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ER-associated retrograde SNAREs and the Dsl1 complex mediate an alternative, Sey1p-independent homotypic ER fusion pathway.

Rogers JV, McMahon C, Baryshnikova A, Hughson FM, Rose MD - Mol. Biol. Cell (2014)

Bottom Line: However, an alternative explanation--that the observed phenotypes arose from perturbed vesicle trafficking--could not be ruled out.In contrast, cytosolic coat protein I (COPI) vesicle coat mutations in sey1Δ cells caused no synthetic defects, excluding perturbed retrograde trafficking as a cause for the previously observed synthetic defects.We conclude that the ER SNAREs and the Dsl1 complex directly mediate Sey1p-independent ER-ER fusion and that, in the absence of both pathways, cell viability depends upon membrane curvature-promoting reticulons.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, Princeton University, Princeton, NJ 08544-1014.

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DSL1 and SEY1 SGAs. (A) Scatterplot of the SGA scores between dsl1ΔL, dsl1ΔE, and the indicated alleles. Genes without an allele designation are deletions. Identical scores would fall on the diagonal line. Alleles listed in purple have similar scores for both Dsl1 alleles (see Materials and Methods for classification metric), alleles listed in blue are classified as dsl1ΔL-specific, and alleles listed in red are classified as dsl1ΔE-specific. A few genes were tested multiple times independently during the SGA experiment (e.g., bre5Δ) and therefore appear multiple times in the scatterplot. (B) As in A, except dsl1ΔE is compared with independently derived sey1Δ SGA data (Costanzo et al., 2010; unpublished data [version 13-04-22], Boone lab). Only alleles with data present for both dsl1ΔE and sey1Δ are plotted. Alleles of genes encoding subunits of the COPI coat complex are shown in red. The strongest sey1Δ-interacting alleles are shown in black.
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Figure 2: DSL1 and SEY1 SGAs. (A) Scatterplot of the SGA scores between dsl1ΔL, dsl1ΔE, and the indicated alleles. Genes without an allele designation are deletions. Identical scores would fall on the diagonal line. Alleles listed in purple have similar scores for both Dsl1 alleles (see Materials and Methods for classification metric), alleles listed in blue are classified as dsl1ΔL-specific, and alleles listed in red are classified as dsl1ΔE-specific. A few genes were tested multiple times independently during the SGA experiment (e.g., bre5Δ) and therefore appear multiple times in the scatterplot. (B) As in A, except dsl1ΔE is compared with independently derived sey1Δ SGA data (Costanzo et al., 2010; unpublished data [version 13-04-22], Boone lab). Only alleles with data present for both dsl1ΔE and sey1Δ are plotted. Alleles of genes encoding subunits of the COPI coat complex are shown in red. The strongest sey1Δ-interacting alleles are shown in black.

Mentions: To identify the most informative genetic interactions, we grouped the interactions into three classes: interactions shared between dsl1ΔE and dsl1ΔL, dsl1ΔL-specific interactions, and dsl1ΔE-specific interactions (Figure 2A; full data set available in Supplemental Table S1; classes determined via a cutoff score and a difference threshold; see Materials and Methods). As expected, the strongest interactions shared between dsl1ΔE and dsl1ΔL consisted of known components of vesicle trafficking. In the dsl1ΔL-specific class, we identified five negative interactions: pse1-41, hir1Δ, mps3-1, chk1Δ, and cse2Δ. Four of these genes have nuclear roles: Pse1p interacts with nuclear pore complexes, Mps3p resides at the half-bridge and mediates spindle pole body formation, Hir1p is a subunit of the histone regulation complex, and Cse2p is a subunit of the RNA polymerase II mediator complex. Interestingly, while pse1-41 is by far the strongest dsl1ΔL-specific negative genetic interaction, pse1-34 is among the strongest dsl1ΔL-specific positive interactions. The final interaction, chk1Δ, uniquely interacts both negatively with dsl1ΔL and positively with dsl1ΔE. Chk1p is a checkpoint kinase that mediates cell cycle arrest.


