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Genetic Evidence for the Role of the Vacuole in Supplying Secretory Organelles with Ca2+ in Hansenula polymorpha.

Fokina AV, Chechenova MB, Karginov AV, Ter-Avanesyan MD, Agaphonov MO - PLoS ONE (2015)

Bottom Line: The ret1-27 mutation also exerted phenotypes indicating alterations in transport between the vacuole and secretory organelles.These data indicate that ret1-27, pmc1 and vps35 affect a previously unknown Pmr1-independent route of the Ca2+ delivery to the secretory pathway.We also observed that the vacuolar protein carboxypeptidase Y receives additional modifications of its glycoside chains if it escapes the Vps10-dependent sorting to the vacuole.

View Article: PubMed Central - PubMed

Affiliation: A.N. Bach Institute of Biochemistry, Research Center of Biotechnology of the Russian Academy of Sciences, Moscow, Russia.

ABSTRACT
Processes taking place in the secretory organelles require Ca2+ and Mn2+, which in yeast are supplied by the Pmr1 ion pump. Here we observed that in the yeast Hansenula polymorpha Ca2+ deficiency in the secretory pathway caused by Pmr1 inactivation is exacerbated by (i) the ret1-27 mutation affecting COPI-mediated vesicular transport, (ii) inactivation of the vacuolar Ca2+ ATPase Pmc1 and (iii) inactivation of Vps35, which is a component of the retromer complex responsible for protein transport between the vacuole and secretory organelles. The ret1-27 mutation also exerted phenotypes indicating alterations in transport between the vacuole and secretory organelles. These data indicate that ret1-27, pmc1 and vps35 affect a previously unknown Pmr1-independent route of the Ca2+ delivery to the secretory pathway. We also observed that the vacuolar protein carboxypeptidase Y receives additional modifications of its glycoside chains if it escapes the Vps10-dependent sorting to the vacuole.

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Effect of the vps35-Δ mutation on growth of strains with or without the PMR1 gene.Cell suspensions with equal densities were spotted onto corresponding media and grown for 2 days. The experiment was performed serial dilutions of cell suspensions (S6 Fig) and a representative dilution is shown in this figure. pmr1-Δ vps35-Δ, the 1MA27/12/GP1-Δvps35 strain lacking the plasmid with PMR1; vps35-Δ, the 1MA27/12/GP1-Δvps35 strain, pmr1-Δ VPS35, the 1MA27/12/GP1 strain lacking the plasmid with PMR1; PMR1 VPS35, the 1MA27/12/GP1 strain.
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pone.0145915.g004: Effect of the vps35-Δ mutation on growth of strains with or without the PMR1 gene.Cell suspensions with equal densities were spotted onto corresponding media and grown for 2 days. The experiment was performed serial dilutions of cell suspensions (S6 Fig) and a representative dilution is shown in this figure. pmr1-Δ vps35-Δ, the 1MA27/12/GP1-Δvps35 strain lacking the plasmid with PMR1; vps35-Δ, the 1MA27/12/GP1-Δvps35 strain, pmr1-Δ VPS35, the 1MA27/12/GP1 strain lacking the plasmid with PMR1; PMR1 VPS35, the 1MA27/12/GP1 strain.

Mentions: The role of the vacuole in supplying the secretory organelles with Ca2+ also followed from effects of the inactivation of Vps35, which is a component of the retromer complex responsible for the retrograde trafficking from the prevacuolar compartments to the Golgi apparatus. Similarly to ret1-27 ([18] and Fig 2), the vps35-Δ mutation led to hypersensitivity to Ca2+ shortage in culture medium and exacerbated Ca2+ dependence of the pmr1-Δ mutant (Fig 4). Indeed, the growth of the vps35-Δ mutant was abolished by supplementing of SD* with 20 mM EGTA, while the strain with the wild-type VPS35 allele still could grow. The vps35-Δ effect on Ca2+ dependence of the pmr1-Δ mutant was less pronounced then the effect of ret1-27, since the vps35-Δ pmr1-Δ double mutant was able to grow on regular SD and YPD media. The exacerbation of the pmr1-Δ dependence on Ca2+ by the vps35-Δ mutation indicated involvement of pre-vacuolar compartments in a Pmr1-independent supply of the secretory organelles with Ca2+. At the same time the vps35-Δ mutation exerted hypersensitivity to Mn2+, which masked the ability of Mn2+ to suppress the growth defect caused by the pmr1-Δ mutation. We speculate that Vps35 is involved in degradation of the plasma membrane Mn2+ transporter and thus the loss of Vps35 may increase Mn2+ uptake.


