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The high osmolarity glycerol response (HOG) MAP kinase pathway controls localization of a yeast golgi glycosyltransferase.

Reynolds TB, Hopkins BD, Lyons MR, Graham TR - J. Cell Biol. (1998)

Bottom Line: Biol.We have found that basal signaling through the HOG pathway is required to localize Mnn1-s to the Golgi in standard osmotic conditions.Mutations in HOG1 and LDR1 also perturb localization of intact Mnn1p, resulting in its loss from early Golgi compartments and a concomitant increase of Mnn1p in later Golgi compartments.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, Vanderbilt University, Nashville, Tennessee 37235, USA.

ABSTRACT
The yeast alpha-1,3-mannosyltransferase (Mnn1p) is localized to the Golgi by independent transmembrane and lumenal domain signals. The lumenal domain is localized to the Golgi complex when expressed as a soluble form (Mnn1-s) by exchange of its transmembrane domain for a cleavable signal sequence (Graham, T. R., and V. A. Krasnov. 1995. Mol. Biol. Cell. 6:809-824). Mutants that failed to retain the lumenal domain in the Golgi complex, called lumenal domain retention (ldr) mutants, were isolated by screening mutagenized yeast colonies for those that secreted Mnn1-s. Two genes were identified by this screen, HOG1, a gene encoding a mitogen-activated protein kinase (MAPK) that functions in the high osmolarity glycerol (HOG) pathway, and LDR1. We have found that basal signaling through the HOG pathway is required to localize Mnn1-s to the Golgi in standard osmotic conditions. Mutations in HOG1 and LDR1 also perturb localization of intact Mnn1p, resulting in its loss from early Golgi compartments and a concomitant increase of Mnn1p in later Golgi compartments.

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Mutations in the HOG1 gene do not perturb the transmembrane domain-mediated localization mechanism. Wild-type,  hog1Δ, pbs2Δ, pmr1Δ, hog1-11, and ldr1-2 cells expressing M39I  on a single-copy plasmid (pCM39S) were subjected to a liquid invertase assay (Bankaitis et al., 1986) to quantitate the level of  M39I mislocalized to the cell surface.
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Figure 5: Mutations in the HOG1 gene do not perturb the transmembrane domain-mediated localization mechanism. Wild-type, hog1Δ, pbs2Δ, pmr1Δ, hog1-11, and ldr1-2 cells expressing M39I on a single-copy plasmid (pCM39S) were subjected to a liquid invertase assay (Bankaitis et al., 1986) to quantitate the level of M39I mislocalized to the cell surface.

Mentions: Wild-type, hog1Δ, hog1-11, and ldr1-2 cells were transformed with a single-copy plasmid expressing the M39I fusion protein (Fig. 1 B) and the level of invertase localized to the cell surface was assayed. Wild-type, hog1Δ, and hog1-11 cells expressed the same low percentage of invertase activity at the cell surface (∼6%) indicating that the transmembrane domain mediated localization mechanism was not defective in cells lacking functional Hog1p (Fig. 5). The same was found to be true for cells carrying a allele of PMR1 (Fig. 5). However, ldr1-2 cells mislocalized ∼15% of the invertase activity to the cell surface (Fig. 5). Although a modest increase relative to the wild-type strain, this effect was reproducible, and suggests that the ldr1-2 mutation leads to defects in both the lumenal and transmembrane domain mediated localization mechanisms.


The high osmolarity glycerol response (HOG) MAP kinase pathway controls localization of a yeast golgi glycosyltransferase.

Reynolds TB, Hopkins BD, Lyons MR, Graham TR - J. Cell Biol. (1998)

Mutations in the HOG1 gene do not perturb the transmembrane domain-mediated localization mechanism. Wild-type,  hog1Δ, pbs2Δ, pmr1Δ, hog1-11, and ldr1-2 cells expressing M39I  on a single-copy plasmid (pCM39S) were subjected to a liquid invertase assay (Bankaitis et al., 1986) to quantitate the level of  M39I mislocalized to the cell surface.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2132948&req=5

Figure 5: Mutations in the HOG1 gene do not perturb the transmembrane domain-mediated localization mechanism. Wild-type, hog1Δ, pbs2Δ, pmr1Δ, hog1-11, and ldr1-2 cells expressing M39I on a single-copy plasmid (pCM39S) were subjected to a liquid invertase assay (Bankaitis et al., 1986) to quantitate the level of M39I mislocalized to the cell surface.
Mentions: Wild-type, hog1Δ, hog1-11, and ldr1-2 cells were transformed with a single-copy plasmid expressing the M39I fusion protein (Fig. 1 B) and the level of invertase localized to the cell surface was assayed. Wild-type, hog1Δ, and hog1-11 cells expressed the same low percentage of invertase activity at the cell surface (∼6%) indicating that the transmembrane domain mediated localization mechanism was not defective in cells lacking functional Hog1p (Fig. 5). The same was found to be true for cells carrying a allele of PMR1 (Fig. 5). However, ldr1-2 cells mislocalized ∼15% of the invertase activity to the cell surface (Fig. 5). Although a modest increase relative to the wild-type strain, this effect was reproducible, and suggests that the ldr1-2 mutation leads to defects in both the lumenal and transmembrane domain mediated localization mechanisms.

Bottom Line: Biol.We have found that basal signaling through the HOG pathway is required to localize Mnn1-s to the Golgi in standard osmotic conditions.Mutations in HOG1 and LDR1 also perturb localization of intact Mnn1p, resulting in its loss from early Golgi compartments and a concomitant increase of Mnn1p in later Golgi compartments.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, Vanderbilt University, Nashville, Tennessee 37235, USA.

ABSTRACT
The yeast alpha-1,3-mannosyltransferase (Mnn1p) is localized to the Golgi by independent transmembrane and lumenal domain signals. The lumenal domain is localized to the Golgi complex when expressed as a soluble form (Mnn1-s) by exchange of its transmembrane domain for a cleavable signal sequence (Graham, T. R., and V. A. Krasnov. 1995. Mol. Biol. Cell. 6:809-824). Mutants that failed to retain the lumenal domain in the Golgi complex, called lumenal domain retention (ldr) mutants, were isolated by screening mutagenized yeast colonies for those that secreted Mnn1-s. Two genes were identified by this screen, HOG1, a gene encoding a mitogen-activated protein kinase (MAPK) that functions in the high osmolarity glycerol (HOG) pathway, and LDR1. We have found that basal signaling through the HOG pathway is required to localize Mnn1-s to the Golgi in standard osmotic conditions. Mutations in HOG1 and LDR1 also perturb localization of intact Mnn1p, resulting in its loss from early Golgi compartments and a concomitant increase of Mnn1p in later Golgi compartments.

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