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Predominantly Cytoplasmic Localization in Yeast of ASR1, a Non-Receptor Transcription Factor from Plants.

Urtasun N, Correa García S, Iusem ND, Bermúdez Moretti M - Open Biochem J (2010)

Bottom Line: It is associated with water-deficit stress and is involved in adaptation to dry climates.For that purpose, we employed an in vivo eukaryotic expression system, the heterologous model Saccharomyces cerevisiae, including wild type strains as well as mutants in which the variant ASR1 previously proved to be functionally protective against osmotic stress.The results are discussed in terms of a plausible dual (cytoplasmic and nuclear) role of ASR proteins.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, Ciudad Universitaria (1428), Buenos Aires, Argentina.

ABSTRACT
The Asr gene family (named after abscisic acid, stress and ripening), currently classified as a novel group of the LEA superfamily, is exclusively present in the genomes of seed plants, except for the Brassicaceae family. It is associated with water-deficit stress and is involved in adaptation to dry climates. Motivated by separate reports depicting ASR proteins as either transcription factors or chaperones, we decided to determine the intracellular localization of ASR proteins. For that purpose, we employed an in vivo eukaryotic expression system, the heterologous model Saccharomyces cerevisiae, including wild type strains as well as mutants in which the variant ASR1 previously proved to be functionally protective against osmotic stress. Our methodology involved immunofluorescence-based confocal microscopy, without artificially altering the native structure of the protein under study. Results show that, in both normal and osmotic stress conditions, recombinant ASR1 turned out to localize mainly to the cytoplasm, irrespective of the genotype used, revealing a scattered distribution in the form of dots or granules. The results are discussed in terms of a plausible dual (cytoplasmic and nuclear) role of ASR proteins.

No MeSH data available.


Related in: MedlinePlus

Intracellular localization of ASR1 in yeast cells. Wild type cells (YPH102 strain) (panels A1-E3) and hog1Δ (JBY13 strain) (F1-I3 panels) transformed with pYES2-Asr1 were incubated in 1% galactose SC medium. After 20 minutes, samples were withdrawn (panels A, E and F) and remaining cells were transferred to 2% glucose fresh SC medium with (panels B, C, G and H) or without (panels D and I) 0.5 M NaCl. Samples were withdrawn at 10 minutes (panels B and G) or 20 minutes (panels C, D, H and I). Nuclei were stained with propidium iodide (panels 1). Cells were processed for indirect immunofluorescence with ASR1-specific antiserum (panels 2), except for panels E where a pre-immune serum was used. Merged images are shown in panels 3.
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Figure 2: Intracellular localization of ASR1 in yeast cells. Wild type cells (YPH102 strain) (panels A1-E3) and hog1Δ (JBY13 strain) (F1-I3 panels) transformed with pYES2-Asr1 were incubated in 1% galactose SC medium. After 20 minutes, samples were withdrawn (panels A, E and F) and remaining cells were transferred to 2% glucose fresh SC medium with (panels B, C, G and H) or without (panels D and I) 0.5 M NaCl. Samples were withdrawn at 10 minutes (panels B and G) or 20 minutes (panels C, D, H and I). Nuclei were stained with propidium iodide (panels 1). Cells were processed for indirect immunofluorescence with ASR1-specific antiserum (panels 2), except for panels E where a pre-immune serum was used. Merged images are shown in panels 3.

Mentions: In normal, non-stressed conditions, ASR1 localized mainly to the cytoplasm (Fig. 2, panels A and E). When cells were subjected to osmotic stress (0.5 M NaCl), ASR1 remained cytoplasmic at least up to 20 minutes (Fig. 2, panels B. C, F and G).


Predominantly Cytoplasmic Localization in Yeast of ASR1, a Non-Receptor Transcription Factor from Plants.

Urtasun N, Correa García S, Iusem ND, Bermúdez Moretti M - Open Biochem J (2010)

Intracellular localization of ASR1 in yeast cells. Wild type cells (YPH102 strain) (panels A1-E3) and hog1Δ (JBY13 strain) (F1-I3 panels) transformed with pYES2-Asr1 were incubated in 1% galactose SC medium. After 20 minutes, samples were withdrawn (panels A, E and F) and remaining cells were transferred to 2% glucose fresh SC medium with (panels B, C, G and H) or without (panels D and I) 0.5 M NaCl. Samples were withdrawn at 10 minutes (panels B and G) or 20 minutes (panels C, D, H and I). Nuclei were stained with propidium iodide (panels 1). Cells were processed for indirect immunofluorescence with ASR1-specific antiserum (panels 2), except for panels E where a pre-immune serum was used. Merged images are shown in panels 3.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2908927&req=5

Figure 2: Intracellular localization of ASR1 in yeast cells. Wild type cells (YPH102 strain) (panels A1-E3) and hog1Δ (JBY13 strain) (F1-I3 panels) transformed with pYES2-Asr1 were incubated in 1% galactose SC medium. After 20 minutes, samples were withdrawn (panels A, E and F) and remaining cells were transferred to 2% glucose fresh SC medium with (panels B, C, G and H) or without (panels D and I) 0.5 M NaCl. Samples were withdrawn at 10 minutes (panels B and G) or 20 minutes (panels C, D, H and I). Nuclei were stained with propidium iodide (panels 1). Cells were processed for indirect immunofluorescence with ASR1-specific antiserum (panels 2), except for panels E where a pre-immune serum was used. Merged images are shown in panels 3.
Mentions: In normal, non-stressed conditions, ASR1 localized mainly to the cytoplasm (Fig. 2, panels A and E). When cells were subjected to osmotic stress (0.5 M NaCl), ASR1 remained cytoplasmic at least up to 20 minutes (Fig. 2, panels B. C, F and G).

Bottom Line: It is associated with water-deficit stress and is involved in adaptation to dry climates.For that purpose, we employed an in vivo eukaryotic expression system, the heterologous model Saccharomyces cerevisiae, including wild type strains as well as mutants in which the variant ASR1 previously proved to be functionally protective against osmotic stress.The results are discussed in terms of a plausible dual (cytoplasmic and nuclear) role of ASR proteins.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, Ciudad Universitaria (1428), Buenos Aires, Argentina.

ABSTRACT
The Asr gene family (named after abscisic acid, stress and ripening), currently classified as a novel group of the LEA superfamily, is exclusively present in the genomes of seed plants, except for the Brassicaceae family. It is associated with water-deficit stress and is involved in adaptation to dry climates. Motivated by separate reports depicting ASR proteins as either transcription factors or chaperones, we decided to determine the intracellular localization of ASR proteins. For that purpose, we employed an in vivo eukaryotic expression system, the heterologous model Saccharomyces cerevisiae, including wild type strains as well as mutants in which the variant ASR1 previously proved to be functionally protective against osmotic stress. Our methodology involved immunofluorescence-based confocal microscopy, without artificially altering the native structure of the protein under study. Results show that, in both normal and osmotic stress conditions, recombinant ASR1 turned out to localize mainly to the cytoplasm, irrespective of the genotype used, revealing a scattered distribution in the form of dots or granules. The results are discussed in terms of a plausible dual (cytoplasmic and nuclear) role of ASR proteins.

No MeSH data available.


Related in: MedlinePlus