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Loss of the Arabidopsis thaliana P₄-ATPase ALA3 reduces adaptability to temperature stresses and impairs vegetative, pollen, and ovule development.

McDowell SC, López-Marqués RL, Poulsen LR, Palmgren MG, Harper JF - PLoS ONE (2013)

Bottom Line: We also demonstrate that ala3 mutants have reduced fecundity resulting from a combination of decreased ovule production and pollen tube growth defects.In-vitro pollen tube growth assays showed that ala3 pollen germinated ∼2 h slower than wild-type and had approximately 2-fold reductions in both maximal growth rate and overall length.Together, these results support a model in which ALA3 functions to modify endomembranes in multiple cell types, enabling structural changes, or signaling functions that are critical in plants for normal development and adaptation to varied growth environments.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, University of Nevada, Reno, Nevada, United States of America.

ABSTRACT
Members of the P4 subfamily of P-type ATPases are thought to help create asymmetry in lipid bilayers by flipping specific lipids between the leaflets of a membrane. This asymmetry is believed to be central to the formation of vesicles in the secretory and endocytic pathways. In Arabidopsis thaliana, a P4-ATPase associated with the trans-Golgi network (ALA3) was previously reported to be important for vegetative growth and reproductive success. Here we show that multiple phenotypes for ala3 knockouts are sensitive to growth conditions. For example, ala3 rosette size was observed to be dependent upon both temperature and soil, and varied between 40% and 80% that of wild-type under different conditions. We also demonstrate that ala3 mutants have reduced fecundity resulting from a combination of decreased ovule production and pollen tube growth defects. In-vitro pollen tube growth assays showed that ala3 pollen germinated ∼2 h slower than wild-type and had approximately 2-fold reductions in both maximal growth rate and overall length. In genetic crosses under conditions of hot days and cold nights, pollen fitness was reduced by at least 90-fold; from ∼18% transmission efficiency (unstressed) to less than 0.2% (stressed). Together, these results support a model in which ALA3 functions to modify endomembranes in multiple cell types, enabling structural changes, or signaling functions that are critical in plants for normal development and adaptation to varied growth environments.

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The length of ala3 roots relative to wild-type varies with growth conditions.Seedlings were grown under 24 h fluorescent light on ½×MS media until the longest roots reached the bottom of the plate (∼7 cm). The column labels represent the conditions used in the assays. For experiments testing variations in growth media, plants were all grown at 26°C and media was amended with either 15 mM NH4NO3 (pH 5.7), KOH to adjust media to pH 5.0 or 6.5, or 4.5% sucrose to create an osmotic challenge. Root lengths were normalized to the wild-type mean and average results (±SE) for three independent experiments (n≥19 for all conditions except: n = 9 for 15°C, and n = 6 for 30°C) are presented for ala3-1 (crosshatched bars) and ala3-4 (filled bars). * Significantly different from ala3 root growth at 26° on unmodified media (p<0.05, Welch’s t-test).
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pone-0062577-g003: The length of ala3 roots relative to wild-type varies with growth conditions.Seedlings were grown under 24 h fluorescent light on ½×MS media until the longest roots reached the bottom of the plate (∼7 cm). The column labels represent the conditions used in the assays. For experiments testing variations in growth media, plants were all grown at 26°C and media was amended with either 15 mM NH4NO3 (pH 5.7), KOH to adjust media to pH 5.0 or 6.5, or 4.5% sucrose to create an osmotic challenge. Root lengths were normalized to the wild-type mean and average results (±SE) for three independent experiments (n≥19 for all conditions except: n = 9 for 15°C, and n = 6 for 30°C) are presented for ala3-1 (crosshatched bars) and ala3-4 (filled bars). * Significantly different from ala3 root growth at 26° on unmodified media (p<0.05, Welch’s t-test).

Mentions: To determine if the ala3 root growth phenotype [22], [26] also varies with growth conditions, wild-type and ala3 seedlings were germinated and grown at different temperatures or on modified media. The reduction in ala3 root growth was observed to be strongly dependent upon temperature (Figure 3). The phenotype was the least pronounced at 26°C, with ala3 roots growing 63% as long as wild-type. However, at 30°C or 15°C, ala3 roots were 34% and 10% as long as wild-type, respectively. Prolonged growth at 15°C was lethal to ala3 seedlings, as they did not recover after being returned to 23°C, whereas the wild-type controls were all viable. Conditions other than temperature were also found to exacerbate the root growth phenotype (Figure 3). An additional 10–20% reduction in relative root length was observed for high pH (pH 6.5), low pH (pH 5.0), and high osmolarity (4.5% sucrose).


