Granular Loops Research Articles

C3-Independent Glomerulonephritis in Guinea Pigs: Dependence upon Primary Humoral Response

We used C3-deficient (C3D) guinea pigs to evaluate the role of C3 in an active model of experimental nephritis. Normal strain 2 (C3N, n = 13) and C3D (n = 6) guinea pigs were immunized with cationized bovine γ-globulin (CBGG). Fourteen days later (Day 0), daily intravenous injections of CBGG were given for 3 to 7 days and the animals were sacrificed on Day 10 or 21. Immunofluorescence (IF) microscopy of renal tissue revealed two patterns of glomerular IgG deposition: granular loop (11/13 C3N, 3/6 C3D), and predominantly mesangial (2/13 C3N, 3/6 C3D). Codeposited C3 was seen in all C3N and in no C3D animals. Electron microscopy showed subepithelial deposits in all. A significant correlation (P < 0.005) was seen between an animal's IF pattern and its level of serum antibodies to CBGG; those with lower antibody levels exhibited the mesangial pattern. C3D animals had lower mean antibody levels than C3N (P < 0.01), but both IF patterns were represented. Urine protein concentration, which was increased relative to controls, did not differ between C3N and C3D groups, but was significantly greater in those with loop IF. Serum albumin was significantly reduced in animals with loop IF. C3N animals showed a significant reduction in mean serum C3. In this model, immune deposit location and degree of proteinuria are independent of C3 deposition and dependent upon the level of antibody response to CBGG. Induction of antibody to CBGG is impaired in the absence of C3.

Ultrastructural similarity in landmark loops of amphibian lampbrush chromosomes

Summary— Simultaneous transmission and scanning electron microscopy studies were performed on lampbrush chromosomes of Notophthalmus viridescens and Xenopus laevis. The organization of their normal and landmark loop ribonucleoprotein (RNP) matrices was compared to that of Pleurodeles waltl lampbrush loops, previously described. Ultrastructural observations clearly showed that in the three species, the RNP matrix of normal and landmark loops displayed a common basic structure: an RNP fibril packed into tightly juxtaposed RNP particles of remarkably uniform size, ie 30 nm. Furthermore, analysis of the spatial arrangement of these constitutive RNP fibrils allowed us to establish ultrastructural similarities between the different types of loop matrices of the three species studied. Thus, granular loops with the same organization were found to be present in the three species, whereas Pleurodeles was the only one to exhibit, in its lampbrush chromosomes, the typical globular matrices previously described. “Sequential labelling loops” of Notophthelmus were shown to be similar of both “convoluted dense loops” of Xenopus and “dense loops” of Pleurodeles.

Effects of cold shock treatment on lampbrush chromosomes of amphibian oocytes

When females of the newt Pleurodeles waltl, normally raised in our laboratory at 20 °C, were placed in 8 °C water for several days, striking modifications occurred in the structure of oocyte lampbrush chromosomes: numerous normal-type lateral loops were partially reduced in size and number, while hyperdeveloped loops of a new morphological type occurred at constant, reproducible loci. However, induction of these cold loops apparently was not accompanied by preferential RNA synthesis at their levels. Cold loops resulted from the decompaction of specific landmarks, the granular loops. Autoradiography revealed a significant drop in lampbrush chromosome transcriptional activity at 8 °C. Both structural and transcriptional modifications were fully reversible when oocytes were returned to normal temperature. These modifications are discussed in relation to processes which could determine the morphological appearance of landmark loops.

Correlation between isolated lampbrush chromosomes and nuclear structures of Pleurodeles waltl oocytes: An electron microscopic study

The organization of the lampbrush chromosomes of Pleurodeles waltl was studied by fixation and embedding of oocytes in toto and correlated with that observed in end-embedded preparations of isolated chromosomes. Particular attention was focused on marker loops, like the granular and globular loops, and atypical structures known as spheres (S) and M. In both types of preparations, the majority of the loops, the so-called normal loops, and the granular loops appeared to share a common basic organization, with ribonucleoprotein (RNP) fibrils appearing as strings of 30 nm particles, as described by earlier authors. Some new types of loop organization were observed: (i) P loops with 45 nm RNP particles; (ii) dense granular loops; (iii) loops with a cylindrical organization. RNP fibrils formed by 60 nm particles were found to occur in association with the globular loops. EDTA staining suggested the presence of large amounts of RNP in the sphere but very little in M. Three morphologically different types of RNP granules could be observed free in the nucleoplasm.

An analog of Xenopus N1N2 protein in Pleurodeles waltl.

