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Processing of endogenous pre-mRNAs in association with SC-35 domains is gene specific.

Smith KP, Moen PT, Wydner KL, Coleman JR, Lawrence JB - J. Cell Biol. (1999)

Bottom Line: These differences do not simply correlate with the complexity, nuclear abundance, or position within overall nuclear space.The distribution of spliceosome assembly factor SC-35 did not simply mirror the distribution of individual pre-mRNAs, but rather suggested that individual domains contain both specific pre-mRNA(s) as well as excess splicing factors.This is consistent with a multifunctional compartment, to which some gene loci and their RNAs have access and others do not.

View Article: PubMed Central - PubMed

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

ABSTRACT
Analysis of six endogenous pre-mRNAs demonstrates that localization at the periphery or within splicing factor-rich (SC-35) domains is not restricted to a few unusually abundant pre-mRNAs, but is apparently a more common paradigm of many protein-coding genes. Different genes are preferentially transcribed and their RNAs processed in different compartments relative to SC-35 domains. These differences do not simply correlate with the complexity, nuclear abundance, or position within overall nuclear space. The distribution of spliceosome assembly factor SC-35 did not simply mirror the distribution of individual pre-mRNAs, but rather suggested that individual domains contain both specific pre-mRNA(s) as well as excess splicing factors. This is consistent with a multifunctional compartment, to which some gene loci and their RNAs have access and others do not. Despite similar molar abundance in muscle fiber nuclei, nascent transcript "trees" of highly complex dystrophin RNA are cotranscriptionally spliced outside of SC-35 domains, whereas posttranscriptional "tracks" of more mature myosin heavy chain transcripts overlap domains. Further analyses supported that endogenous pre-mRNAs exhibit distinct structural organization that may reflect not only the expression and complexity of the gene, but also constraints of its chromosomal context and kinetics of its RNA metabolism.

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Tracks versus trees. (A and B) MyHC RNA forms a  track that moves away from the gene. (A) MyHC RNA foci (red)  form a track with the MyHC gene signal (green, appear yellow in  overlap) at the border. (B) Model showing posttranscriptional  MyHC RNA molecules (red) moving away from the gene (dark  blue). Although gene position relative to SC-35 (light blue) domains is directly demonstrated elsewhere (Moen et al., in preparation) the relationship of the track relative to the domain is consistent with published results for collagen (Xing et al., 1995) and  fits with results presented here. (C and D) Dystrophin RNA  forms a tree around the gene. (C) Dystrophin RNA (green) appears to surround the two foci corresponding to the 5′ and midgene regions of the dystrophin gene (red, appear yellow in overlap). (D) Model indicating the tree of nascent dystrophin  transcripts (green) attached to the dystrophin gene (red and  blue). Dark blue segments refer to the portions of the gene detected with the two different genomic probes.
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Figure 6: Tracks versus trees. (A and B) MyHC RNA forms a track that moves away from the gene. (A) MyHC RNA foci (red) form a track with the MyHC gene signal (green, appear yellow in overlap) at the border. (B) Model showing posttranscriptional MyHC RNA molecules (red) moving away from the gene (dark blue). Although gene position relative to SC-35 (light blue) domains is directly demonstrated elsewhere (Moen et al., in preparation) the relationship of the track relative to the domain is consistent with published results for collagen (Xing et al., 1995) and fits with results presented here. (C and D) Dystrophin RNA forms a tree around the gene. (C) Dystrophin RNA (green) appears to surround the two foci corresponding to the 5′ and midgene regions of the dystrophin gene (red, appear yellow in overlap). (D) Model indicating the tree of nascent dystrophin transcripts (green) attached to the dystrophin gene (red and blue). Dark blue segments refer to the portions of the gene detected with the two different genomic probes.

Mentions: To address this directly, we investigated the spatial configuration of the MyHC and dystrophin genes relative to their respective RNA accumulations using a sequential RNA and DNA hybridization strategy (Xing et al., 1995). MyHC RNA and gene were visualized in two different colors using a 32-kb β-cMyHC–specific genomic probe (see Materials and Methods and Fig. 6, A and B). Dystrophin RNA was visualized by using the 5′ cDNA probe while different segments of the dystrophin gene (∼10–15 kb each) were delineated using the 5′ and midgenomic probes (see Materials and Methods and Fig. 6, C and D).