Loss of the Arabidopsis thaliana P₄-ATPase ALA3 reduces adaptability to temperature stresses and impairs vegetative, pollen, and ovule development.

McDowell SC, López-Marqués RL, Poulsen LR, Palmgren MG, Harper JF - PLoS ONE (2013)

The length of ala3 roots relative to wild-type varies with growth conditions.Seedlings were grown under 24 h fluorescent light on ½×MS media until the longest roots reached the bottom of the plate (∼7 cm). The column labels represent the conditions used in the assays. For experiments testing variations in growth media, plants were all grown at 26°C and media was amended with either 15 mM NH4NO3 (pH 5.7), KOH to adjust media to pH 5.0 or 6.5, or 4.5% sucrose to create an osmotic challenge. Root lengths were normalized to the wild-type mean and average results (±SE) for three independent experiments (n≥19 for all conditions except: n = 9 for 15°C, and n = 6 for 30°C) are presented for ala3-1 (crosshatched bars) and ala3-4 (filled bars). * Significantly different from ala3 root growth at 26° on unmodified media (p<0.05, Welch’s t-test).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3646830&req=5

pone-0062577-g003: The length of ala3 roots relative to wild-type varies with growth conditions.Seedlings were grown under 24 h fluorescent light on ½×MS media until the longest roots reached the bottom of the plate (∼7 cm). The column labels represent the conditions used in the assays. For experiments testing variations in growth media, plants were all grown at 26°C and media was amended with either 15 mM NH4NO3 (pH 5.7), KOH to adjust media to pH 5.0 or 6.5, or 4.5% sucrose to create an osmotic challenge. Root lengths were normalized to the wild-type mean and average results (±SE) for three independent experiments (n≥19 for all conditions except: n = 9 for 15°C, and n = 6 for 30°C) are presented for ala3-1 (crosshatched bars) and ala3-4 (filled bars). * Significantly different from ala3 root growth at 26° on unmodified media (p<0.05, Welch’s t-test).
Mentions: To determine if the ala3 root growth phenotype [22], [26] also varies with growth conditions, wild-type and ala3 seedlings were germinated and grown at different temperatures or on modified media. The reduction in ala3 root growth was observed to be strongly dependent upon temperature (Figure 3). The phenotype was the least pronounced at 26°C, with ala3 roots growing 63% as long as wild-type. However, at 30°C or 15°C, ala3 roots were 34% and 10% as long as wild-type, respectively. Prolonged growth at 15°C was lethal to ala3 seedlings, as they did not recover after being returned to 23°C, whereas the wild-type controls were all viable. Conditions other than temperature were also found to exacerbate the root growth phenotype (Figure 3). An additional 10–20% reduction in relative root length was observed for high pH (pH 6.5), low pH (pH 5.0), and high osmolarity (4.5% sucrose).

Bottom Line: We also demonstrate that ala3 mutants have reduced fecundity resulting from a combination of decreased ovule production and pollen tube growth defects.In-vitro pollen tube growth assays showed that ala3 pollen germinated ∼2 h slower than wild-type and had approximately 2-fold reductions in both maximal growth rate and overall length.Together, these results support a model in which ALA3 functions to modify endomembranes in multiple cell types, enabling structural changes, or signaling functions that are critical in plants for normal development and adaptation to varied growth environments.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, University of Nevada, Reno, Nevada, United States of America.

ABSTRACT
Members of the P4 subfamily of P-type ATPases are thought to help create asymmetry in lipid bilayers by flipping specific lipids between the leaflets of a membrane. This asymmetry is believed to be central to the formation of vesicles in the secretory and endocytic pathways. In Arabidopsis thaliana, a P4-ATPase associated with the trans-Golgi network (ALA3) was previously reported to be important for vegetative growth and reproductive success. Here we show that multiple phenotypes for ala3 knockouts are sensitive to growth conditions. For example, ala3 rosette size was observed to be dependent upon both temperature and soil, and varied between 40% and 80% that of wild-type under different conditions. We also demonstrate that ala3 mutants have reduced fecundity resulting from a combination of decreased ovule production and pollen tube growth defects. In-vitro pollen tube growth assays showed that ala3 pollen germinated ∼2 h slower than wild-type and had approximately 2-fold reductions in both maximal growth rate and overall length. In genetic crosses under conditions of hot days and cold nights, pollen fitness was reduced by at least 90-fold; from ∼18% transmission efficiency (unstressed) to less than 0.2% (stressed). Together, these results support a model in which ALA3 functions to modify endomembranes in multiple cell types, enabling structural changes, or signaling functions that are critical in plants for normal development and adaptation to varied growth environments.

Show MeSH
Related in: MedlinePlus