Role of Nudel protease activation in triggering dorsoventral polarization of the Drosophila embryo

Development ◽  
1998 ◽  
Vol 125 (20) ◽  
pp. 4045-4053 ◽  
Author(s):  
E.K. LeMosy ◽  
D. Kemler ◽  
C. Hashimoto
Keyword(s):  
Serine Protease ◽  
Ovarian Follicle ◽  
Positional Information ◽  
Ventral Side ◽  
Drosophila Embryo ◽  
Follicle Cells ◽  
Signaling Mechanism ◽  
Dorsoventral Polarity ◽  
Serine Protease Domain ◽  
Protease Activation

The establishment of embryonic dorsoventral polarity in Drosophila depends on a signaling mechanism in which the signal for ventral development is locally produced. This mechanism requires the activity of the nudel gene in ovarian follicle cells, which provide dorsoventral positional information for the embryo. The nudel gene product, a large mosaic protein with a central serine protease domain, has been proposed to function in locally triggering a protease cascade that produces the ventral signal. Here we provide evidence that the serine protease activity of the Nudel protein is essential for embryonic dorsoventral polarity and that the active Nudel protease is generated by autoproteolytic cleavage of a zymogen form. Activation of the Nudel protease is independent of the other known proteases involved in dorsoventral polarity establishment and appears to occur symmetrically on the surface of the embryo. Our findings suggest that Nudel protease activation initiates the protease cascade that produces the ventral signal, but that spatial regulation occurring downstream of Nudel protease activation localizes the cascade to the ventral side of the embryo.

Biochemical Defects of Mutant nudel Alleles Causing Early Developmental Arrest or Dorsalization of the Drosophila Embryo

Genetics ◽  
2000 ◽  
Vol 154 (1) ◽  
pp. 247-257
Author(s):  
Ellen K LeMosy ◽  
Cynthia L Leclerc ◽  
Carl Hashimoto
Keyword(s):  
Serine Proteases ◽  
Structural Integrity ◽  
Ovarian Follicle ◽  
Drosophila Embryo ◽  
Follicle Cells ◽  
Dorsoventral Patterning ◽  
Protease Domain ◽  
Serine Protease Domain ◽  
Embryonic Arrest ◽  
Processing Stability

Abstract The nudel gene of Drosophila is maternally required both for structural integrity of the egg and for dorsoventral patterning of the embryo. It encodes a structurally modular protein that is secreted by ovarian follicle cells. Genetic and molecular studies have suggested that the Nudel protein is also functionally modular, with a serine protease domain that is specifically required for ventral development. Here we describe biochemical and immunolocalization studies that provide insight into the molecular basis for the distinct phenotypes produced by nudel mutations and for the interactions between these alleles. Mutations causing loss of embryonic dorsoventral polarity result in a failure to activate the protease domain of Nudel. Our analyses support previous findings that catalytic activity of the protease domain is required for dorsoventral patterning and that the Nudel protease is auto-activated and reveal an important role for a region adjacent to the protease domain in Nudel protease function. Mutations causing egg fragility and early embryonic arrest result in a significant decrease in extracellular Nudel protein, due to defects in post-translational processing, stability, or secretion. On the basis of these and other studies of serine proteases, we suggest potential mechanisms for the complementary and antagonistic interactions between the nudel alleles.

Genes that control dorsoventral polarity affect gene expression along the anteroposterior axis of the Drosophila embryo

Development ◽  
1987 ◽  
Vol 99 (3) ◽  
pp. 327-332 ◽  
Author(s):  
S.B. Carroll ◽  
G.M. Winslow ◽  
V.J. Twombly ◽  
M.P. Scott
Keyword(s):  
Gene Expression ◽  
Cell Fate ◽  
Maternal Effect ◽  
Positional Information ◽  
Drosophila Embryo ◽  
Dorsoventral Polarity ◽  
Anteroposterior Axis ◽  
Positional Marker ◽  
Control Pattern ◽  
The Relationship

At least 13 genes control the establishment of dorsoventral polarity in the Drosophila embryo and more than 30 genes control the anteroposterior pattern of body segments. Each group of genes is thought to control pattern formation along one body axis, independently of the other group. We have used the expression of the fushi tarazu (ftz) segmentation gene as a positional marker to investigate the relationship between the dorsoventral and anteroposterior axes. The ftz gene is normally expressed in seven transverse stripes. Changes in the striped pattern in embryos mutant for other genes (or progeny of females homozygous for maternal-effect mutations) can reveal alterations of cell fate resulting from such mutations. We show that in the absence of any of ten maternal-effect dorsoventral polarity gene functions, the characteristic stripes of ftz protein are altered. Normally there is a difference between ftz stripe spacing on the dorsal and ventral sides of the embryo; in dorsalized mutant embryos the ftz stripes appear to be altered so that dorsal-type spacing occurs on all sides of the embryo. These results indicate that cells respond to dorsoventral positional information in establishing early patterns of gene expression along the anteroposterior axis and that there may be more significant interactions between the different axes of positional information than previously determined.

