Expression of the [beta]-Glucuronidase Gene in Pollen of Lily (Lilium longiflorum), Tobacco (Nicotiana tabacum), Nicotiana rustica, and Peony (Paeonia lactiflora) by Particle Bombardment

Nishihara, Masahiro; Ito, Masanori; Tanaka, Ichiro; Kyo, Masaharu; Ono, Kazuhisa; Irifune, Kohei; Morikawa, Hiromichi

doi:10.1104/pp.102.2.357

Cited by 58 publications

(34 citation statements)

References 13 publications

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“…LlANK-GFP fusion gene was put under the control of double-cauliflower mosaic virus (CaMV)-35S promoter, which works moderately in lily pollen (Nishihara et al, 1993). Using an increasing amount of the doubleCaMV-35STGFP DNA, we found that a GFP signal was readily detectable when no less than 5 mg DNA was used per bombardment.…”

Section: Llank Protein Is Associated With Membrane-enclosed Organellementioning

confidence: 99%

An Ankyrin Repeat-Containing Protein, Characterized as a Ubiquitin Ligase, Is Closely Associated with Membrane-Enclosed Organelles and Required for Pollen Germination and Pollen Tube Growth in Lily

et al. 2006

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Exhibiting rapid polarized growth, the pollen tube delivers the male gametes into the ovule for fertilization in higher plants. To get an overall picture of gene expression during pollen germination and pollen tube growth, we profiled the transcription patterns of 1,536 pollen cDNAs from lily (Lilium longiflorum) by microarray. Among those that exhibited significant differential expression, a cDNA named lily ankyrin repeat-containing protein (LlANK) was thoroughly studied. The full-length LlANK cDNA sequence predicts a protein containing five tandem ankyrin repeats and a RING zinc-finger domain. The LlANK protein possesses ubiquitin ligase activity in vitro. RNA blots demonstrated that LlANK transcript is present in mature pollen and its level, interestingly contrary to most pollen mRNAs, up-regulated significantly during pollen germination and pollen tube growth. When fused with green fluorescent protein and transiently expressed in pollen, LlANK was found dominantly associated with membrane-enclosed organelles as well as the generative cell. Overexpression of LlANK, however, led to abnormal growth of the pollen tube. On the other hand, transient silencing of LlANK impaired pollen germination and tube growth. Taken together, these results showed that LlANK is a ubiquitin ligase associated with membrane-enclosed organelles and required for polarized pollen tube growth.

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Section: Llank Protein Is Associated With Membrane-enclosed Organellementioning

confidence: 99%

An Ankyrin Repeat-Containing Protein, Characterized as a Ubiquitin Ligase, Is Closely Associated with Membrane-Enclosed Organelles and Required for Pollen Germination and Pollen Tube Growth in Lily

et al. 2006

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“…An important technology that has greatly facilitated the direct analysis of gene regulatory mechanisms in pollen is particle bombardment (Klein et al 1987), which provides a facile and efficient method for delivery of DNA into pollen (Twell et al 1989(Twell et al , 1991b. All currently published studies using this tech nology have been carried out with pollen of angiosperms, including both monocotyledonous and dicotyledonous species (Twell et al 1989, 1991a, b, Hamilton et al 1992, van der Leede-Plegt et al 1992, Nishihara et al 1993, which has led to the identification of conserved regu latory elements among several genes which are prefer entially expressed in maturing pollen (Twell et al 1991b). Thus, established technologies have not been applied to investigate gene regulation in species of gymnosperms, such that the extent to which pollen-expressed genes and their regulatory mechanisms are conserved between angiospermous and gymnospermous species is unknown.…”

Section: Introductionmentioning

confidence: 99%

Analysis of gene regulation in growing pollen tubes of angiosperm and gymnosperm species using microprojectile bombardment

Martinussen¹,

Bate²,

Weterings³

et al. 1995

Physiol Plant

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A microprojectile based transient expression assay was used to investigate the func tional conservation of gene regulatory mechanisms in the male gametophytes of an angiosperm (.Nicotiana tabacum) and two gymnospermous (Picea abies and Pinus pinaster) species. The activities of two angiosperm gene promoters, which have previously shown to be either preferentially expressed in the male gametophyte (lat52) or highly expressed in both the sporophyte and male gametophyte (A ct\), were analysed. The results showed that in P. abies and P. : pinaster, activity of the Actl promoter was significantly higher than the activity of the lat52 promoter, while the converse was observed in N. tabacum. Detailed analysis of Iat52 5' promoter deletions demonstrated that although the minimal -67 bp lat52 core promoter was active at low levels in all three species, upstream regulatory elements conserved among several pollen-expressed genes, including the PBI element, were not functional in P. abies and P. pinaster. These results suggest that both taxa-specific and conserved regulatory mechanisms operate Lo control gene expression during pollen germination and tube growth.

