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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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Dystrophin and MyHC RNA signal positions in reiterated myotube nuclei. Three examples of the diagrams used to analyze 51 nuclei in 15 separate myotubes are shown. Myotubes  were from female 077 myoblasts. M, MyHC RNA; D, dystrophin  RNA. Only one dystrophin RNA track is seen in each nucleus  due to inactivation of one X chromosome. Relative positions of  the RNA signals were found to be quite different among nuclei  of a common myotube.
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Figure 4: Dystrophin and MyHC RNA signal positions in reiterated myotube nuclei. Three examples of the diagrams used to analyze 51 nuclei in 15 separate myotubes are shown. Myotubes were from female 077 myoblasts. M, MyHC RNA; D, dystrophin RNA. Only one dystrophin RNA track is seen in each nucleus due to inactivation of one X chromosome. Relative positions of the RNA signals were found to be quite different among nuclei of a common myotube.

Mentions: The relative positions of MyHC and dystrophin RNA foci were examined to determine whether any association or pattern of spatial arrangement was apparent for these coordinately expressed sequences within reiterated myofiber nuclei. Cultured myofibers most often do not have well-aligned nuclei, hence, for part of our analysis we attempted to focus on nuclei arrayed in single file and apparently aligned relative to the linear axis of the myotube. Cells were probed for MyHC and dystrophin RNA simultaneously, and the relative positions of signals in 51 nuclei from 15 different 077 myotubes were recorded on drawings as represented in Fig. 4. The three nuclear RNA accumulations (two MyHC and one dystrophin) showed highly variable locations relative to one another. Since sometimes only one MyHC RNA focus was detected, we cannot rule out the possibility that the two MyHC alleles associate in a minority of cells. Within the limits of our analysis, neither nuclear RNA accumulation showed precise coordinates or had a clearly preferred pattern of distribution. Although the dystrophin RNA focus tended to be slightly more peripheral than MyHC (data not shown), neither RNA showed the marked peripheral location commonly seen for inactive neurotensin and albumin genes (Xing et al., 1995). Dystrophin RNA was clearly not confined to the peripheral region, from which SC-35 domains are excluded in these cells (Carter et al., 1993).


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)

Dystrophin and MyHC RNA signal positions in reiterated myotube nuclei. Three examples of the diagrams used to analyze 51 nuclei in 15 separate myotubes are shown. Myotubes  were from female 077 myoblasts. M, MyHC RNA; D, dystrophin  RNA. Only one dystrophin RNA track is seen in each nucleus  due to inactivation of one X chromosome. Relative positions of  the RNA signals were found to be quite different among nuclei  of a common myotube.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2132926&req=5

Figure 4: Dystrophin and MyHC RNA signal positions in reiterated myotube nuclei. Three examples of the diagrams used to analyze 51 nuclei in 15 separate myotubes are shown. Myotubes were from female 077 myoblasts. M, MyHC RNA; D, dystrophin RNA. Only one dystrophin RNA track is seen in each nucleus due to inactivation of one X chromosome. Relative positions of the RNA signals were found to be quite different among nuclei of a common myotube.
Mentions: The relative positions of MyHC and dystrophin RNA foci were examined to determine whether any association or pattern of spatial arrangement was apparent for these coordinately expressed sequences within reiterated myofiber nuclei. Cultured myofibers most often do not have well-aligned nuclei, hence, for part of our analysis we attempted to focus on nuclei arrayed in single file and apparently aligned relative to the linear axis of the myotube. Cells were probed for MyHC and dystrophin RNA simultaneously, and the relative positions of signals in 51 nuclei from 15 different 077 myotubes were recorded on drawings as represented in Fig. 4. The three nuclear RNA accumulations (two MyHC and one dystrophin) showed highly variable locations relative to one another. Since sometimes only one MyHC RNA focus was detected, we cannot rule out the possibility that the two MyHC alleles associate in a minority of cells. Within the limits of our analysis, neither nuclear RNA accumulation showed precise coordinates or had a clearly preferred pattern of distribution. Although the dystrophin RNA focus tended to be slightly more peripheral than MyHC (data not shown), neither RNA showed the marked peripheral location commonly seen for inactive neurotensin and albumin genes (Xing et al., 1995). Dystrophin RNA was clearly not confined to the peripheral region, from which SC-35 domains are excluded in these cells (Carter et al., 1993).

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