Limits...
The Nuclear Pore-Associated TREX-2 Complex Employs Mediator to Regulate Gene Expression.

Schneider M, Hellerschmied D, Schubert T, Amlacher S, Vinayachandran V, Reja R, Pugh BF, Clausen T, Köhler A - Cell (2015)

Bottom Line: Transcriptome and phenotypic profiling confirm that TREX-2 and Med31 are functionally interdependent at specific genes.TREX-2 additionally uses its Mediator-interacting surface to regulate mRNA export suggesting a mechanism for coupling transcription initiation and early steps of mRNA processing.Our data provide mechanistic insight into how an NPC-associated adaptor complex accesses the core transcription machinery.

View Article: PubMed Central - PubMed

Affiliation: Max F. Perutz Laboratories, Medical University of Vienna, Vienna Biocenter Campus (VBC), Dr. Bohr-Gasse 9/3, 1030 Vienna, Austria.

Show MeSH
Sac3 Localization and Cell Growth in Mediator Mutants, Related to Figure 6(A) Localization of N-terminally GFP-tagged wild-type Sac3 expressed from endogenous promoter in sac3Δ cells or cells carrying an additional deletion of CDK8 or MED31. Sac3 localizes mainly to the nuclear periphery, where it exhibits a punctate staining pattern that is typical for NPCs and their associated proteins. Scale bar 3μm.(B) Growth analyses of the indicated strains on medium prepared with glucose or galactose and different temperatures. Cell density was normalized and cells were spotted onto plates in 10-fold serial dilutions. Plates were incubated for 2-3 days.
© Copyright Policy - CC BY-NC-ND
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4644235&req=5

figs6: Sac3 Localization and Cell Growth in Mediator Mutants, Related to Figure 6(A) Localization of N-terminally GFP-tagged wild-type Sac3 expressed from endogenous promoter in sac3Δ cells or cells carrying an additional deletion of CDK8 or MED31. Sac3 localizes mainly to the nuclear periphery, where it exhibits a punctate staining pattern that is typical for NPCs and their associated proteins. Scale bar 3μm.(B) Growth analyses of the indicated strains on medium prepared with glucose or galactose and different temperatures. Cell density was normalized and cells were spotted onto plates in 10-fold serial dilutions. Plates were incubated for 2-3 days.

Mentions: Given the role of TREX-2 in promoting the targeting of the highly inducible GAL1 gene to NPCs (“gene gating”) (Cabal et al., 2006), we asked whether the functional interface between TREX-2 and Mediator could be involved. To this end, we monitored the location of GAL1 relative to the nuclear periphery in transcriptionally repressed and activated conditions. Notably, as for sac3Δ cells (Cabal et al., 2006), the sac3 R288D mutation interferes with the repositioning of the activated GAL1 gene to NPCs (Figure 6). Impaired NPC targeting was seen in med31Δ cells, but not in cdk8Δ cells (Figure 6), in which GAL1 repositioning to the periphery occurred normally despite reduced GAL1 mRNA levels (Figure S3A). To test another “gated” model gene, we analyzed the subtelomeric HXK1, which becomes activated and targeted to NPCs by removing glucose (e.g., growth in galactose) (Taddei et al., 2006). Promoter activation did not significantly increase HXK1 relocalization to the periphery in sac3Δ cells (Figure S5A; see Figure S5B for expression levels). In contrast to GAL1 regulation, HXK1 repositioning was impaired by the deletion of both MED31 and CDK8 (Figure S5A). The gene-NPC targeting defects in these mutants are not caused by indirect effects on the NPC anchor element of TREX-2 (Figure 1A), because Sac3 remained properly attached to NPCs in med31Δ and cdk8Δ cells (Figure S6A; see Figure S6B for phenotypes). In sum, Mediator and TREX-2 are both required for NPC-targeting of the highly inducible GAL1 and HXK1 genes. Differences in Cdk8-dependancy likely reflect how Mediator “interprets” a gene-specific context of transcription factors, illustrated by the fact that Hxk2, a HXK1-specific transcriptional repressor, binds to Mediator via Med8 (de la Cera et al., 2002), whereas the GAL1 transcription activator Gal4 interacts with Med17 and Cdk8 (Traven et al., 2006).


