Differential nuclear scaffold/matrix attachment marks expressed genes

Abstract

It is well established that nuclear architecture plays a key role in poising regions of the genome for transcription. This may be achieved using scaffold/matrix attachment regions (S/MARs) that establish loop domains. However, the relationship between changes in the physical structure of the genome as mediated by attachment to the nuclear scaffold/matrix and gene expression is not clearly understood. To define the role of S/MARs in organizing our genome and to resolve the often contradictory loci-specific studies, we have surveyed the S/MARs in HeLa S3 cells on human chromosomes 14-18 by array comparative genomic hybridization. Comparison of LIS (lithium 3,5-diiodosalicylate) extraction to identify SARs and 2 m NaCl extraction to identify MARs revealed that approximately one-half of the sites were in common. The results presented in this study suggest that SARs 5' of a gene are associated with transcript presence whereas MARs contained within a gene are associated with silenced genes. The varied functions of the S/MARs as revealed by the different extraction methods highlights their unique functional contribution.

Figure 1.

Nuclear Scaffold/Matrix extraction reveals isolation specific differences. (A) Isolation of loop and scaffold/matrix DNA. (B) HeLa log₂ CGH signal ratios of LIS isolated loop:scaffold and NaCl-isolated loop:matrix fractions. The log₂ CGH ratios are indicated as bars along chromosome 16. Loop regions (green) are indicated as positive signals, whereas nuclear scaffold/matrix-associated regions (blue) are represented as negative signals respective of the zero axis. Gene density (orange bars) and an ideogram representative of G-banding were then overlaid along the chromosome. The NaCl-extracted nuclear matrix revealed global chromatin organization such that gene density correlated with looping. It is apparent that the LIS-isolated nuclear scaffold organizes the chromosome in a different manner.

Figure 2.

Spacing of S/MARs along human chromosome 16. The binned distance between S/MARs on human chromosome 16 is shown for nuclear scaffolds/matrices isolated by LIS or NaCl extraction with the peak distance averages for each indicated. The frequency of both SAR and MAR spacing shows a bimodal distribution with two groups of average inferred loop sizes at 558 bp and ∼88 kb for SARs and 762 bp and ∼94 kb for MARs.

Figure 3.

Intergenic nuclear scaffold/matrix attachment and expression. SARs and MARs were mapped according to their distance from both the 5′ and 3′ ends of genes on chromosome 16. Analysis of intergenic S/MAR distance from all genes reveals similar overall distributions from LIS and NaCl extractions.

Figure 4.

LIS and NaCl extractions reveal different profiles relative to gene density/expression. The nuclear scaffold/matrix aCGH profiles in a 30–31 Mb, 16p11.2 gene-rich and 60–61 Mb, 16q21 gene-poor region are shown Log₂ loop enrichment is indicated in green and scaffold/matrix association in blue. Regions identified as S/MARs (black bars) were compared with orange: silent and gray: expressed genes (indicated by asterisks). Gene-dense regions show little, if any nuclear matrix attachment and rather are bound to a LIS extracted nuclear scaffold. Gene-poor regions show significant nuclear matrix attachment with fewer-attachment sites to the LIS-isolated nuclear scaffold.

Figure 5.

SAR and MAR correlations with gene expression. The presence of S/MARs encompassing 20 kb region 5′ and 3′ of a gene, as well as within genes was compared with transcript presence using chi-square analysis (95% CI, 1 df with Yate's correction). The resulting P-values are displayed for each measurement with significance <0.05 indicated in bold. A SAR within 10 kb of the 5′ end region of a gene significantly correlates with gene expression. In contrast a MAR within a gene correlates with silencing of that gene. Direction of transcription is indicated by the arrow.