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Sul1 and Sul2 sulfate transceptors signal to protein kinase A upon exit of sulfur starvation.

Kankipati HN, Rubio-Texeira M, Castermans D, Diallinas G, Thevelein JM - J. Biol. Chem. (2015)

Bottom Line: Overall, our data suggest that transceptors can undergo independent conformational changes, each responsible for triggering different downstream processes.The Sul1 and Sul2 transceptors are the first identified plasma membrane sensors for extracellular sulfate.High affinity transporters induced upon starvation for their substrate may generally act as transceptors during exit from starvation.

View Article: PubMed Central - PubMed

Affiliation: From the Laboratory of Molecular Cell Biology, Institute of Botany and Microbiology, KU Leuven, Kasteelpark Arenberg 31, B-3001 Leuven-Heverlee, Flanders, Belgium, the Department of Molecular Microbiology, VIB, Kasteelpark Arenberg 31, B-3001 Leuven-Heverlee, Flanders, Belgium, and.

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Sulfate transceptors Sul1 and Sul2 are regulated at the transcriptional and post-transcriptional level.A, relative mRNA levels of SUL1 (●) and SUL2 (○) before and after the addition of 3 mm sulfate to sulfur-starved cells. B, uptake rate of [35S]sulfate in Sul1-HA-expressing (●) and Sul2-HA-expressing (○) cells before and after the addition of sulfate to sulfur-starved cells. The cells were harvested at the indicated time points, and the short term uptake rate was then measured with a 1-min uptake assay using radioactive sulfate. C, immunoprecipitation of Sul1-HA and Sul2-HA from membrane-enriched (P13) fractions, isolated before and at different time intervals after the addition of 3 mm sulfate. The immunoprecipitated samples were blotted and immunodetected with anti-HA antibody. The predicted size of the Sul1-HA and Sul2-HA proteins is indicated with an arrow. D, localization of Sul1-HA or Sul2-HA was monitored in the absence of sulfate (−) and after the addition of 3 mm sulfate (+) by immunofluorescence and confocal microscopy imaging. HA-tagged Sul1 and Sul2 proteins were detected by treatment with anti-HA rat primary antibody followed by treatment with Alexa Fluor 488-conjugated anti-rat secondary antibody. DIC, differential interference contrast.
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Figure 7: Sulfate transceptors Sul1 and Sul2 are regulated at the transcriptional and post-transcriptional level.A, relative mRNA levels of SUL1 (●) and SUL2 (○) before and after the addition of 3 mm sulfate to sulfur-starved cells. B, uptake rate of [35S]sulfate in Sul1-HA-expressing (●) and Sul2-HA-expressing (○) cells before and after the addition of sulfate to sulfur-starved cells. The cells were harvested at the indicated time points, and the short term uptake rate was then measured with a 1-min uptake assay using radioactive sulfate. C, immunoprecipitation of Sul1-HA and Sul2-HA from membrane-enriched (P13) fractions, isolated before and at different time intervals after the addition of 3 mm sulfate. The immunoprecipitated samples were blotted and immunodetected with anti-HA antibody. The predicted size of the Sul1-HA and Sul2-HA proteins is indicated with an arrow. D, localization of Sul1-HA or Sul2-HA was monitored in the absence of sulfate (−) and after the addition of 3 mm sulfate (+) by immunofluorescence and confocal microscopy imaging. HA-tagged Sul1 and Sul2 proteins were detected by treatment with anti-HA rat primary antibody followed by treatment with Alexa Fluor 488-conjugated anti-rat secondary antibody. DIC, differential interference contrast.

Mentions: Starvation of yeast cells for sulfur causes strong induction of SUL1 and SUL2 and higher levels of the Sul1 and Sul2 high affinity sulfate transporters at the plasma membrane, whereas the addition of sulfate causes rapid down-regulation of sulfate uptake (20, 25). We confirmed that a strong drop in the mRNA level for both SUL1 and SUL2 takes place shortly after the addition of 3 mm sulfate to cells starved for sulfur, indicating a tight negative regulation of the sulfate transporters at the transcriptional level and/or sudden stimulation of mRNA breakdown upon substrate availability (Fig. 7A). Moreover, upon the readdition of sulfate to sulfur-starved cells, the sulfate uptake rate also decreased rapidly and dramatically over time. In particular, the uptake rate of a sul1Δ sul2Δ strain expressing only Sul1-HA dropped to 60% in only 5 min and then slowly but progressively declined further, suggesting the involvement of two different inactivation processes. The uptake rate of the sul1Δ sul2Δ strain expressing only Sul2-HA dropped much more sharply. It declined to below 20% within 5 min and then remained constant at this low level (Fig. 7B). These observations indicate a tight regulation of sulfate transporting capacity, possibly both by allosteric regulation and intracellular sorting, as well as a differential regulation of Sul1 and Sul2 upon the readdition of the substrate.


