Transgenic mice with inactive alleles for procollagen N-proteinase (ADAMTS-2) develop fragile skin and male sterility

2001 ◽  
Vol 355 (2) ◽  
pp. 271-278 ◽  
Author(s):  
Shi-Wu LI ◽  
Machiko ARITA ◽  
Andrzej FERTALA ◽  
Yunhua BAO ◽  
Gene C. KOPEN ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Transgenic Mice ◽  
Large Fraction ◽  
Polarized Light ◽  
Collagen Fibrils ◽  
Type I ◽  
Type Ii ◽  
Newborn Mice ◽  
Type I Procollagen ◽  
Thin Skin

Transgenic mice were prepared with inactive alleles for procollagen N-proteinase (ADAMTS-2; where ADAMTS stands for adisintegrin and metalloproteinase with thrombospondin repeats). Homozygous mice were grossly normal at birth, but after 1-2 months they developed thin skin that tore after gentle handling. Although the gene was inactivated, a large fraction of the N-propeptides of type I procollagen in skin and the N-propeptides of type II procollagen in cartilage were cleaved. Therefore the results suggested the tissues contained one or more additional enzymes that slowly process the proteins. Electron microscopy did not reveal any defects in the morphology of collagen fibrils in newborn mice. However, in two-month-old mice, the collagen fibrils in skin were seen as bizarre curls in cross-section and the mean diameters of the fibrils were approx. half of the controls. Although a portion of the N-propeptides of type II procollagen in cartilage were not cleaved, no defects in the morphology of the fibrils were seen by electron microscopy or by polarized-light microscopy. Female homozygous mice were fertile, but male mice were sterile with a marked decrease in testicular sperm. Therefore the results indicated that ADAMTS-2 plays an essential role in the maturation of spermatogonia.

Specific and selective inhibition of translation of mRNA for type II procollagen in cells of transgenic mice that synthesize type I procollagen. A possible role for endogneous antisense RNA

1996 ◽  
Vol 15 (3) ◽  
pp. 203-210
Author(s):  
Leena Ala-Kokko ◽  
Constance Yuan ◽  
Dominique Le Guellec ◽  
Suzanne Franc ◽  
Andrzej Fertala ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Antisense Rna ◽  
Selective Inhibition ◽  
Type I ◽  
Type Ii ◽  
Type I Procollagen ◽  
In Cells

scholarly journals Lack of a phenotype in transgenic mice aberrantly expressing COL2A1 mRNA because of highly selective post-transcriptional down-regulation

2000 ◽  
Vol 345 (2) ◽  
pp. 377-384 ◽  
Author(s):  
Constance M. YUAN ◽  
Leena ALA-KOKKO ◽  
Dominique LE GUELLEC ◽  
Suzanne FRANC ◽  
Andrzej FERTALA ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Type I Collagen ◽  
Type Ii Collagen ◽  
Type I ◽  
Mrna Levels ◽  
Type Ii ◽  
Col2a1 Gene ◽  
Type I Procollagen ◽  
Steady State Levels ◽  
Hybridization In Situ

We reported previously that a 1.9-kb 5ʹ-fragment from the human COL1A1 gene drove transcription of a promoterless human COL2A1 gene in tissues of transgenic mice that normally express the COL1A1 but not the COL2A1 gene. In the present study, we have established that the aberrant transcription of the COL2A1 gene did not produce any gross or microscopic phenotype, because the transcripts were not efficiently translated in cells that do not normally express the COL2A1 gene. In two lines of transgenic mice, the mRNA levels from the transgene were 30% to 45% of the mRNA for the proα1(I) chain of type I procollagen, the most abundant mRNA in the same tissues. Analysis of collagens extracted from skin of the transgenic mice indicated that triple-helical type II collagen, with the normal pattern of cyanogen bromide peptides, was synthesized from the transgene. However, the level of type II collagen in skin was less than 2% of the level of type I collagen. Hybridization in situ indicated the presence of mRNA for both COL2A1 and COL1A1 in the same cells. Immunofluorescence staining for type II collagen, however, was negative in the same tissues. The results, therefore, indicated that many mesenchymal cells in the transgenic mice had high steady-state levels of the homologous mRNAs for type I and type II procollagen, but only the mRNAs for type I procollagen were efficiently translated.

scholarly journals Substitutions of aspartic acid for glycine-220 and of arginine for glycine-664 in the triple helix of the pro α1(I) chain of type I procollagen produce lethal osteogenesis imperfecta and disrupt the ability of collagen fibrils to incorporate crystalline hydroxyapatite

