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Clustering of multiple specific genes and gene-rich R-bands around SC-35 domains: evidence for local euchromatic neighborhoods.

Shopland LS, Johnson CV, Byron M, McNeil J, Lawrence JB - J. Cell Biol. (2003)

Bottom Line: Certain bands showed extensive contact, often aligning with or encircling an SC-35 domain, whereas others did not.All three gene-rich reverse bands showed this more than the gene-poor Giemsa dark bands, and morphometric analyses demonstrated statistically significant differences.Rather than random reservoirs of splicing factors, or factors accumulated on an individual highly active gene, we propose a model of SC-35 domains as functional centers for a multitude of clustered genes, forming local euchromatic "neighborhoods."

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, University of Massachusetts Medical Center, Worcester, MA 01655, USA.

ABSTRACT
Typically, eukaryotic nuclei contain 10-30 prominent domains (referred to here as SC-35 domains) that are concentrated in mRNA metabolic factors. Here, we show that multiple specific genes cluster around a common SC-35 domain, which contains multiple mRNAs. Nonsyntenic genes are capable of associating with a common domain, but domain "choice" appears random, even for two coordinately expressed genes. Active genes widely separated on different chromosome arms associate with the same domain frequently, assorting randomly into the 3-4 subregions of the chromosome periphery that contact a domain. Most importantly, visualization of six individual chromosome bands showed that large genomic segments ( approximately 5 Mb) have striking differences in organization relative to domains. Certain bands showed extensive contact, often aligning with or encircling an SC-35 domain, whereas others did not. All three gene-rich reverse bands showed this more than the gene-poor Giemsa dark bands, and morphometric analyses demonstrated statistically significant differences. Similarly, late-replicating DNA generally avoids SC-35 domains. These findings suggest a functional rationale for gene clustering in chromosomal bands, which relates to nuclear clustering of genes with SC-35 domains. Rather than random reservoirs of splicing factors, or factors accumulated on an individual highly active gene, we propose a model of SC-35 domains as functional centers for a multitude of clustered genes, forming local euchromatic "neighborhoods."

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Differential organization of early- and later-replicating DNA relative to SC-35 domains. (A) An early S-phase nucleus pulse labeled with BrdU (green) shows the typical pattern of hundreds of small early-replication foci throughout the nuclear interior, excluding the nucleolus. All SC-35 domains (red) are contacted by multiple early-replicating foci, which correspond to gene-rich DNA. Nuclei in mid (B) and late (C) S-phase are identified by replication sites of gene-poor DNA at the nuclear periphery or in large clumps that are more interior but on different focal planes than SC-35 domains. Arrows indicate SC-35 domains enlarged in insets.
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fig5: Differential organization of early- and later-replicating DNA relative to SC-35 domains. (A) An early S-phase nucleus pulse labeled with BrdU (green) shows the typical pattern of hundreds of small early-replication foci throughout the nuclear interior, excluding the nucleolus. All SC-35 domains (red) are contacted by multiple early-replicating foci, which correspond to gene-rich DNA. Nuclei in mid (B) and late (C) S-phase are identified by replication sites of gene-poor DNA at the nuclear periphery or in large clumps that are more interior but on different focal planes than SC-35 domains. Arrows indicate SC-35 domains enlarged in insets.

Mentions: Because we were practically limited in the number of specific R- and G-bands we could examine directly, we expanded our analyses to obtain a general overview of the genome's gene-rich and gene-poor DNA. When cells are pulse labeled with halogenated dNTPs during S-phase, gene-rich early-replicating DNA, typically in R-bands, is detected in hundreds of small foci throughout the nuclear interior (Fig. 5 A, green), whereas the gene-poor middle- and late-replicating DNA (most G-bands) correspond respectively to similar small foci at the nucleolar and nuclear periphery (Fig. 5 B, green) and to ∼30 larger patches at the periphery and in the interior (Fig. 5 C, green; Ferreira et al., 1997). We found that SC-35 domains are rarely contacted by the two later-replicating classes of DNA (Fig. 5, B and C), even those signals within the nuclear interior. In contrast, SC-35 domains are typically surrounded by early-replicating DNA foci (Fig. 5 A, insets), though these foci do not exclusively localize around SC-35 domain peripheries. This is consistent with our findings that only a portion of any given R-band directly abuts SC-35 domains, and the fact that even early-replicating R-band DNA is substantially comprised of intergenic sequences and nonexpressed genes. In addition, some active genes are consistently expressed in the nucleoplasm between the SC-35 domains (Smith et al., 1999).


