.

Control of polygalacturonase synthesis inFusarium oxyspotumf.sp.radicis lycopersici

1997 ◽  
Vol 43 (11) ◽  
pp. 1084-1090 ◽  
Author(s):  
Belén Patiño ◽  
Martha Lucía Posada ◽  
Covadonga Vázquez ◽  
María Teresa González-Jaén ◽  
Álvaro Martínez del Pozo
Keyword(s):  
Fusarium Oxysporum ◽  
Blot Analysis ◽  
Genomic Dna ◽  
Southern Blot Analysis ◽  
Northern Blot Analysis ◽  
Single Copy ◽  
Transcriptional Level ◽  
High Similarity ◽  
Apple Pectin ◽  
Sds Page

Genetic control of polygalacturonase (PG) activity from Fusarium oxysporum f.sp. radicis lycopersici was analyzed on pectin and glucose cultures. One exopolygalacturonase from F. oxysporum f.sp. radicis lycopersici was strongly induced, in stationary culture, when the fungus was grown on apple pectin, while on glucose no extracellular PG activity could be detected. Although SDS–PAGE detected the presence of a putative PG band (66 kDa) in both conditions, specific antibodies obtained against the purified PG only detected it in PG-inducing conditions, that is to say, when apple pectin was used as the carbon source. Northern blot analysis of RNA of two isolates of F. oxysporum f.sp. radicis lycopersici (r6and r2) confirmed that this regulation of PG synthesis was exerted at the transcriptional level. Only one single mRNA species of around 1400 nucleotides was detected on the cultures containing pectin and was absent in glucose-grown cultures. Southern blot analysis of genomic DNA indicated that pg gene seems to be present in a single copy in the genomes of F. oxysporum f.sp. radicis lycopersici r6and r2and Fusarium oxysporum f.sp. lycopersici, showing similar hybridization patterns in all species. The partial sequence of this pg gene from F. oxysporum f.sp. radicis lycopersici r6, which is also reported, showed high similarity to diverse PGs already reported. Exopolygalacturonase of F. oxysporum f.sp. radicis lycopersici r6is heavily glycosylated; its deglycosylated form had a molecular mass of 50 kDa.Key words: polygalacturonase, Fusarium oxysporum f.sp. radicis lycopersici, regulation.

scholarly journals Emergence of Fosfomycin-Resistant Isolates of Shiga-Like Toxin-Producing Escherichia coli O26

1999 ◽  
Vol 43 (4) ◽  
pp. 789-793 ◽  
Author(s):  
Toshinobu Horii ◽  
Taku Kimura ◽  
Kumiko Sato ◽  
Keigo Shibayama ◽  
Michio Ohta
Keyword(s):  
Escherichia Coli ◽  
Northern Blot ◽  
Blot Analysis ◽  
Northern Blot Analysis ◽  
Transport Systems ◽  
Transcriptional Level ◽  
Gene Encoding ◽  
E Coli ◽  
Fosfomycin Resistance

ABSTRACT We evaluated the susceptibilities of 129 Shiga-like toxin-producingEscherichia coli (STEC) isolates to various antibiotics. The numbers of isolates for which MICs were high (≧128 μg/ml) were as follows: 5 for fosfomycin, 14 for ampicillin, 1 for cefaclor, 6 for kanamycin, 22 for tetracycline, and 2 for doxycycline. For two isolates of STEC O26 MICs of fosfomycin were high (1,024 and 512 μg/ml, respectively). Conjugation experiments and glutathioneS-transferase assays suggested that the fosfomycin resistance in these isolates was determined not by a plasmid but chromosomally. The amount of active intracellular fosfomycin in these STEC isolates was 100- to 200-fold less than that in E. coli C600 harboring pREFTT47B408 in the presence of eitherl-α-glycerophosphate or glucose-6-phosphate. Cloning, sequencing, and Northern blot analysis demonstrated that the transcriptional level of the murA gene encoding UDP-N-acetylglucosamine enolpyruvoyl transferase in these isolates was greater than that in E. coli C600. Our results suggest that the fosfomycin resistance in these STEC isolates is due to concurrent effects of alteration of the glpT and/oruhp transport systems and of the enhanced transcription of the murA gene.

scholarly journals Controlled-atmosphere Suppression of ACC Synthase and ACC Oxidase in `Golden Delicious' Apples during Long-term Cold Storage

