scholarly journals Intragenic recombination in an E beta gene for a murine Ia antigen.

1982 ◽  
Vol 155 (3) ◽  
pp. 937-942 ◽  
Author(s):  
M L Plunkett ◽  
J E Coligan ◽  
C S David ◽  
J H Freed
Keyword(s):  
Plasma Membrane ◽  
Recombinant Strain ◽  
Intragenic Recombination ◽  
Two Dimensional ◽  
Beta Chain ◽  
Terminal Portion ◽  
Recombinational Event ◽  
Amino Terminal ◽  
Normal Behavior ◽  
Beta Gene

The recombinant strain D2.GD was originally typed as I-Ad by serological methods. Indeed, the A alpha and A beta chains of the I-A antigens appear to exhibit normal behavior by the criteria of serology and two dimensional gel analysis. However, the E beta chain encoded by the I-A subregion of this strain, one of the two components of the plasma membrane located I-E antigens produced in D2.GD X A.TFR5)F1 animals, has been demonstrated to be the product of an intragenic recombinational event between E beta genes from the d and b haplotypes. Sequence analysis suggests that the amino-terminal portion of the Eg2 beta chain is derived from the d haplotype and, therefore, that the coding strand for this gene is oriented centromeric leads to telomeric (5' to 3' direction). Finally, these data combined with the data of Rose and Cullen (17) allow the ordering of the genes within the I-A subregion as (H-2K), A alpha, A beta, E beta ... (H-2D).

The extracellular matrix of echinoderm and amphibian eggs: Visualization in quick-frozen, deep-etched, and rotary-shadowed specimens

1990 ◽  
Vol 48 (3) ◽  
pp. 60-61
Author(s):  
Barry Bonnell ◽  
Carolyn Larabell ◽  
Douglas Chandler
Keyword(s):  
Extracellular Matrix ◽  
Plasma Membrane ◽  
Sea Urchin ◽  
Crystalline Structure ◽  
Cortical Granule ◽  
Two Dimensional ◽  
Small Peptides ◽  
Deep Etching ◽  
Quick Freezing ◽  
Acrosomal Reaction

Eggs of many species including those of echinoderms, amphibians and mammals exhibit an extensive extracellular matrix (ECM) that is important both in the reception of sperm and in providing a block to polyspermy after fertilization.In sea urchin eggs there are two distinctive coats, the vitelline layer which contains glycoprotein sperm receptors and the jelly layer that contains fucose sulfate glycoconjugates which trigger the acrosomal reaction and small peptides which act as chemoattractants for sperm. The vitelline layer (VL), as visualized by quick-freezing, deep-etching, and rotary-shadowing (QFDE-RS), is a fishnet-like structure, anchored to the plasma membrane by short posts. Orbiting above the VL are horizontal filaments which are thought to anchor the thicker jelly layer to the egg. Upon fertilization, the VL elevates and is transformed by cortical granule secretions into the fertilization envelope (FE). The rounded casts of microvilli in the VL are transformed into angular peaks and the envelope becomes coated inside and out with sheets of paracrystalline protein having a quasi-two dimensional crystalline structure.

Parathyroid hormone metabolism and its potential as a uremic toxin

1980 ◽  
Vol 239 (1) ◽  
pp. F1-F12 ◽  
Author(s):  
E. Slatopolsky ◽  
K. Martin ◽  
K. Hruska
Keyword(s):  
Renal Failure ◽  
Parathyroid Hormone ◽  
Chronic Renal Failure ◽  
Biological Effects ◽  
Uremic Toxin ◽  
Terminal Portion ◽  
Single Chain ◽  
Biochemical Alterations ◽  
Amino Terminal ◽  
Carboxy Terminal