Clustering of multiple specific genes and gene-rich R-bands around SC-35 domains: evidence for local euchromatic neighborhoods.

Shopland LS, Johnson CV, Byron M, McNeil J, Lawrence JB - J. Cell Biol. (2003)

Differential organization of early- and later-replicating DNA relative to SC-35 domains. (A) An early S-phase nucleus pulse labeled with BrdU (green) shows the typical pattern of hundreds of small early-replication foci throughout the nuclear interior, excluding the nucleolus. All SC-35 domains (red) are contacted by multiple early-replicating foci, which correspond to gene-rich DNA. Nuclei in mid (B) and late (C) S-phase are identified by replication sites of gene-poor DNA at the nuclear periphery or in large clumps that are more interior but on different focal planes than SC-35 domains. Arrows indicate SC-35 domains enlarged in insets.
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Related In: Results  -  Collection

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fig5: Differential organization of early- and later-replicating DNA relative to SC-35 domains. (A) An early S-phase nucleus pulse labeled with BrdU (green) shows the typical pattern of hundreds of small early-replication foci throughout the nuclear interior, excluding the nucleolus. All SC-35 domains (red) are contacted by multiple early-replicating foci, which correspond to gene-rich DNA. Nuclei in mid (B) and late (C) S-phase are identified by replication sites of gene-poor DNA at the nuclear periphery or in large clumps that are more interior but on different focal planes than SC-35 domains. Arrows indicate SC-35 domains enlarged in insets.
Mentions: Because we were practically limited in the number of specific R- and G-bands we could examine directly, we expanded our analyses to obtain a general overview of the genome's gene-rich and gene-poor DNA. When cells are pulse labeled with halogenated dNTPs during S-phase, gene-rich early-replicating DNA, typically in R-bands, is detected in hundreds of small foci throughout the nuclear interior (Fig. 5 A, green), whereas the gene-poor middle- and late-replicating DNA (most G-bands) correspond respectively to similar small foci at the nucleolar and nuclear periphery (Fig. 5 B, green) and to ∼30 larger patches at the periphery and in the interior (Fig. 5 C, green; Ferreira et al., 1997). We found that SC-35 domains are rarely contacted by the two later-replicating classes of DNA (Fig. 5, B and C), even those signals within the nuclear interior. In contrast, SC-35 domains are typically surrounded by early-replicating DNA foci (Fig. 5 A, insets), though these foci do not exclusively localize around SC-35 domain peripheries. This is consistent with our findings that only a portion of any given R-band directly abuts SC-35 domains, and the fact that even early-replicating R-band DNA is substantially comprised of intergenic sequences and nonexpressed genes. In addition, some active genes are consistently expressed in the nucleoplasm between the SC-35 domains (Smith et al., 1999).

Bottom Line: Certain bands showed extensive contact, often aligning with or encircling an SC-35 domain, whereas others did not.All three gene-rich reverse bands showed this more than the gene-poor Giemsa dark bands, and morphometric analyses demonstrated statistically significant differences.Rather than random reservoirs of splicing factors, or factors accumulated on an individual highly active gene, we propose a model of SC-35 domains as functional centers for a multitude of clustered genes, forming local euchromatic "neighborhoods."

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, University of Massachusetts Medical Center, Worcester, MA 01655, USA.

ABSTRACT
Typically, eukaryotic nuclei contain 10-30 prominent domains (referred to here as SC-35 domains) that are concentrated in mRNA metabolic factors. Here, we show that multiple specific genes cluster around a common SC-35 domain, which contains multiple mRNAs. Nonsyntenic genes are capable of associating with a common domain, but domain "choice" appears random, even for two coordinately expressed genes. Active genes widely separated on different chromosome arms associate with the same domain frequently, assorting randomly into the 3-4 subregions of the chromosome periphery that contact a domain. Most importantly, visualization of six individual chromosome bands showed that large genomic segments ( approximately 5 Mb) have striking differences in organization relative to domains. Certain bands showed extensive contact, often aligning with or encircling an SC-35 domain, whereas others did not. All three gene-rich reverse bands showed this more than the gene-poor Giemsa dark bands, and morphometric analyses demonstrated statistically significant differences. Similarly, late-replicating DNA generally avoids SC-35 domains. These findings suggest a functional rationale for gene clustering in chromosomal bands, which relates to nuclear clustering of genes with SC-35 domains. Rather than random reservoirs of splicing factors, or factors accumulated on an individual highly active gene, we propose a model of SC-35 domains as functional centers for a multitude of clustered genes, forming local euchromatic "neighborhoods."

Show MeSH