1995 ◽  
Vol 311 (3) ◽  
pp. 815-820 ◽  
Author(s):  
A A Culbert ◽  
M P Lowe ◽  
M Atkinson ◽  
P H Byers ◽  
G A Wallis ◽  
...  
Keyword(s):  
Osteogenesis Imperfecta ◽  
Aspartic Acid ◽  
Triple Helix ◽  
Type I Collagen ◽  
Collagen Fibrils ◽  
Morphological Data ◽  
Type I ◽  
Type Ii ◽  
Col1a1 Gene ◽  
Type I Procollagen

We identified two infants with lethal (type II) osteogenesis imperfecta (OI) who were heterozygous for mutations in the COL1A1 gene that resulted in substitutions of aspartic acid for glycine at position 220 and arginine for glycine at position 664 in the product of one COL1A1 allele in each individual. In normal age- and site-matched bone, approximately 70% (by number) of the collagen fibrils were encrusted with plate-like crystallites of hydroxyapatite. In contrast, approximately 5% (by number) of the collagen fibrils in the probands' bone contained crystallites. In contrast with normal bone, the c-axes of hydroxyapatite crystallites were sometimes poorly aligned with the long axis of fibrils obtained from OI bone. Chemical analysis showed that the OI samples contained normal amounts of calcium. The probands' bone samples contained type I collagen, overmodified type I collagen and elevated levels of type III and V collagens. On the basis of biochemical and morphological data, the fibrils in the OI samples were co-polymers of normal and mutant collagen. The results are consistent with a model of fibril mineralization in which the presence of abnormal type I collagen prevents normal collagen in the same fibril from incorporating hydroxyapatite crystallites.

scholarly journals Two Distinct Notch1 Mutant Alleles Are Involved in the Induction of T-Cell Leukemia in c-myc Transgenic Mice

2000 ◽  
Vol 20 (11) ◽  
pp. 3831-3842 ◽  
Author(s):  
C. D. Hoemann ◽  
N. Beaulieu ◽  
L. Girard ◽  
N. Rebai ◽  
P. Jolicoeur
Keyword(s):  
Cell Surface ◽  
Transgenic Mice ◽  
Intracellular Signaling ◽  
Transmembrane Domain ◽  
Tumor Formation ◽  
Type I ◽  
Dependent Manner ◽  
Type Ii ◽  
Extracellular Medium ◽  
Calcium Dependent

ABSTRACT We have previously characterized a large panel of provirus insertion Notch1 mutant alleles and their products arising in thymomas of MMTVD/myc transgenic mice. Here, we show that these Notch1 mutations represent two clearly distinct classes. In the first class (type I), proviral integrations were clustered just upstream of sequences encoding the transmembrane domain. Type I Notch1 alleles produced two types of mutantNotch1 RNA, one of which encoded the entire Notch1 cytoplasmic domain [N(IC)] and the other of which encoded a soluble ectodomain [N(EC)Mut] which, in contrast to the processed wild-type ectodomain [N(EC)WT], did not reside at the cell surface and became secreted in a temperature-dependent manner. A second, novel class of mutant Notch1 allele (type II) encoded a Notch1 receptor with the C-terminal PEST motif deleted (ΔCT). The type II Notch1ΔCT protein was expressed as a normally processed receptor [N(EC)WT and N(IC)ΔCT] at the cell surface, and its ectodomain was found to be shed into the extracellular medium in a temperature- and calcium-dependent manner. These data suggest that both type I and type II mutations generate two structurally distinct Notch1 N(EC) and N(IC) proteins that may participate in tumor formation, in collaboration with the c-myc oncogene, through distinct mechanisms. Constitutive type I N(IC) and type II N(IC)ΔCT expression may enhance Notch1 intracellular signaling, while secreted or shed type I N(EC)Mut and type II N(EC) proteins may differentially interact in an autocrine or paracrine fashion with ligands of Notch1 and affect their signaling.

scholarly journals Expression of a Truncated, Kinase-Defective TGF-β Type II Receptor in Mouse Skeletal Tissue Promotes Terminal Chondrocyte Differentiation and Osteoarthritis

1997 ◽  
Vol 139 (2) ◽  
pp. 541-552 ◽  
Author(s):  
Rosa Serra ◽  
Mahlon Johnson ◽  
Ellen H. Filvaroff ◽  
James LaBorde ◽  
Daniel M. Sheehan ◽  
...  
Keyword(s):  
Articular Cartilage ◽  
Transgenic Mice ◽  
Growth Plate ◽  
Dominant Negative Mutant ◽  
Type I ◽  
Type Ii ◽  
Type X Collagen ◽  
Skeletal Tissue ◽  
Hypertrophic Cartilage ◽  
Growth Plate Cartilage