Sul1 and Sul2 sulfate transceptors signal to protein kinase A upon exit of sulfur starvation.

Kankipati HN, Rubio-Texeira M, Castermans D, Diallinas G, Thevelein JM - J. Biol. Chem. (2015)

Sulfate transceptors Sul1 and Sul2 are regulated at the transcriptional and post-transcriptional level.A, relative mRNA levels of SUL1 (●) and SUL2 (○) before and after the addition of 3 mm sulfate to sulfur-starved cells. B, uptake rate of [35S]sulfate in Sul1-HA-expressing (●) and Sul2-HA-expressing (○) cells before and after the addition of sulfate to sulfur-starved cells. The cells were harvested at the indicated time points, and the short term uptake rate was then measured with a 1-min uptake assay using radioactive sulfate. C, immunoprecipitation of Sul1-HA and Sul2-HA from membrane-enriched (P13) fractions, isolated before and at different time intervals after the addition of 3 mm sulfate. The immunoprecipitated samples were blotted and immunodetected with anti-HA antibody. The predicted size of the Sul1-HA and Sul2-HA proteins is indicated with an arrow. D, localization of Sul1-HA or Sul2-HA was monitored in the absence of sulfate (−) and after the addition of 3 mm sulfate (+) by immunofluorescence and confocal microscopy imaging. HA-tagged Sul1 and Sul2 proteins were detected by treatment with anti-HA rat primary antibody followed by treatment with Alexa Fluor 488-conjugated anti-rat secondary antibody. DIC, differential interference contrast.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
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Figure 7: Sulfate transceptors Sul1 and Sul2 are regulated at the transcriptional and post-transcriptional level.A, relative mRNA levels of SUL1 (●) and SUL2 (○) before and after the addition of 3 mm sulfate to sulfur-starved cells. B, uptake rate of [35S]sulfate in Sul1-HA-expressing (●) and Sul2-HA-expressing (○) cells before and after the addition of sulfate to sulfur-starved cells. The cells were harvested at the indicated time points, and the short term uptake rate was then measured with a 1-min uptake assay using radioactive sulfate. C, immunoprecipitation of Sul1-HA and Sul2-HA from membrane-enriched (P13) fractions, isolated before and at different time intervals after the addition of 3 mm sulfate. The immunoprecipitated samples were blotted and immunodetected with anti-HA antibody. The predicted size of the Sul1-HA and Sul2-HA proteins is indicated with an arrow. D, localization of Sul1-HA or Sul2-HA was monitored in the absence of sulfate (−) and after the addition of 3 mm sulfate (+) by immunofluorescence and confocal microscopy imaging. HA-tagged Sul1 and Sul2 proteins were detected by treatment with anti-HA rat primary antibody followed by treatment with Alexa Fluor 488-conjugated anti-rat secondary antibody. DIC, differential interference contrast.
Mentions: Starvation of yeast cells for sulfur causes strong induction of SUL1 and SUL2 and higher levels of the Sul1 and Sul2 high affinity sulfate transporters at the plasma membrane, whereas the addition of sulfate causes rapid down-regulation of sulfate uptake (20, 25). We confirmed that a strong drop in the mRNA level for both SUL1 and SUL2 takes place shortly after the addition of 3 mm sulfate to cells starved for sulfur, indicating a tight negative regulation of the sulfate transporters at the transcriptional level and/or sudden stimulation of mRNA breakdown upon substrate availability (Fig. 7A). Moreover, upon the readdition of sulfate to sulfur-starved cells, the sulfate uptake rate also decreased rapidly and dramatically over time. In particular, the uptake rate of a sul1Δ sul2Δ strain expressing only Sul1-HA dropped to 60% in only 5 min and then slowly but progressively declined further, suggesting the involvement of two different inactivation processes. The uptake rate of the sul1Δ sul2Δ strain expressing only Sul2-HA dropped much more sharply. It declined to below 20% within 5 min and then remained constant at this low level (Fig. 7B). These observations indicate a tight regulation of sulfate transporting capacity, possibly both by allosteric regulation and intracellular sorting, as well as a differential regulation of Sul1 and Sul2 upon the readdition of the substrate.

Bottom Line: Overall, our data suggest that transceptors can undergo independent conformational changes, each responsible for triggering different downstream processes.The Sul1 and Sul2 transceptors are the first identified plasma membrane sensors for extracellular sulfate.High affinity transporters induced upon starvation for their substrate may generally act as transceptors during exit from starvation.

View Article: PubMed Central - PubMed

Affiliation: From the Laboratory of Molecular Cell Biology, Institute of Botany and Microbiology, KU Leuven, Kasteelpark Arenberg 31, B-3001 Leuven-Heverlee, Flanders, Belgium, the Department of Molecular Microbiology, VIB, Kasteelpark Arenberg 31, B-3001 Leuven-Heverlee, Flanders, Belgium, and.

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Related in: MedlinePlus