Intranuclear relocation of the Plc β 3 sequence in cerebellar Purkinje neurons: temporal association with de novo expression during development

Abstract

Interphase nuclei exhibit a cell type-specific topology of chromatin domains. This topology has been proposed to be established at a specific developmental stage and to be associated, in turn, with cell type-specific gene expression. Using murine, cerebellar Purkinje neurons, we have shown previously that the number and the extent of clustering as well as the spatial, intranuclear distribution of centromeric domains change as a function of postnatal development. Specifically, the redistribution of centromeric domains was determined to be associated temporally with major changes in gene expression. Given that centromeric sequences are not transcribed, we tested the hypothesis that the de novo expression of a specific sequence is similarly associated with a change in its spatial, intranuclear position. In Purkinje neurons, Plc beta3 is expressed de novo between postnatal day 2 and 7. In contrast, the level of expression of Rora remains constant throughout development, following its initial expression at embryonic day 15. Plc beta3 and Rora were labeled by fluorescence in situ hybridization within intact nuclei and their intranuclear, spatial positions quantified by confocal microscopy. When analyzed as the distance from the nuclear centroid, the mean fraction of radial distance of Plc beta3 signals changed from 57.3%+/-2.35 (+/-SEM) (n=50) at P3 to 37.9%+/-2.35 (n=50) at P5. In contrast, the mean fraction of the radial distance of Rora signals did not change during postnatal development, remaining at a mean of 60.1%+/-2.01 (n=208) from the nuclear centroid. While the results do not support a causal relationship between the spatial relocation of Plc beta3 and its de novo expression, their temporal association, as described herein, may be taken to support the hypothesis that its intranuclear, spatial positioning may represent one level of transcriptional control.

Cell-type specific proximity of centromeric domains of one homologue each of chromosomes 2 and 11 in nuclei of cerebellar Purkinje neurons

“…The distinct centromere proximity pattern of chromosomes 2 and 11 in Purkinje and granule neurons may indicate specific roles of these centromeres for neuron-specific function. The structural association of centromeres in Purkinje neurons may be associated with the de novo expression of specific gene sequences, as shown previously (Martou et al 2002). This also supports the concept that the centromeres of chromosomes may be sufficiently unique, as previously hypothesized (Karpen and Allshire 1997), and may indicate that these differences are recognized during the chromosomespecific formation of centromeres.…”

“…Fluorescence in situ hybridization (FISH) studies showed that the Purkinje-neuron-specific Plcβ3 sequence is relocated within the nuclear space, temporally coincident with its de novo expression. In contrast, Rorα, a sequence that is not expressed during the same time frame, did not alter its spatial intranuclear position (Martou et al 2002). However, studies of the relative positioning between centromeric domains in neurons have not been reported.…”

Chromobility: the rapid movement of chromosomes in interphase nuclei

“…Individual chromosomes and gene loci have been shown to change their nuclear location upon stimuli, whether that be stimuli to induce differentiation [11,[33][34][35][36], signals that change growth status [10,37,38], environmental stimuli [12] or compounds to induce exogenous gene expression [39]. The consensus hypothesis in the field is that whole chromosomes are moved to allow some control over genes that are housed upon that chromosome, whether for expression or for repression.…”

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

“…Enhanced synaptic input certainly means increasing physiological activity of the developing PCs involving active transcription of PC-specific genes. Martou et al (2002) described an example of intranuclear sequence relocation correlated with onset of gene expression by showing radial relocation of the Plcß3 gene between P3 and P5 coincident with its de novo expression between P2 and P7. Whether changes in higher-order chromatin arrangements in postmitotic PCs during the postnatal development of the cerebellum are a causal necessity or a consequence of changes of the gene expression pattern remains an intriguing question.…”