Members of the TGF-β superfamily are important regulators of skeletal development. TGF-βs signal through heteromeric type I and type II receptor serine/threonine kinases. When over-expressed, a cytoplasmically truncated type II receptor can compete with the endogenous receptors for complex formation, thereby acting as a dominant-negative mutant (DNIIR). To determine the role of TGF-βs in the development and maintenance of the skeleton, we have generated transgenic mice (MT-DNIIR-4 and -27) that express the DNIIR in skeletal tissue. DNIIR mRNA expression was localized to the periosteum/perichondrium, syno-vium, and articular cartilage. Lower levels of DNIIR mRNA were detected in growth plate cartilage. Transgenic mice frequently showed bifurcation of the xiphoid process and sternum. They also developed progressive skeletal degeneration, resulting by 4 to 8 mo of age in kyphoscoliosis and stiff and torqued joints. The histology of affected joints strongly resembled human osteo-arthritis. The articular surface was replaced by bone or hypertrophic cartilage as judged by the expression of type X collagen, a marker of hypertrophic cartilage normally absent from articular cartilage. The synovium was hyperplastic, and cartilaginous metaplasia was observed in the joint space. We then tested the hypothesis that TGF-β is required for normal differentiation of cartilage in vivo. By 4 and 8 wk of age, the level of type X collagen was increased in growth plate cartilage of transgenic mice relative to wild-type controls. Less proteoglycan staining was detected in the growth plate and articular cartilage matrix of transgenic mice. Mice that express DNIIR in skeletal tissue also demonstrated increased Indian hedgehog (IHH) expression. IHH is a secreted protein that is expressed in chondrocytes that are committed to becoming hypertrophic. It is thought to be involved in a feedback loop that signals through the periosteum/ perichondrium to inhibit cartilage differentiation. The data suggest that TGF-β may be critical for multifaceted maintenance of synovial joints. Loss of responsiveness to TGF-β promotes chondrocyte terminal differentiation and results in development of degenerative joint disease resembling osteoarthritis in humans.

Collagen expression in embryonic chicken chondrocytes treated with phorbol myristate acetate

1985 ◽  
Vol 5 (6) ◽  
pp. 1415-1424
Author(s):  
M H Finer ◽  
L C Gerstenfeld ◽  
D Young ◽  
P Doty ◽  
H Boedtker
Keyword(s):  
Morphological Change ◽  
Type I Collagen ◽  
Late Phase ◽  
Tumor Promoter ◽  
Type I ◽  
Mrna Levels ◽  
Type Ii ◽  
Collagen Mrna ◽  
Type I Procollagen ◽  
Embryonic Chicken

Growth of embryonic chicken sternal chondrocytes in the presence of phorbol-12-myristate-13-acetate (PMA), a potent tumor promoter, resulted in a dramatic morphological change from spherical floating cells to adherent fibroblastic cells. This morphological change was accompanied by a quantitative switch from synthesis of cartilage-specific type II procollagen to type I procollagen. Type II procollagen mRNA levels decreased 10-fold in PMA-treated cells. Activation of type I collagen genes led to the accumulation of type I procollagen mRNA levels comparable to those of type II mRNA in these cells. However, only type I procollagen mRNA was translated. In addition to gene activation, unprocessed pro alpha 1(I) transcripts present at low levels in control chondrocytes were processed to mature mRNA species. Redifferentiation of PMA-treated chondrocytes was possible if cells were removed from PMA after the morphological change and cessation of type II procollagen synthesis but before detectable amounts of type I procollagen were synthesized. Production of type I collagen thus marks a late phase of chondrocyte "dedifferentiation" from which reversion is no longer possible. Redifferentiated cell populations contained 24-fold more pro alpha 1(II) collagen mRNA than pro alpha 1(I) collagen mRNA, but the rates of procollagen synthesis were comparable. This suggests that the PMA-mediated dedifferentiation of chondrocytes as well as their redifferentiation is under both transcriptional and posttranscriptional regulation.

Protein Tomography Analysis Shows That CD20 Molecules Bound by Different Type I and Type II Anti-CD20 Antibodies Are Organized in Different Protein Complexes.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4782-4782
Author(s):  
Gerhard J Niederfellner ◽  
Annika Braennstroem ◽  
Frida Lindstrom ◽  
Magnus Jansson ◽  
Marika Lundin ◽  
...  
Keyword(s):  
Protein Complexes ◽  
Large Fraction ◽  
Type I ◽  
Type Ii ◽  
Employment Equity ◽  
Downstream Signaling ◽  
Equity Ownership ◽  
Anti Cd20 ◽  
Tomography Analysis ◽  
Cd20 Antibodies