Processing of endogenous pre-mRNAs in association with SC-35 domains is gene specific.

Smith KP, Moen PT, Wydner KL, Coleman JR, Lawrence JB - J. Cell Biol. (1999)

Tracks versus trees. (A and B) MyHC RNA forms a  track that moves away from the gene. (A) MyHC RNA foci (red)  form a track with the MyHC gene signal (green, appear yellow in  overlap) at the border. (B) Model showing posttranscriptional  MyHC RNA molecules (red) moving away from the gene (dark  blue). Although gene position relative to SC-35 (light blue) domains is directly demonstrated elsewhere (Moen et al., in preparation) the relationship of the track relative to the domain is consistent with published results for collagen (Xing et al., 1995) and  fits with results presented here. (C and D) Dystrophin RNA  forms a tree around the gene. (C) Dystrophin RNA (green) appears to surround the two foci corresponding to the 5′ and midgene regions of the dystrophin gene (red, appear yellow in overlap). (D) Model indicating the tree of nascent dystrophin  transcripts (green) attached to the dystrophin gene (red and  blue). Dark blue segments refer to the portions of the gene detected with the two different genomic probes.
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Related In: Results  -  Collection

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Figure 6: Tracks versus trees. (A and B) MyHC RNA forms a track that moves away from the gene. (A) MyHC RNA foci (red) form a track with the MyHC gene signal (green, appear yellow in overlap) at the border. (B) Model showing posttranscriptional MyHC RNA molecules (red) moving away from the gene (dark blue). Although gene position relative to SC-35 (light blue) domains is directly demonstrated elsewhere (Moen et al., in preparation) the relationship of the track relative to the domain is consistent with published results for collagen (Xing et al., 1995) and fits with results presented here. (C and D) Dystrophin RNA forms a tree around the gene. (C) Dystrophin RNA (green) appears to surround the two foci corresponding to the 5′ and midgene regions of the dystrophin gene (red, appear yellow in overlap). (D) Model indicating the tree of nascent dystrophin transcripts (green) attached to the dystrophin gene (red and blue). Dark blue segments refer to the portions of the gene detected with the two different genomic probes.
Mentions: To address this directly, we investigated the spatial configuration of the MyHC and dystrophin genes relative to their respective RNA accumulations using a sequential RNA and DNA hybridization strategy (Xing et al., 1995). MyHC RNA and gene were visualized in two different colors using a 32-kb β-cMyHC–specific genomic probe (see Materials and Methods and Fig. 6, A and B). Dystrophin RNA was visualized by using the 5′ cDNA probe while different segments of the dystrophin gene (∼10–15 kb each) were delineated using the 5′ and midgenomic probes (see Materials and Methods and Fig. 6, C and D).

Bottom Line: These differences do not simply correlate with the complexity, nuclear abundance, or position within overall nuclear space.The distribution of spliceosome assembly factor SC-35 did not simply mirror the distribution of individual pre-mRNAs, but rather suggested that individual domains contain both specific pre-mRNA(s) as well as excess splicing factors.This is consistent with a multifunctional compartment, to which some gene loci and their RNAs have access and others do not.

View Article: PubMed Central - PubMed

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

ABSTRACT
Analysis of six endogenous pre-mRNAs demonstrates that localization at the periphery or within splicing factor-rich (SC-35) domains is not restricted to a few unusually abundant pre-mRNAs, but is apparently a more common paradigm of many protein-coding genes. Different genes are preferentially transcribed and their RNAs processed in different compartments relative to SC-35 domains. These differences do not simply correlate with the complexity, nuclear abundance, or position within overall nuclear space. The distribution of spliceosome assembly factor SC-35 did not simply mirror the distribution of individual pre-mRNAs, but rather suggested that individual domains contain both specific pre-mRNA(s) as well as excess splicing factors. This is consistent with a multifunctional compartment, to which some gene loci and their RNAs have access and others do not. Despite similar molar abundance in muscle fiber nuclei, nascent transcript "trees" of highly complex dystrophin RNA are cotranscriptionally spliced outside of SC-35 domains, whereas posttranscriptional "tracks" of more mature myosin heavy chain transcripts overlap domains. Further analyses supported that endogenous pre-mRNAs exhibit distinct structural organization that may reflect not only the expression and complexity of the gene, but also constraints of its chromosomal context and kinetics of its RNA metabolism.

Show MeSH
Related in: MedlinePlus