1996 ◽  
Vol 121 (4) ◽  
pp. 751-755 ◽  
Author(s):  
James R. Gorny ◽  
Adel A. Kader
Keyword(s):  
Blot Analysis ◽  
Northern Blot Analysis ◽  
Acc Oxidase ◽  
Acc Synthase ◽  
Transcript Abundance ◽  
Ethylene Biosynthesis ◽  
Transcriptional Level ◽  
Golden Delicious

Preclimacteric `Golden Delicious' apples (Malus domestica Borkh.) were stored at 0 °C in: air; air + 5% CO2; 2% O2 + 98% N2; or 2% O2 + 5% CO2 + 93% N2, and sampled monthly for 4 months to investigate the mechanism(s) by which reduced O2 and/or elevated CO2 atmospheres inhibit C2H4 biosynthesis. Ethylene biosynthesis rates and in vitro ACS activity were closely correlated in all treatments, while in vitro ACO activity significantly increased over time regardless of the treatment. Only a small amount of C2H4 biosynthesis inhibition by lowered O2 and/or elevated CO2 atmospheres could be accounted for by suppressed induction of ACO activity. Western blot analysis demonstrated that apples held for 2 months in lowered O2 and/or elevated CO2 atmospheres had significantly reduced abundance of ACO protein, compared to fruit held in air. Northern blot analysis of ACS and ACO transcript abundance revealed that reduced O2 and/or elevated CO2 atmospheres delay induction and reduce the abundance of both transcripts. Reduced O2 and/or elevated CO2 atmospheres reduce C2H4 biosynthesis by delaying and suppressing expression of ACS at the transcriptional level and by reducing the abundance of active ACO protein. Chemical names used: 1-aminocyclopropane-1-carboxylic acid (ACC), ACC synthase (ACS), ACC oxidase (ACO), ethylene (C2H4), S-adenosylmethionine (AdoMet).

scholarly journals IL-8 and MCP Gene Expression and Production by LPS-Stimulated Human Corneal Stromal Cells

2012 ◽  
Vol 2012 ◽  
pp. 1-4 ◽  
Author(s):  
Roni M. Shtein ◽  
Susan G. Elner ◽  
Zong-Mei Bian ◽  
Victor M. Elner
Keyword(s):  
Gene Expression ◽  
Northern Blot ◽  
Stromal Cells ◽  
Blot Analysis ◽  
Time Course ◽  
Northern Blot Analysis ◽  
Statistical Significance ◽  
Dependent Manner ◽  
Corneal Inflammation ◽  
Chemotactic Protein

Purpose. To determine time course of effect of lipopolysaccharide (LPS) on production of interleukin-8 (IL-8) and monocyte chemotactic protein (MCP) by cultured human corneal stromal cells.Methods. Human corneal stromal cells were harvested from donor corneal specimens, and fourth to sixth passaged cells were used. Cell cultures were stimulated with LPS for 2, 4, 8, and 24 hours. Northern blot analysis of IL-8 and MCP gene expression and ELISA for IL-8 and MCP secretion were performed. ELISA results were analyzed for statistical significance using two-tailed Student'st-test.Results. Northern blot analysis demonstrated significantly increased IL-8 and MCP gene expression after 4 and 8 hours of exposure to LPS. ELISA for secreted IL-8 and MCP demonstrated statistically significant increases (P<0.05) after corneal stromal cell stimulation with LPS.Conclusions. This paper suggests that human corneal stromal cells may participate in corneal inflammation by secreting potent leukocyte chemotactic and activating proteins in a time-dependent manner when exposed to LPS.

scholarly journals A rapid and accurate method for quantitating total RNA transferred during Northern blot analysis

1988 ◽  
Vol 16 (5) ◽  
pp. 2354-2354 ◽  
Author(s):  
Nathalie Denis ◽  
Daniel Corcos ◽  
Jacques Kruh ◽  
Alain Kitzis
Keyword(s):  
Northern Blot ◽  
Blot Analysis ◽  
Northern Blot Analysis ◽  
Accurate Method ◽  
Total Rna

The mouse prostaglandin E receptor EP2 subtype: cloning, expression, and Northern blot analysis

FEBS Letters ◽  
1995 ◽  
Vol 372 (2-3) ◽  
pp. 151-156 ◽  
Author(s):  
Masato Katsuyama ◽  
Nobuhiro Nishigaki ◽  
Yukihiko Sugimoto ◽  
Kimiko Morimoto ◽  
Manabu Negishi ◽  
...  
Keyword(s):  
Northern Blot ◽  
Blot Analysis ◽  
Northern Blot Analysis ◽  
Prostaglandin E
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