The Nuclear Pore-Associated TREX-2 Complex Employs Mediator to Regulate Gene Expression.

Schneider M, Hellerschmied D, Schubert T, Amlacher S, Vinayachandran V, Reja R, Pugh BF, Clausen T, Köhler A - Cell (2015)

Sac3 Localization and Cell Growth in Mediator Mutants, Related to Figure 6(A) Localization of N-terminally GFP-tagged wild-type Sac3 expressed from endogenous promoter in sac3Δ cells or cells carrying an additional deletion of CDK8 or MED31. Sac3 localizes mainly to the nuclear periphery, where it exhibits a punctate staining pattern that is typical for NPCs and their associated proteins. Scale bar 3μm.(B) Growth analyses of the indicated strains on medium prepared with glucose or galactose and different temperatures. Cell density was normalized and cells were spotted onto plates in 10-fold serial dilutions. Plates were incubated for 2-3 days.
© Copyright Policy - CC BY-NC-ND
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4644235&req=5

figs6: Sac3 Localization and Cell Growth in Mediator Mutants, Related to Figure 6(A) Localization of N-terminally GFP-tagged wild-type Sac3 expressed from endogenous promoter in sac3Δ cells or cells carrying an additional deletion of CDK8 or MED31. Sac3 localizes mainly to the nuclear periphery, where it exhibits a punctate staining pattern that is typical for NPCs and their associated proteins. Scale bar 3μm.(B) Growth analyses of the indicated strains on medium prepared with glucose or galactose and different temperatures. Cell density was normalized and cells were spotted onto plates in 10-fold serial dilutions. Plates were incubated for 2-3 days.
Mentions: Given the role of TREX-2 in promoting the targeting of the highly inducible GAL1 gene to NPCs (“gene gating”) (Cabal et al., 2006), we asked whether the functional interface between TREX-2 and Mediator could be involved. To this end, we monitored the location of GAL1 relative to the nuclear periphery in transcriptionally repressed and activated conditions. Notably, as for sac3Δ cells (Cabal et al., 2006), the sac3 R288D mutation interferes with the repositioning of the activated GAL1 gene to NPCs (Figure 6). Impaired NPC targeting was seen in med31Δ cells, but not in cdk8Δ cells (Figure 6), in which GAL1 repositioning to the periphery occurred normally despite reduced GAL1 mRNA levels (Figure S3A). To test another “gated” model gene, we analyzed the subtelomeric HXK1, which becomes activated and targeted to NPCs by removing glucose (e.g., growth in galactose) (Taddei et al., 2006). Promoter activation did not significantly increase HXK1 relocalization to the periphery in sac3Δ cells (Figure S5A; see Figure S5B for expression levels). In contrast to GAL1 regulation, HXK1 repositioning was impaired by the deletion of both MED31 and CDK8 (Figure S5A). The gene-NPC targeting defects in these mutants are not caused by indirect effects on the NPC anchor element of TREX-2 (Figure 1A), because Sac3 remained properly attached to NPCs in med31Δ and cdk8Δ cells (Figure S6A; see Figure S6B for phenotypes). In sum, Mediator and TREX-2 are both required for NPC-targeting of the highly inducible GAL1 and HXK1 genes. Differences in Cdk8-dependancy likely reflect how Mediator “interprets” a gene-specific context of transcription factors, illustrated by the fact that Hxk2, a HXK1-specific transcriptional repressor, binds to Mediator via Med8 (de la Cera et al., 2002), whereas the GAL1 transcription activator Gal4 interacts with Med17 and Cdk8 (Traven et al., 2006).

Bottom Line: Transcriptome and phenotypic profiling confirm that TREX-2 and Med31 are functionally interdependent at specific genes.TREX-2 additionally uses its Mediator-interacting surface to regulate mRNA export suggesting a mechanism for coupling transcription initiation and early steps of mRNA processing.Our data provide mechanistic insight into how an NPC-associated adaptor complex accesses the core transcription machinery.

View Article: PubMed Central - PubMed

Affiliation: Max F. Perutz Laboratories, Medical University of Vienna, Vienna Biocenter Campus (VBC), Dr. Bohr-Gasse 9/3, 1030 Vienna, Austria.

Show MeSH