Positional changes of pericentromeric heterochromatin and nucleoli in postmitotic Purkinje cells during murine cerebellum development

Abstract

Previous studies revealed changes of pericentromeric heterochromatin arrangements in postmitotic Purkinje cells (PCs) during postnatal development in the mouse cerebellum (Manuelidis, 1985; Martou and De Boni, 2000). Here, we performed vibratome sections of mouse cerebellum (vermis) at P0 (day of birth), at various stages of the postnatal development (P2–P21), as well as in very young (P28) and 17-months-old adults. FISH was carried out on these sections with major mouse satellite DNA in combination with immunostaining of the nucleolar protein B23 (nucleophosmin). Laser confocal microscopy, 3D reconstructions and quantitative image analysis were employed to describe changes in the number and topology of chromocenters and nucleoli. At all stages of postnatal PC development heterochromatin clusters were typically associated either with nucleoli or with the nuclear periphery, while non-associated clusters were rare (<1% at P0 to P21 and about 3% in adult stages). At P0, about 2–4 nucleoli and 7–8 pericentromeric heterochromatin clusters were variably located within PC nuclei. The relative volume of heterochromatin clusters associated with the nucleoli (about 50%) was roughly equal to the volume of clusters associated with the nuclear periphery. Positional changes of both nucleoli and centromeres towards the nuclear center occurred between P0 and P6. At P6 the average number of chromocenters per PC nucleus had decreased to about five. In agreement with previous studies, one or occasionally two nucleoli were noted at the nuclear center surrounded by major perinucleolar heterochromatin clusters. The relative volume of these perinucleolar clusters increased to about 84%, while the volume of clusters in the nuclear periphery decreased to about 15%. At subsequent postnatal stages, the arrangement of most pericentromeric heterochromatin around a central nucleolus was maintained. In adult animals, however, we observed a partial redistribution of heterochromatin towards the nuclear periphery. The average total number of pericentromeric heterochromatin signals increased again to about ten. The volume of heterochromatin associated with the nuclear periphery roughly doubled (30%), while the volume of the perinucleolar heterochromatin decreased correspondingly.

Transcriptional Activation of Mouse Major Satellite Regions during Neuronal Differentiation

“…Although it has been previously shown that the localization, number, and size of major satellite regions change during neural development (Billia et al, 1992;Solovei et al, 2004Solovei et al, , 2009, we now show the first evidence that these changes are accompanied by a change in the chromatin state and transcriptional activity of this region. The major satellite regions are traditionally considered to be "silent" regions located within the heterochromatin and marked by repressive histone modifications.…”

“…Histone H3 lysine-4 trimethylation becomes localized to chromocenters during neuronal differentiation Because of the previous observation that the number, location and size of major satellite foci change during neuronal development, we asked whether neuronal differentiation is also accompanied by changes in the chromatin state of these regions (Billia et al, 1992;Solovei et al, 2004Solovei et al, , 2009). We used immunostaining to examine histone modifications, a reflection of chromatin state, in major satellite regions during neuronal differentiation.…”

“…However, previous evidence suggests a possible role for major satellite regions during neuronal differentiation. DNA-fluorescence in situ hybridization (DNA-FISH) analysis indicated that the number, localization, and size of major satellite foci change during development of post-mitotic Purkinje neurons or rod photoreceptors (Solovei et al, 2004(Solovei et al, , 2009). Furthermore, long-term potentiation-inducing stimuli were shown to decrease the number of major satellite foci in post-mitotic hippocampal neurons (Billia et al, 1992), and transcripts from major satellite regions have been detected in the developing neocortex (Rudert et al, 1995).…”

Building up the nucleus: nuclear organization in the establishment of totipotency and pluripotency during mammalian development

“…genomic loci. Indeed, the composition of the nuclear lamina, the position and architectural changes of the nucleolus, and the sequestration of genomic loci and regulatory proteins into promyelocytic leukemia (PML) bodies, nuclear speckles, histone locus bodies, transcription factories, Cajal bodies, and others have all been linked to gene regulation across a variety of cell types (Solovei et al 2004;Peric-Hupkes et al 2010;Sleeman and Trinkle-Mulcahy 2014).…”

Maintenance of imprinting and nuclear architecture in cycling cells

“…Live-cell observations of movements of a GFP-tagged transgene array in a mammalian cell line (CHO) yielded long-range movements over distances of 1-5 m at velocities from 0.1 to 0.9 m/min (34). Large-scale movements were also observed in nuclei during postmitotic cell differentiation, but these movements required a time scale of days rather than hours (35). In conclusion, it seems unlikely that directed chromatin movements of an extent required for kissing events between widely separated loci take place during interphase.…”