Abstract Abstract 4782 Ramos cells were labeled with Type I (Rituximab and Ocrelizumab) and Type II (B1/H299 and GA101) anti-CD20 antibodies and then fixed and stained with marker gold labeled 2ary antibodies. Protein complexes connected to marker gold were analyzed by Protein TomographyTM at SIDEC and the corresponding 3D-structures visualized (∼100 structures per antibody). Less than 5 % of the refined structures were dimeric. While most (60 – 90%) CD20 molecules were present in tetramers or even higher order defined complexes, a sizeable proportion was also engaged in large protein networks (11 -38%). For the multimeric complexes, we could clearly distinguish between extended (or open) and ring-like (or closed) conformations. Although cells had been labeled with an excess of antibody, as suggested by FACS binding curves, the antibodies bound CD20 monovalently in most refined structures. Bivalent binding was overall more prevalent with Rituximab and Ocrelizumab than with B1 and GA101 (1/3 vs 1/6 structures). The proportion of CD20 molecules present in ringlike complexes was higher for the Type II than for the Type I antibodies. In co-localization experiments, Rituximab-CD20 and GA101-CD20 complexes were also found to only partially colocalize, while a large fraction of the two antibodies were found in separate cell surface compartments. These findings suggest that the different antibodies favor different CD20 conformations that seem to be associated with different protein complexes and might form the basis for initiation of different downstream signaling processes. Disclosures: Niederfellner: Roche: Employment. Braennstroem:SIDEC: Employment. Lindstrom:SIDEC: Employment. Jansson:SIDEC: Employment. Lundin:SIDEC: Employment. Schaefer:Roche: Employment. Mundigl:Roche: Employment. Umana:Glycart: Employment, Equity Ownership, Patents & Royalties. Klein:Roche: Employment, Equity Ownership, Patents & Royalties.

Structure and synthesis of the peritrophic membrane in Trichoptera larvae

Zoosymposia ◽  
2011 ◽  
Vol 5 (1) ◽  
pp. 63-70
Author(s):  
CARLA CORALLINI
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Cardiac Valve ◽  
Food Type ◽  
Peritrophic Membrane ◽  
Type I ◽  
Type Ii ◽  
Food Ingestion ◽  
Transmission Electron ◽  
Anterior Midgut

The peritrophic membrane (PM) in Trichoptera larvae was examined by light, scanning and transmission electron microscopy. The gut of most insects produces 2 fundamental types of PM: Type I is synthesized and secreted by the entire midgut epithelium in response to the ingestion of food; type II is synthesized by specialized cells of the cardiac valve located in the anterior midgut independent from food ingestion. Corallini (2003) described, in the midgut of Limnephilidae larvae, a type I PM which is also secreted by unfed larvae. In this study, both types of PM were observed. Type I PM was evident in larvae of Rhyacophilidae, Leptoceridae, Sericostomatidae and Odontoceridae; the type II PM was observed in larvae of Philopotamidae, Polycentropodidae and Hydropsychidae.

scholarly journals Stepwise proteolytic activation of type I procollagen to collagen within the secretory pathway of tendon fibroblasts in situ

2011 ◽  
Vol 441 (2) ◽  
pp. 707-717 ◽  
Author(s):  
Elizabeth G. Canty-Laird ◽  
Yinhui Lu ◽  
Karl E. Kadler
Keyword(s):  
Extracellular Matrix ◽  
Secretory Pathway ◽  
Brefeldin A ◽  
Collagen Fibrils ◽  
Proteinase Activity ◽  
Type I ◽  
Proteolytic Activation ◽  
Type I Procollagen ◽  
Tail Tendon

Proteolytic cleavage of procollagen I to collagen I is essential for the formation of collagen fibrils in the extracellular matrix of vertebrate tissues. Procollagen is cleaved by the procollagen N- and C-proteinases, which remove the respective N- and C-propeptides from procollagen. Procollagen processing is initiated within the secretory pathway in tendon fibroblasts, which are adept in assembling an ordered extracellular matrix of collagen fibrils in vivo. It was thought that intracellular processing was restricted to the TGN (trans-Golgi network). In the present study, brefeldin A treatment of tendon explant cultures showed that N-proteinase activity is present in the resulting fused ER (endoplasmic reticulum)–Golgi compartment, but that C-proteinase activity is restricted to the TGN in embryonic chick tendon fibroblasts. In late embryonic and postnatal rat tail and postnatal mouse tail tendon, C-proteinase activity was detected in TGN and pre-TGN compartments. Preventing activation of the procollagen N- and C-proteinases with the furin inhibitor Dec-RVKR-CMK (decanoyl-Arg-Val-Lys-Arg-chloromethylketone) indicated that only a fraction of intracellular procollagen cleavage was mediated by newly activated proteinases. In conclusion, the N-propeptides are removed earlier in the secretory pathway than the C-propeptides. The removal of the C-propeptides in post-Golgi compartments most probably indicates preparation of collagen molecules for fibril formation at the cell–matrix interface.

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