Marina Ferraro

Ag staining of the nucleolus organizer (NO) and its relationship to satellite association

SummaryThe frequency of involvement in satellite association and the frequency of selective staining of the secondary constrictions with silver solutions have been studied in five phenotypically normal individuals, all carriers of morphological variants of the nucleolus organizing region (NOR). The results show the preferential involvement of some morphological markers in satellite association, and also their preferential staining with Ag-I. It has also been shown that acrocentric chromosomes involved in satellite association are always stained by silver.

DNA methylation changes during mouse spermatogenesis

Genomic imprinting in mammals is thought to be mediated by differences in the methylation level of cytosine residues in the genome. These differences in DNA methylation are thought to be generated during the development of the germ line. To characterize the profile of global methylation of the mouse genome during male gametogenesis, we have quantified the relative level of methylation in individual cells during meiosis and spermatogenesis. A decrease in the level of DNA methylation is observed from meiotic cells to elongated spermatids. The erasure of the somatic pattern of methylation during spermatogenesis suggests the existence of a subsequent mechanism generating the parental specific methylation patterns leading to genomic imprinting of specific alleles.

Silver staining of the nucleolus organizer regions (NOR) requires clusters of sulfhydryl groups.

Silver stainability of nucleolus organizer regions (NORs) appears to be correlated with the presence of grouped sulfhydryl (SH) side chains of proteins. In fact, heavy metals with high affinity for SH groups, such as Hg and Cu, do prevent the silver staining reaction. Ferricyanide, which is known to oxidize SH to disulfides, also prevents any further reaction with silver. On the other hand, alkali and reducing agents (mercaptoethanol, cyanide) do not affect silver stainability of the NORs. These results show that the silver staining reaction is not related to disulfide or persulfide groups and that alkali-soluble, acidic nuclear proteins per se do not play a major role in this process.

Activation of human ribosomal genes by 5-azacytidine☆

Cultured human fibroblasts were exposed to 5-azacytidine which inhibits methylation of newly synthesized DNA. A significant increase in the mean number of Ag-stained active nucleolus organizers has been observed in treated cells. This suggests that DNA methylation is involved in modulation of human rRNA synthesis.

Involvement of histone H4 acetylation in the epigenetic inheritance of different activity states of maternally and paternally derived genomes in the mealybug Planococcus citri

Abstract. Modification of histones by acetylation is a well-known mechanism for the establishment and maintenance of specific chromatin structures with different activity states. In Planococcus citri males the paternal genome, early in development, becomes mostly inactive and heterochromatic. As we had not found methylation in the genome of P. citri, we analyzed the acetylation state of histone H4. We report here that, in males, differences in the level of histone H4 acetylation are indeed present in the two genomes of different parental origin; these differences were confirmed by treatment with the histone deacetylase inhibitor Trichostatin A. There is also evidence of acetylation of histone H4 on metaphase chromosomes. Our data therefore suggest a role of histone H4 acetylation in the imprinting of the paternal genome in P. citri males, thus supporting a role of modification of chromatin-related structural proteins in the epigenetic transmission of imprinting.

Clonal inheritance of rRNA gene activity: Cytological evidence in human cells

Clone-specific silver staining patterns of the nucleolus organizers (NOs) were observed in clones deriving from a human diploid fibroblast culture. Each specific staining pattern corresponds to one of the nine patterns observed in the general population. Since silver stainability of the NOs is related to rRNA gene function, these data demonstrate the clonal inheritance of rRNA gene activity.

Germline cyst development and imprinting in male mealybug Planococcus citri.

In the epigenetic modifications involved in the phenomenon of imprinting, which is thought to take place during gametogenesis, one of the primary roles is exerted by histone tail modifications acting on chromatin structure. What is more, in insects like mealybugs, with a lecanoid chromosome system, imprinting is strictly related to sex determination. In many diverse species gametes originate in specific, highly evolutionarily conserved structures called germline cysts. The use of staining techniques specific for fusomal components like F-actin has allowed us to describe for the first time the morphogenesis of male germline cysts in the mealybug Planococcus citri. Antibodies to anti-methylated lysine 9 of histone H3 (MeLy9-H3) and anti-heterochromatin protein 1 (HP1) were used during cyst formation to investigate the involvement of these epigenetic modifications in the phenomenon of imprinting and their possible concerted action in sex determination in P. citri. These observations indicate: (i) a specific role for F-actin in the segregation, typical of the lecanoid chromosome system, of genomes of paternal origin; (ii) that the two vital gametes originating from a given meiosis, although carrying the same genome, differ in the levels of both MeLy9-H3 and HP1, one of them being more heavily labelled by both antibodies.

Nucleolus organizer and N-band distribution in morphologic and fluorescence variants of human chromosomes

SummaryThree cases of morphologic variants of human D- or G-group chromosomes have been studied by N- and Ag-AS banding techniques. The results confirm our previous findings about the localization of nucleolus organizers on the secondary constrictions of acrocentric chromosomes. Preliminary results on the distribution and number of N bands in D- and G-group chromosomes with morphologic and fluorescence variants are reported

Cytogenetic characterization of the genome of mealybug Planococcus citri (Homoptera, Coccoidea)

Summary Mealybugs although being agriculturally harmful insects have been very poorly studied by modern cytogenetics techniques, and no cytotaxonomic criteria to distinguish between closely related species is available yet. In the mealybug Planococcus citri (2n=10) male and female individuals are both diploid, however in males, at the stage of blastula, the haploid chromosome set of paternal origin becomes heterochromatic, even though its complete inertia has been considered questionable. Here we present data on the cytogenetic characterization of the chromosomes of Planococcus citri. We report on (i) the fluorescence karyotype (D287/170), which to our knowledge is the first banded karyotype of a mealybug to be described; (ii) the chromosome localization of constitutive heterochromatin; (iii) the chromosome localization of rDNA sites; (iv) NORs activity. Our data also show, for the first time, that in the heterochromatic chromosome set ribosomal genes are still active.

Nucleolus organizer and satellite association in a variant D-group chromosome.

SummaryA No. 15 chromosome with a short arm longer than usual is observed in two phenotypically normal brothers. This chromosome appears to have no visible satellite, shows no N-band staining, and is never involved in satellite association. These results have led us to the conclusion that this chromosome lacks the nucleolus organizer region.