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Yeast homologues of tomosyn and lethal giant larvae function in exocytosis and are associated with the plasma membrane SNARE, Sec9.

Lehman K, Rossi G, Adamo JE, Brennwald P - J. Cell Biol. (1999)

Bottom Line: In contrast to a previous report, we see no defect in actin polarity under conditions where we see a dramatic effect on secretion.Genetic analysis suggests that Sro7 and Sec9 function together in a pathway downstream of the Rho3 GTPase.Taken together, our studies suggest that members of the lethal giant larvae/tomosyn/Sro7 family play an important role in polarized exocytosis by regulating SNARE function on the plasma membrane.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Weill Medical College of Cornell University, New York, New York 10021, USA.

ABSTRACT
We have identified a pair of related yeast proteins, Sro7p and Sro77p, based on their ability to bind to the plasma membrane SNARE (SNARE) protein, Sec9p. These proteins show significant similarity to the Drosophila tumor suppressor, lethal giant larvae and to the neuronal syntaxin-binding protein, tomosyn. SRO7 and SRO77 have redundant functions as loss of both gene products leads to a severe cold-sensitive growth defect that correlates with a severe defect in exocytosis. We show that similar to Sec9, Sro7/77 functions in the docking and fusion of post-Golgi vesicles with the plasma membrane. In contrast to a previous report, we see no defect in actin polarity under conditions where we see a dramatic effect on secretion. This demonstrates that the primary function of Sro7/77, and likely all members of the lethal giant larvae family, is in exocytosis rather than in regulating the actin cytoskeleton. Analysis of the association of Sro7p and Sec9p demonstrates that Sro7p directly interacts with Sec9p both in the cytosol and in the plasma membrane and can associate with Sec9p in the context of a SNAP receptor complex. Genetic analysis suggests that Sro7 and Sec9 function together in a pathway downstream of the Rho3 GTPase. Taken together, our studies suggest that members of the lethal giant larvae/tomosyn/Sro7 family play an important role in polarized exocytosis by regulating SNARE function on the plasma membrane.

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SRO7 and SRO77 form a functionally redundant gene family and the sro7Δ, sro77Δ mutant exhibits a severe cold-sensitive phenotype. (A) Tetrad dissection of a cross between sro7Δ and sro77Δ single disruptants. Tetrads were dissected and grown on YPD plates at 25°C. Pinpoint colonies correspond to haploid segregants containing disruptions of both SRO7 and SRO77. Large colonies correspond to haploid segregants containing wild-type copies of both SRO7 and SRO77 or individual disruptions in either gene. T refers to a tetratype and NPD refers to nonparental ditype segregation patterns observed after scoring for auxotrophic markers for each disruption. (B) Cold sensitivity of the sro7Δ, sro77Δ double-disruptant strain. Each strain of a tetratype tetrad was struck out onto YPD plates and grown at either 37, 25, or 14°C. The genotype of the four strains is as indicated.
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Figure 2: SRO7 and SRO77 form a functionally redundant gene family and the sro7Δ, sro77Δ mutant exhibits a severe cold-sensitive phenotype. (A) Tetrad dissection of a cross between sro7Δ and sro77Δ single disruptants. Tetrads were dissected and grown on YPD plates at 25°C. Pinpoint colonies correspond to haploid segregants containing disruptions of both SRO7 and SRO77. Large colonies correspond to haploid segregants containing wild-type copies of both SRO7 and SRO77 or individual disruptions in either gene. T refers to a tetratype and NPD refers to nonparental ditype segregation patterns observed after scoring for auxotrophic markers for each disruption. (B) Cold sensitivity of the sro7Δ, sro77Δ double-disruptant strain. Each strain of a tetratype tetrad was struck out onto YPD plates and grown at either 37, 25, or 14°C. The genotype of the four strains is as indicated.

Mentions: To examine the effect of loss of Sro7 and Sro77 function in vivo, we disrupted each of the chromosomal copies of these genes individually in a diploid yeast strain. After sporulation and tetrad dissection, there was no observable effect on growth of the resulting haploid disruptants containing either the sro7Δ or sro77Δ allele. However, when sro7 disruptants were crossed with sro77 disruptants, a pronounced growth defect was observed at 25°C in haploid segregants containing both alleles. Therefore, these genes represent a functionally redundant gene family (Fig. 2 A). Growth of these strains at lower temperatures, such as 14 or 19°C (not shown), further enhances this growth defect, whereas growth at 37°C reduces the growth defect, demonstrating that the sro7Δ and sro77Δ double mutants have a severe cold-sensitive growth defect.