The oocyte nucleus of Pleurodeles waltl contains a major 185-kDa protein analog of Xenopus N1N2. Rabbit antibodies were raised against the 185-kDa protein. Affinity-purified antibody directed against the acid part (pI 4.7) of the protein was prepared using antigens separated by two-dimensional electrophoresis. Specificity of the antibody was controlled on two-dimensional gels of nuclear proteins. It was shown that the 185-kDa protein was separated into 2 forms: an acid form of pI 4.7-5.3, and a base form of pI 7. Peptide maps of the 2 spots revealed that they were closely related. The antibody was tested on: a) spread nuclear content, b) on sections of embryonic stages from stage 2 to stage 34, and c) the sections of adult tissues. The 185-kDa protein appeared to be associated with the RNP matrix of a particular type of lampbrush chromosome loop, the granular loop. This protein was present in the nuclei of all embryonic cells. In adult tissues, it was present only in the nuclei of cells which presented high mitotic activity. These results confirm that, like N1N2, the 185-kDa protein interacts with the constitution of chromatin; furthermore, they provide evidence for the role of this protein in transcriptional activity.

Function of lampbrush chromosomes: A hypothesis

A hypothesis about lampbrush chromosome function is proposed which takes into account several aspects of their morphology and behavior. It is suggested that in lampbrush loops, alterations of the deoxyribonucleoprotein fiber take place which reprogram the chromosome for development. Such reprogramming could involve exchange or enzymatic modification of regulatory molecules (i.e., non-histone proteins). Since inhibition of RNA synthesis causes the loops to collapse, it is possible that transcription largely occurs to generate and maintain the loop, allowing the DNP fiber to “enter” the nucleoplasm. Lampbrush chromosomes are also involved in the production of “masked” mRNAs for early development. The giant granular loops could either be the sites where such mRNAs are packaged or they could be sites where specific alterations of the deoxyribonucleoprotein fiber take place.

The maps of the lampbrush chromosomes of Triturus (Amphibia Urodela). IV. Triturus vulgaris meridionalis.

The lampbrush chromosomes of Triturus vulgaris meridionalis were isolated from the germinal vesicle of medium and large-sized oocytes and studied with phase-contrast microscope. The maps were constructed on the basis of the lengths and major morphological features of the chromosomes. The length of each map is equal to the mean of the relative lengths of the corresponding chromosome from different oocytes (the relative length of each chromosome is represented by the ratio between its absolute length and that of chromosome XII from the same complement, conventionally considered as 100 units long). The maps arranged in decreasing length order, were oriented according to the most frequent position of chiasmata, as centromeres were not always evident. — Chromosomes VI and XI bear a sphere in subterminal position. Landmarks typical for T. vulgaris meridionalis are the loops inserted on chromosomes VIII (47 units), X (23 units), XI (34 units) and XII (34 units) frequently presenting themselves under the form of double loop bridges of considerable extension. On chromosomes I (4 units), VI (13 units), X (4 units) and XI (36 units) giant bodies were found that are sometimes comparable to dense-matrix loops. Chromosome XI includes a nucleolus-organizing region, sometimes identifiable by the presence of an inserted nucleolus. Normal and granular loops (much extended at times), axial granules, globules, and loopless bars supplement the morphology of the lampbrush chromosomes of this species.

Evidence for a polarized movement of the lateral loops of newt lampbrush chromosomes during oogenesis.

ABSTRACT Actinomycin D inhibits RNA synthesis on the lateral loops of newt lampbrush chromsomes. Partial inhibition does not provoke marked morphological alteration of ordinary lateral loops, most of which recover to the full their capacity for RNA synthesis within 2 days of treatment. However, occasional ordinary loops do not recover completely within the first few days after treatment, and in such loops RNA-synthesizing capacity is restricted to a region adjoining the thinner insertion in the parent chromomere. A greater degree of inhibition of RNA synthesis is accompanied by loss of matrix from ordinary lateral loops, and in the extreme case the loop axes retract to their parent chromomeres and neighbouring chromomeres coalesce; for the ordinary loops, full recovery from this stripped condition is nevertheless possible. Some 20 μ per loop extends during the first day following exposure to actinomycin, and normal morphology and RNA-synthesizing capacity are regained within 2–4 days. The giant granular loop of Triturus cristatus cristatus chromosome XII responds to extreme actinomycin D poisoning in different fashion. Matrix does not at once slough off its loop axis, but the loop present at the time of treatment is progressively replaced by a new granular loop which develops between the parent chromomere and the original loop’s dense tip. These observations support the theory that the DNA-containing axes of all lateral loops of lampbrush chromosomes continually extend from their parent chromomeres, engage in RNA synthesis while extended, and carry the associated RNP matrix along as they move towards the return insertions in the parent chromomeres, where loop axis retraction occurs.