Windbeutel is required for function and correct subcellular localization of the Drosophila patterning protein Pipe

Development ◽  
2000 ◽  
Vol 127 (24) ◽  
pp. 5541-5550 ◽  
Author(s):  
J. Sen ◽  
J.S. Goltz ◽  
M. Konsolaki ◽  
T. Schupbach ◽  
D. Stein
Keyword(s):  
Subcellular Localization ◽  
Ovarian Follicle ◽  
Growth Factor Receptor ◽  
Signal Transduction Pathway ◽  
Follicle Cell ◽  
Ventral Side ◽  
Follicle Cells ◽  
Culture Cells ◽  
Proteolytic Cascade ◽  
Pattern Forming

Drosophila embryonic dorsal-ventral polarity originates in the ovarian follicle through the restriction of pipe gene expression to a ventral subpopulation of follicle cells. Pipe, a homolog of vertebrate glycosaminoglycan-modifying enzymes, directs the ventral activation of an extracellular serine proteolytic cascade which defines the ventral side of the embryo. When pipe is expressed uniformly in the follicle cell layer, a strong ventralization of the resulting embryos is observed. Here, we show that this ventralization is dependent on the other members of the dorsal group of genes controlling dorsal-ventral polarity, but not on the state of the Epidermal Growth Factor Receptor signal transduction pathway which defines egg chamber polarity. Pipe protein expressed in vertebrate tissue culture cells localizes to the endoplasmic reticulum. Strikingly, coexpression of the dorsal group gene windbeutel in those cells directs Pipe to the Golgi. Similarly, Pipe protein exhibits an altered subcellular localization in the follicle cells of females mutant for windbeutel. Thus, Windbeutel protein enables the correct subcellular distribution of Pipe to facilitate its pattern-forming activity.

An Enzyme-Linked Immunosorbent Assay for Urokinase-Type Plasminogen Activator (u-PA) and Mutants and Chimeras Containing the Serine Protease Domain of u-PA

1992 ◽  
Vol 67 (01) ◽  
pp. 095-100 ◽  
Author(s):  
Paul J Declerck ◽  
Leen Van Keer ◽  
Maria Verstreken ◽  
Désiré Collen
Keyword(s):  
Serine Protease ◽  
Plasminogen Activator ◽  
Active Site ◽  
Enzyme Linked Immunosorbent Assay ◽  
Single Chain ◽  
Protease Domain ◽  
Serine Protease Domain ◽  
Type Plasminogen Activator ◽  
Urokinase Type Plasminogen Activator ◽  
Normal Human

SummaryAn enzyme-linked immunosorbent assay (ELISA) for quantitation of natural and recombinant plasminogen activators containing the serine protease domain (B-chain) of urokinase-type plasminogen activator (u-PA) was developed, based on two murine monoclonal antibodies, MA-4D1E8 and MA-2L3, raised against u-PA and reacting with non-overlapping epitopes in the B-chain. MA-4D1E8 was coated on microtiter plates and bound antigen was quantitated with MA-2L3 conjugated with horseradish peroxidase. The intra-assay, inter-assay and inter-dilution coefficients of variation of the assay were 6%, 15% and 9%, respectively. Using recombinant single-chain u-PA (rscu-PA) as a standard, the u-PA-related antigen level in normal human plasma was 1.4 ± 0.6 ng/ml (mean ± SD, n = 27).The ELISA recognized the following compounds with comparable sensitivity: intact scu-PA (amino acids, AA, 1 to 411), scu-PA-32k (AA 144 to 411), a truncated (thrombin-derived) scu-PA comprising A A 157 to 411, and chimeric t-PA/u-PA molecules including t-PA(AA1-263)/scu-PA(AA144-411), t-PA(AA1-274)/scu-PA(AA138-411) and t-PA(AA87-274)/scu-PA(AA138-411). Conversion of single-chain to two-chain forms of u-PA or inhibition of active two-chain forms with plasminogen activator inhibitor-1 or with the active site serine inhibitor phenyl-methyl-sulfonyl fluoride, did not alter the reactivity in the assay. In contrast, inactivation with α2-antiplasmin or with the active site histidine inhibitor Glu-Gly-Arg-CH2Cl resulted in a 3- to 5-fold reduction of the reactivity. When purified scu-PA-32k was added to pooled normal human plasma at final concentrations ranging from 20 to 1,000 ng/ml, recoveries in the ELISA were between 84 and 110%.The assay was successfully applied for the quantitation of pharmacological levels of scu-PA and t-PA(AA87_274)/scu-PA(AA138-411) in plasma during experimental thrombolysis in baboons.Thus the present ELISA, which is specifically dependent on the presence of the serine protease part of u-PA, is useful for measurement of a wide variety of variants and chimeras of u-PA which are presently being developed for improved thrombolytic therapy.

