Positional changes of pericentromeric heterochromatin and nucleoli in postmitotic Purkinje cells during murine cerebellum developmentSolovei I.a · Grandi N.a · Knoth R.b · Volk B.b · Cremer T.a
aDepartment of Biology II, Human Genetics, Ludwig Maximilians University (LMU), Munich; bDepartment of Neuropathology, Neurocentre, University of Freiburg, Freiburg (Germany)
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.
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