Abstract
Calcium signals are critical for the regulation of polarized growth in many eukaryotic cells, including pollen tubes and neurons. In plants, the regulatory pathways that code and decode Ca(2+) signals are poorly understood. In Arabidopsis thaliana, genetic evidence presented here indicates that pollen tube tip growth involves the redundant activity of two Ca(2+)-dependent protein kinases (CPKs), isoforms CPK17 and -34. Both isoforms appear to target to the plasma membrane, as shown by imaging of CPK17-yellow fluorescent protein (YFP) and CPK34-YFP in growing pollen tubes. Segregation analyses from two independent sets of T-DNA insertion mutants indicate that a double disruption of CPK17 and -34 results in an approximately 350-fold reduction in pollen transmission efficiency. The near sterile phenotype of homozygous double mutants could be rescued through pollen expression of a CPK34-YFP fusion. In contrast, a transgene rescue was blocked by mutations engineered to disrupt the Ca(2+)-activation mechanism of CPK34 (CPK34-YFP-E465A,E500A), providing in vivo evidence linking Ca(2+) activation to a biological function of a CPK. While double mutant pollen tubes displayed normal morphology, relative growth rates for the most rapidly growing tubes were reduced by more than three-fold compared with wild type. In addition, while most mutant tubes appeared to grow far enough to reach ovules, the vast majority (>90%) still failed to locate and fertilize ovules. Together, these results provide genetic evidence that CPKs are essential to pollen fitness, and support a mechanistic model in which CPK17 and -34 transduce Ca(2+) signals to increase the rate of pollen tube tip growth and facilitate a response to tropism cues.
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Claims 8
Claim · model-written summary
CPK17 and CPK34 appeared to target the plasma membrane in growing pollen tubes.
- Taxon
- Arabidopsis thaliana
Arabidopsis thalianae0 - Organ / stage
- growing pollen tubese1
- Gene
- CPK17-yellow fluorescent protein (YFP) and CPK34-YFPe1
- Outcome
- target to the plasma membranee1
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“In Arabidopsis thaliana”
e1“Both isoforms appear to target to the plasma membrane, as shown by imaging of CPK17-yellow fluorescent protein (YFP) and CPK34-YFP in growing pollen tubes”
Claim · model-written summary
Double disruption of CPK17 and CPK34 reduced pollen transmission efficiency approximately 350-fold.
- Taxon
- Arabidopsis thalianaArabidopsis thalianae0
- Treatment
- double disruption of CPK17 and -34e1
- Gene
- CPK17 and -34e1
- Outcome
- pollen transmission efficiencye1
- Direction
- reductione1
- Value
- approximately 350-folde1
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“In Arabidopsis thaliana”
e1“a double disruption of CPK17 and -34 results in an approximately 350-fold reduction in pollen transmission efficiency”
Claim · model-written summary
Pollen expression of CPK34-YFP rescued near sterility of homozygous double mutants.
- Taxon
- Arabidopsis thalianaArabidopsis thalianae0
- Treatment
- pollen expression of a CPK34-YFP fusione1
- Gene
- CPK34-YFPe1
- Outcome
- near sterile phenotypee1
- Direction
- rescuede1
- Comparator
- homozygous double mutantse1
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“In Arabidopsis thaliana”
e1“The near sterile phenotype of homozygous double mutants could be rescued through pollen expression of a CPK34-YFP fusion”
Claim · model-written summary
Mutations disrupting CPK34 calcium activation blocked transgene rescue.
- Taxon
- Arabidopsis thalianaArabidopsis thalianae0
- Treatment
- mutations engineered to disrupt the Ca(2+)-activation mechanism of CPK34e1
- Gene
- CPK34-YFP-E465A,E500Ae1
- Outcome
- transgene rescuee1
- Direction
- blockede1
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“In Arabidopsis thaliana”
e1“a transgene rescue was blocked by mutations engineered to disrupt the Ca(2+)-activation mechanism of CPK34 (CPK34-YFP-E465A,E500A)”
Claim · model-written summary
Double mutant pollen tubes had normal morphology.
- Taxon
- Arabidopsis thalianaArabidopsis thalianae0
- Material
- double mutante1
- Organ / stage
- pollen tubese1
- Outcome
- normal morphologye1
Not reportedDirection
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“In Arabidopsis thaliana”
e1“double mutant pollen tubes displayed normal morphology”
Claim · model-written summary
The fastest growing double mutant pollen tubes grew more than three-fold slower than wild type.
- Taxon
- Arabidopsis thalianaArabidopsis thalianae0
- Outcome
- relative growth ratese1
- Direction
- reducede1
- Value
- more than three-folde1
- Comparator
- wild typee1
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“In Arabidopsis thaliana”
e1“relative growth rates for the most rapidly growing tubes were reduced by more than three-fold compared with wild type”
Claim · model-written summary
Most mutant tubes appeared to reach ovules.
- Taxon
- Arabidopsis thalianaArabidopsis thalianae0
- Organ / stage
- mutant tubese1
- Outcome
- reach ovulese1
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“In Arabidopsis thaliana”
e1“most mutant tubes appeared to grow far enough to reach ovules”
Claim · model-written summary
More than 90% of mutant tubes failed to locate and fertilize ovules.
- Taxon
- Arabidopsis thalianaArabidopsis thalianae0
- Outcome
- failed to locate and fertilize ovulese1
- Value
- >90%e1
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“In Arabidopsis thaliana”
e1“the vast majority (>90%) still failed to locate and fertilize ovules”
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