Secondary hyperparathyroidism is a universal complication of chronic renal failure. It has been proposed that the markedly elevated levels of immunoreactive parathyroid hormone (i-PTH) in uremia may represent a “uremic toxin” responsible for many of the abnormalities of the uremic state. Plasma i-PTH consists of a mixture of intact hormone, a single-chain polypeptide of 84 amino acids, and smaller molecular weight hormonal fragments from both the carboxy- and amino-terminal portion of the PTH molecule. The hormonal fragments arise from metabolism of intact PTH by peripheral organs as well as from secretion of fragments from the parathyroid glands. The structural requirements for the known biological actions of PTH reside in the amino-terminal portion of the PTH molecule. Carboxy-terminal fragments, biologically inactive at least in terms of adenylate cyclase activation, hypercalcemia, or phosphaturia, depend on the kidney for their removal from plasma, and thus accumulate in the circulation in chronic renal failure. It is unknown at the present time if other biological effects of these carboxy-terminal fragments may contribute to some of the biochemical alterations observed in uremia. The most significant consequence of increased PTH levels in uremia is the development of bone disease characterized by osteitis fibrosa. In addition, it would appear that PTH plays an important role in some of the abnormal electroencephalographic patterns observed in uremia. This may be due to a potential role of PTH in increasing calcium content of brain. Parathyroid hormone also has been implicated as a pathogenetic factor in many other alterations present in uremia, i.e., peripheral neuropathy, carbohydrate intolerance, hyperlipidemia, and other alterations. Unfortunately, outstanding clinical research is lacking in this field and conclusive experimental data are practically nonexistent. Further studies are necessary if one is to accept the concept of PTH being a significant “uremic toxin.”

Oocyte proteomics: localisation of mouse zona pellucida protein 3 to the plasma membrane of ovulated mouse eggs

2004 ◽  
Vol 16 (2) ◽  
pp. 69 ◽  
Author(s):  
S. A. Coonrod ◽  
M. E. Calvert ◽  
P. P. Reddi ◽  
E. N. Kasper ◽  
L. C. Digilio ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Indirect Immunofluorescence ◽  
Zona Pellucida ◽  
Surface Expression ◽  
Surface Proteins ◽  
2D Electrophoresis ◽  
Dependent Manner ◽  
Two Dimensional ◽  
Phase Partitioning ◽  
Proteomic Approach

In order to gain a deeper understanding of the molecular underpinnings of sperm–egg interaction and early development, we have used two-dimensional (2D) electrophoresis, avidin blotting and tandem mass spectrometry to identify, clone and characterise abundant molecules from the mouse egg proteome. Two-dimensional avidin blots of biotinylated zona-free eggs revealed an abundant approximately 75-kDa surface-labelled heterogeneous protein possessing a staining pattern similar to that of the zona pellucida glycoprotein, mouse ZP3 (mZP3). In light of this observation, we investigated whether mZP3 specifically localises to the plasma membrane of mature eggs. Zona pellucidae of immature mouse oocytes and mature eggs were removed using acid Tyrode’s solution, chymotrypsin or mechanical shearing. Indirect immunofluorescence using the mZP3 monoclonal antibody (mAb) IE-10 demonstrated strong continuous staining over the entire surface of immature oocytes and weak microvillar staining on ovulated eggs, regardless of the method of zona removal. Interestingly, in mature eggs, increased fluorescence intensity was observed following artificial activation and fertilisation, whereas little to no fluorescence was observed in degenerated eggs. The surface localisation of ZP3 on mature eggs was supported by the finding that the IE-10 mAb immunoprecipitated an approximate 75-kDa protein from lysates of biotinylated zona-free eggs. To further investigate the specificity of the localisation of mZP3 to the oolemma, indirect immunofluorescence was performed using the IE-10 mAb on both CV-1 and CHO cells transfected with full-length recombinant mZP3 (re-mZP3). Plasma membrane targeting of the expressed re-mZP3 protein was observed in both cell lines. The membrane association of re-mZP3 was confirmed by the finding that biotinylated re-mZP3 (approximately 75 kDa) is immunoprecipitated from the hydrophobic phase of Triton X-114 extracts of transfected cells following phase partitioning. Immunoprecipitation assays also demonstrated that surface re-mZP3 was released from transfected CV-1 in a time-dependent manner. These results demonstrate that ZP3 is specifically associated with the surface of mature eggs and its subsequent release from the cell surface may represent one mechanism by which ZP3 is secreted. Furthermore, the increase in ZP3 surface expression following fertilisation suggests that ZP3 may have a functional role during sperm–oolemma binding and fusion. These results also validate the usefulness of using the 2D proteomic approach to identify and characterise egg-surface proteins.

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