scholarly journals Graft-versus-host disease targets ovary and causes female infertility in mice

Blood ◽  
2017 ◽  
Vol 129 (9) ◽  
pp. 1216-1225 ◽  
Author(s):  
Sonoko Shimoji ◽  
Daigo Hashimoto ◽  
Hidetsugu Tsujigiwa ◽  
Kohta Miyawaki ◽  
Koji Kato ◽  
...  
Keyword(s):  
T Cell ◽  
Graft Versus Host Disease ◽  
Ovarian Follicle ◽  
Follicle Cells ◽  
Cell Infiltration ◽  
Ovarian Insufficiency ◽  
Graft Versus Host ◽  
T Cell Infiltration ◽  
Gvhd Prophylaxis ◽  
Key Points

Key Points GVHD mediates donor T-cell infiltration and apoptosis of the ovarian follicle cells, leading to ovarian insufficiency and infertility. Ovarian insufficiency and infertility are independent of conditioning, and pharmacologic GVHD prophylaxis preserves fertility.

Interaction of the A1 Subunit of Factor VIIIa and the Serine Protease Domain of Factor X Identified by Zero-length Cross-linking

1998 ◽  
Vol 80 (09) ◽  
pp. 418-422 ◽  
Author(s):  
Kirsty Lapan ◽  
Philip Fay
Keyword(s):  
Serine Protease ◽  
Heavy Chain ◽  
Solid Phase ◽  
Activated Protein C ◽  
Cross Linking ◽  
Factor X ◽  
Serine Protease Domain ◽  
C Terminus ◽  
Factor Viiia ◽  
Solid Phase Binding

SummaryWe have previously used a solid phase binding assay to localize a Factor X (FX) interactive site to the acidic C-terminus of the A1 subunit of FVIIIa (Lapan KA, Fay PJ. J Biol Chem 1997; 272: 2082-2088). The complex of FVIII-FX was made covalent following reaction with the zero-length cross-linking reagent 1-ethyl-3-(3-dimethylaminopropyl-)carbodiimide hydrochloride (EDC). Western blotting of the thrombin-cleaved complex showed that the A1 subunit of FVIIIa associated with FX heavy chain. The FX-A1 product was also detected following cross-linking to the A1/A3-C1-C2 dimer, but not the activated protein C-cleaved A1336/A3-C1-C2 form, indicating that a residue(s) in the region spanning Met337-Arg372 contributed to the intermolecular ion pair(s). A synthetic peptide to this acidic region (FVIII337-372) cross-linked to FX and the product was alkaline resistant indicating that amide linkage(s) were formed. Sequence analysis of the FX-FVIII337-372 adduct suggested that the first 12 NH2-terminal residues of the FX and peptide do not participate in cross-link formation. Conversion of the cross-linked product to FXa by RVV-X showed that the peptide was associated with the serine protease-forming domain of the heavy chain. These results indicate that the association of FVIIIa and FX occurs from a salt linkage(s) formed between residues of the A1 acidic C-terminus of the cofactor (within residues 349-372) and the serine protease-forming domain of the substrate.

Role of ovarian follicle cells in vitellogenesis and oocyte resorption inCapitella sp. I (Polychaeta)

1984 ◽  
Vol 79 (2) ◽  
pp. 133-144 ◽  
Author(s):  
K. J. Eckelbarger ◽  
P. A. Linley ◽  
J. P. Grassle
Keyword(s):  
Ovarian Follicle ◽  
Follicle Cells ◽  
Oocyte Resorption
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