Genetic Evidence for the Role of the Vacuole in Supplying Secretory Organelles with Ca2+ in Hansenula polymorpha.

Fokina AV, Chechenova MB, Karginov AV, Ter-Avanesyan MD, Agaphonov MO - PLoS ONE (2015)

Effect of the vps35-Δ mutation on growth of strains with or without the PMR1 gene.Cell suspensions with equal densities were spotted onto corresponding media and grown for 2 days. The experiment was performed serial dilutions of cell suspensions (S6 Fig) and a representative dilution is shown in this figure. pmr1-Δ vps35-Δ, the 1MA27/12/GP1-Δvps35 strain lacking the plasmid with PMR1; vps35-Δ, the 1MA27/12/GP1-Δvps35 strain, pmr1-Δ VPS35, the 1MA27/12/GP1 strain lacking the plasmid with PMR1; PMR1 VPS35, the 1MA27/12/GP1 strain.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4696657&req=5

pone.0145915.g004: Effect of the vps35-Δ mutation on growth of strains with or without the PMR1 gene.Cell suspensions with equal densities were spotted onto corresponding media and grown for 2 days. The experiment was performed serial dilutions of cell suspensions (S6 Fig) and a representative dilution is shown in this figure. pmr1-Δ vps35-Δ, the 1MA27/12/GP1-Δvps35 strain lacking the plasmid with PMR1; vps35-Δ, the 1MA27/12/GP1-Δvps35 strain, pmr1-Δ VPS35, the 1MA27/12/GP1 strain lacking the plasmid with PMR1; PMR1 VPS35, the 1MA27/12/GP1 strain.
Mentions: The role of the vacuole in supplying the secretory organelles with Ca2+ also followed from effects of the inactivation of Vps35, which is a component of the retromer complex responsible for the retrograde trafficking from the prevacuolar compartments to the Golgi apparatus. Similarly to ret1-27 ([18] and Fig 2), the vps35-Δ mutation led to hypersensitivity to Ca2+ shortage in culture medium and exacerbated Ca2+ dependence of the pmr1-Δ mutant (Fig 4). Indeed, the growth of the vps35-Δ mutant was abolished by supplementing of SD* with 20 mM EGTA, while the strain with the wild-type VPS35 allele still could grow. The vps35-Δ effect on Ca2+ dependence of the pmr1-Δ mutant was less pronounced then the effect of ret1-27, since the vps35-Δ pmr1-Δ double mutant was able to grow on regular SD and YPD media. The exacerbation of the pmr1-Δ dependence on Ca2+ by the vps35-Δ mutation indicated involvement of pre-vacuolar compartments in a Pmr1-independent supply of the secretory organelles with Ca2+. At the same time the vps35-Δ mutation exerted hypersensitivity to Mn2+, which masked the ability of Mn2+ to suppress the growth defect caused by the pmr1-Δ mutation. We speculate that Vps35 is involved in degradation of the plasma membrane Mn2+ transporter and thus the loss of Vps35 may increase Mn2+ uptake.

Bottom Line: The ret1-27 mutation also exerted phenotypes indicating alterations in transport between the vacuole and secretory organelles.These data indicate that ret1-27, pmc1 and vps35 affect a previously unknown Pmr1-independent route of the Ca2+ delivery to the secretory pathway.We also observed that the vacuolar protein carboxypeptidase Y receives additional modifications of its glycoside chains if it escapes the Vps10-dependent sorting to the vacuole.

View Article: PubMed Central - PubMed

Affiliation: A.N. Bach Institute of Biochemistry, Research Center of Biotechnology of the Russian Academy of Sciences, Moscow, Russia.

ABSTRACT
Processes taking place in the secretory organelles require Ca2+ and Mn2+, which in yeast are supplied by the Pmr1 ion pump. Here we observed that in the yeast Hansenula polymorpha Ca2+ deficiency in the secretory pathway caused by Pmr1 inactivation is exacerbated by (i) the ret1-27 mutation affecting COPI-mediated vesicular transport, (ii) inactivation of the vacuolar Ca2+ ATPase Pmc1 and (iii) inactivation of Vps35, which is a component of the retromer complex responsible for protein transport between the vacuole and secretory organelles. The ret1-27 mutation also exerted phenotypes indicating alterations in transport between the vacuole and secretory organelles. These data indicate that ret1-27, pmc1 and vps35 affect a previously unknown Pmr1-independent route of the Ca2+ delivery to the secretory pathway. We also observed that the vacuolar protein carboxypeptidase Y receives additional modifications of its glycoside chains if it escapes the Vps10-dependent sorting to the vacuole.

Show MeSH
Related in: MedlinePlus