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“…In contrast, in our system, gene transfer by electroporation was performed after protoplast isolation, and the Zm13 promoter was used for gene expression in pollen protoplasts. Electroporation into isolated pollen protoplasts is more efficient than particle bombardment followed by protoplast isolation because the gene transfer efficiency of particle bombardment is much lower (Miyoshi et al 1995;Nishihara et al 1993). Furthermore, many transfected protoplasts are lost during protoplast isolation from pollen grains.…”

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confidence: 99%

“…Lily pollen protoplasts have been widely used for investigating the proton pump, potassium channel, cytosolic calcium, or osmotic water permeability (Gehwolf et al 2002;Griessner and Obermeyer 2003;Shang et al 2005;Sommer et al 2007). For gene transfer, electroporation and polyethylene glycol-mediated transformation can be used for lily pollen protoplasts, whereas particle bombardment is typically used for lily pollen grains (Miyoshi et al 1995;Nishihara et al 1993). In this study, we propose a novel, convenient experimental system for functional analysis of plant aquaporins, exploiting the advantages of lily pollen protoplasts.…”

mentioning

confidence: 99%

“…The gene transfer efficiency was high (53.4%, the average of three individual experiments) in the case of Zm13::GFP, as also reported by Miyoshi et al (1995) (60-70%). In lily pollen, gene transfer efficiency by electroporation is higher than that by particle bombardment, which results in less than 1% gene transfer efficiency (Miyoshi et al 1995;Nishihara et al 1993). This experimental system obtained a large number of transfected protoplasts because of the high (45-60%) gene transfer efficiency attained using electroporation.…”

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Expression of foreign aquaporin genes in lily pollen protoplasts

Yamada

Yanada

Matsuzawa

et al. 2011

Plant Biotechnology

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Aquaporins facilitate water flux across biomembranes in cells and are involved in various physiological phenomena in several plant tissues. Generally, the water-flux activity of exogenously expressed aquaporins is measured in Xenopus laevis oocytes or yeast. However, heterogeneous systems are not likely to be optimal for plant aquaporin analysis. Thus, we created a new experimental system for functional analysis of plant aquaporins using lily (Lilium longiflorum) pollen protoplasts. Large protoplasts with uniform diameters of approximately 95 mm were isolated from lily pollen grains. No plasma membrane intrinsic protein (PIP) aquaporin was detected in lily pollen. For ectopic expression of PIPs in the lily pollen protoplasts, we constructed plasmids in which Arabidopsis AtPIP1;1 or AtPIP2;1 was under the control of a strong pollen-specific promoter (maize Zm13). The PCR-amplified DNA fragments were transformed into the pollen protoplasts by electroporation. Between 45 and 60% of protoplasts were successfully transformed. The protoplasts expressing AtPIP2;1 significantly increased in volume in a hypotonic solution (350 mM mannitol), compared with the vector control. In contrast, the changes in volume of the protoplasts expressing AtPIP1;1 were similar to that of the vector control. This result suggests that PIP2 induces higher water-flux activity in plant cells, whereas PIP1 does not. Thus, we propose the lily pollen protoplast as a simple and useful experimental system to analyze the function of plant aquaporins.

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Expression of the [beta]-Glucuronidase Gene in Pollen of Lily (Lilium longiflorum), Tobacco (Nicotiana tabacum), Nicotiana rustica, and Peony (Paeonia lactiflora) by Particle Bombardment

Cited by 58 publications

References 13 publications

An Ankyrin Repeat-Containing Protein, Characterized as a Ubiquitin Ligase, Is Closely Associated with Membrane-Enclosed Organelles and Required for Pollen Germination and Pollen Tube Growth in Lily

An Ankyrin Repeat-Containing Protein, Characterized as a Ubiquitin Ligase, Is Closely Associated with Membrane-Enclosed Organelles and Required for Pollen Germination and Pollen Tube Growth in Lily

Analysis of gene regulation in growing pollen tubes of angiosperm and gymnosperm species using microprojectile bombardment

Expression of foreign aquaporin genes in lily pollen protoplasts

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