ER-associated retrograde SNAREs and the Dsl1 complex mediate an alternative, Sey1p-independent homotypic ER fusion pathway.

Rogers JV, McMahon C, Baryshnikova A, Hughson FM, Rose MD - Mol. Biol. Cell (2014)

DSL1 and SEY1 SGAs. (A) Scatterplot of the SGA scores between dsl1ΔL, dsl1ΔE, and the indicated alleles. Genes without an allele designation are deletions. Identical scores would fall on the diagonal line. Alleles listed in purple have similar scores for both Dsl1 alleles (see Materials and Methods for classification metric), alleles listed in blue are classified as dsl1ΔL-specific, and alleles listed in red are classified as dsl1ΔE-specific. A few genes were tested multiple times independently during the SGA experiment (e.g., bre5Δ) and therefore appear multiple times in the scatterplot. (B) As in A, except dsl1ΔE is compared with independently derived sey1Δ SGA data (Costanzo et al., 2010; unpublished data [version 13-04-22], Boone lab). Only alleles with data present for both dsl1ΔE and sey1Δ are plotted. Alleles of genes encoding subunits of the COPI coat complex are shown in red. The strongest sey1Δ-interacting alleles are shown in black.
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Related In: Results  -  Collection

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Figure 2: DSL1 and SEY1 SGAs. (A) Scatterplot of the SGA scores between dsl1ΔL, dsl1ΔE, and the indicated alleles. Genes without an allele designation are deletions. Identical scores would fall on the diagonal line. Alleles listed in purple have similar scores for both Dsl1 alleles (see Materials and Methods for classification metric), alleles listed in blue are classified as dsl1ΔL-specific, and alleles listed in red are classified as dsl1ΔE-specific. A few genes were tested multiple times independently during the SGA experiment (e.g., bre5Δ) and therefore appear multiple times in the scatterplot. (B) As in A, except dsl1ΔE is compared with independently derived sey1Δ SGA data (Costanzo et al., 2010; unpublished data [version 13-04-22], Boone lab). Only alleles with data present for both dsl1ΔE and sey1Δ are plotted. Alleles of genes encoding subunits of the COPI coat complex are shown in red. The strongest sey1Δ-interacting alleles are shown in black.
Mentions: To identify the most informative genetic interactions, we grouped the interactions into three classes: interactions shared between dsl1ΔE and dsl1ΔL, dsl1ΔL-specific interactions, and dsl1ΔE-specific interactions (Figure 2A; full data set available in Supplemental Table S1; classes determined via a cutoff score and a difference threshold; see Materials and Methods). As expected, the strongest interactions shared between dsl1ΔE and dsl1ΔL consisted of known components of vesicle trafficking. In the dsl1ΔL-specific class, we identified five negative interactions: pse1-41, hir1Δ, mps3-1, chk1Δ, and cse2Δ. Four of these genes have nuclear roles: Pse1p interacts with nuclear pore complexes, Mps3p resides at the half-bridge and mediates spindle pole body formation, Hir1p is a subunit of the histone regulation complex, and Cse2p is a subunit of the RNA polymerase II mediator complex. Interestingly, while pse1-41 is by far the strongest dsl1ΔL-specific negative genetic interaction, pse1-34 is among the strongest dsl1ΔL-specific positive interactions. The final interaction, chk1Δ, uniquely interacts both negatively with dsl1ΔL and positively with dsl1ΔE. Chk1p is a checkpoint kinase that mediates cell cycle arrest.

Bottom Line: However, an alternative explanation--that the observed phenotypes arose from perturbed vesicle trafficking--could not be ruled out.In contrast, cytosolic coat protein I (COPI) vesicle coat mutations in sey1Δ cells caused no synthetic defects, excluding perturbed retrograde trafficking as a cause for the previously observed synthetic defects.We conclude that the ER SNAREs and the Dsl1 complex directly mediate Sey1p-independent ER-ER fusion and that, in the absence of both pathways, cell viability depends upon membrane curvature-promoting reticulons.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, Princeton University, Princeton, NJ 08544-1014.

Show MeSH
Related in: MedlinePlus