Abstract
The functional consequences of long-range nuclear reorganization were studied in a cell-by-cell analysis of gene expression and long-range chromosomal interactions in the Drosophila eye and eye imaginal disk. Position-effect variegation was used to stochastically perturb gene expression and probe nuclear reorganization. Variegating genes on rearrangements of Chromosomes X, 2, and 3 were probed for long-range interactions with heterochromatin. Studies were conducted only in tissues known to express the variegating genes. Nuclear structure was revealed by fluorescence in situ hybridization with probes to the variegating gene and heterochromatin. Gene expression was determined alternately by immunofluorescence against specific proteins and by eye pigment autofluorescence. This allowed cell-by-cell comparisons of nuclear architecture between cells in which the variegating gene was either expressed or silenced. Very strong correlations between heterochromatic association and silencing were found. Expressing cells showed a broad distribution of distances between variegating genes and their own centromeric heterochromatin, while silenced cells showed a very tight distribution centered around very short distances, consistent with interaction between the silenced genes and heterochromatin. Spatial and temporal analysis of interactions with heterochromatin indicated that variegating genes primarily associate with heterochromatin in cells that have exited the cell cycle. Differentiation was not a requirement for association, and no differences in association were observed between cell types. Thus, long-range interactions between distal chromosome regions and their own heterochromatin have functional consequences for the organism.
Highlights
From the broad level of the whole chromosome down to the individual gene, interphase chromosomes in every organism studied adhere to common organizational principles
long-range chromsomal interaction (LRCI) Are Strongly Correlated with Gene Silencing Using gene expression as a functional assay, the consequences of LRCIs between chromosomal loci and heterochromatin were probed on a cell-by-cell basis
Silenced cells had a tight distribution of distances centered around 0.5 lm, with virtually all variegating-gene-to-heterochromatin distances being under 1 lm (Figure 2D)
Summary
From the broad level of the whole chromosome down to the individual gene, interphase chromosomes in every organism studied adhere to common organizational principles (reviewed in [1]). An aspect of chromosome structure important for organism function is long-range chromosomal interactions (LRCIs) between distant loci. LRCIs between euchromatic loci and heterochromatin can silence genes (reviewed in [6,7]). LRCIs are not static, for example, the polar organization of Drosophila embryonic chromosomes changes as homologous loci pair and LRCIs within the nucleus form [8,9,10,11]. Mouse immune cells adopt unique contacts between silenced genes and heterochromatin during differentiation and cell fate specification [12,13,14]. These changes appear to be functional rather than merely structural, such that altering LRCIs appears to have profound biological consequences
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