Yeast homologues of tomosyn and lethal giant larvae function in exocytosis and are associated with the plasma membrane SNARE, Sec9.

Lehman K, Rossi G, Adamo JE, Brennwald P - J. Cell Biol. (1999)

SRO7 and SRO77 form a functionally redundant gene family and the sro7Δ, sro77Δ mutant exhibits a severe cold-sensitive phenotype. (A) Tetrad dissection of a cross between sro7Δ and sro77Δ single disruptants. Tetrads were dissected and grown on YPD plates at 25°C. Pinpoint colonies correspond to haploid segregants containing disruptions of both SRO7 and SRO77. Large colonies correspond to haploid segregants containing wild-type copies of both SRO7 and SRO77 or individual disruptions in either gene. T refers to a tetratype and NPD refers to nonparental ditype segregation patterns observed after scoring for auxotrophic markers for each disruption. (B) Cold sensitivity of the sro7Δ, sro77Δ double-disruptant strain. Each strain of a tetratype tetrad was struck out onto YPD plates and grown at either 37, 25, or 14°C. The genotype of the four strains is as indicated.
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Related In: Results  -  Collection

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Figure 2: SRO7 and SRO77 form a functionally redundant gene family and the sro7Δ, sro77Δ mutant exhibits a severe cold-sensitive phenotype. (A) Tetrad dissection of a cross between sro7Δ and sro77Δ single disruptants. Tetrads were dissected and grown on YPD plates at 25°C. Pinpoint colonies correspond to haploid segregants containing disruptions of both SRO7 and SRO77. Large colonies correspond to haploid segregants containing wild-type copies of both SRO7 and SRO77 or individual disruptions in either gene. T refers to a tetratype and NPD refers to nonparental ditype segregation patterns observed after scoring for auxotrophic markers for each disruption. (B) Cold sensitivity of the sro7Δ, sro77Δ double-disruptant strain. Each strain of a tetratype tetrad was struck out onto YPD plates and grown at either 37, 25, or 14°C. The genotype of the four strains is as indicated.
Mentions: To examine the effect of loss of Sro7 and Sro77 function in vivo, we disrupted each of the chromosomal copies of these genes individually in a diploid yeast strain. After sporulation and tetrad dissection, there was no observable effect on growth of the resulting haploid disruptants containing either the sro7Δ or sro77Δ allele. However, when sro7 disruptants were crossed with sro77 disruptants, a pronounced growth defect was observed at 25°C in haploid segregants containing both alleles. Therefore, these genes represent a functionally redundant gene family (Fig. 2 A). Growth of these strains at lower temperatures, such as 14 or 19°C (not shown), further enhances this growth defect, whereas growth at 37°C reduces the growth defect, demonstrating that the sro7Δ and sro77Δ double mutants have a severe cold-sensitive growth defect.

Bottom Line: In contrast to a previous report, we see no defect in actin polarity under conditions where we see a dramatic effect on secretion.Genetic analysis suggests that Sro7 and Sec9 function together in a pathway downstream of the Rho3 GTPase.Taken together, our studies suggest that members of the lethal giant larvae/tomosyn/Sro7 family play an important role in polarized exocytosis by regulating SNARE function on the plasma membrane.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Weill Medical College of Cornell University, New York, New York 10021, USA.

ABSTRACT
We have identified a pair of related yeast proteins, Sro7p and Sro77p, based on their ability to bind to the plasma membrane SNARE (SNARE) protein, Sec9p. These proteins show significant similarity to the Drosophila tumor suppressor, lethal giant larvae and to the neuronal syntaxin-binding protein, tomosyn. SRO7 and SRO77 have redundant functions as loss of both gene products leads to a severe cold-sensitive growth defect that correlates with a severe defect in exocytosis. We show that similar to Sec9, Sro7/77 functions in the docking and fusion of post-Golgi vesicles with the plasma membrane. In contrast to a previous report, we see no defect in actin polarity under conditions where we see a dramatic effect on secretion. This demonstrates that the primary function of Sro7/77, and likely all members of the lethal giant larvae family, is in exocytosis rather than in regulating the actin cytoskeleton. Analysis of the association of Sro7p and Sec9p demonstrates that Sro7p directly interacts with Sec9p both in the cytosol and in the plasma membrane and can associate with Sec9p in the context of a SNAP receptor complex. Genetic analysis suggests that Sro7 and Sec9 function together in a pathway downstream of the Rho3 GTPase. Taken together, our studies suggest that members of the lethal giant larvae/tomosyn/Sro7 family play an important role in polarized exocytosis by regulating SNARE function on the plasma membrane.

Show MeSH
Related in: MedlinePlus