Dedifferentiation of Asparagus officinalis L. mesophyll cells during initiation of cell cultures

Paul, E. M. M.; Harikrishna, K.; Fioroni, O.; Draper, John

doi:10.1016/0168-9452(89)90213-6

Cited by 16 publications

(12 citation statements)

References 12 publications

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“…The AoPR7 promoter is the first 'intracellular' PR gene promoter to be studied in transgenic plants and shown to be inducible by wounding and pathogen invasion as well as expressed in mature pollen. It is hoped that mRNA from mechanically isolated asparagus cells (Harikrishna er a/., 1991;Paul et al, 1989) and IPCR technology may be applied to the isolation and study of many more monocotyledonous wound-induced genes.…”

Section: The Aoprl-gus Fusion Protein Is Not Secretedmentioning

confidence: 99%

Isolation of an asparagus intracellular PR gene (AoPR1) wound‐responsive promoter by the inverse polymerase chain reaction and its characterization in transgenic tobacco

1993

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SummaryThe Asparagus officinalis intracellular PR1 (AoPRl) gene is expressed in response to wounding and pathogen attack. We utilized the inverse polymerase chain reaction (IPCR) to isolate the cis-acting regulatory sequences of the AoPRl gene following unsuccessful attempts to identify hybridizing clones in genomic libraries. Sequence analysis of two IPCR products revealed that a 347 bp intron was present in the AoPRl gene and that it was probable that the AoPR7 regulatory sequence had been amplified. To test the AoPRl cis acting sequences for biological function a translational fusion was constructed with the p-glucuronidase (GUS) reporter gene and tested in tobacco. These data demonstrated that sequences 982 bp from the probable start of transcription are sufficient to direct woundinducible transcription and that there is no signal peptide encoded by the first 31 residues of the predicted AoPRl protein. Histochemical localization of GUS activity in transgenic tobacco demonstrated strong activity localized to wound and pathogen invasion sites. GUS activity was also found in mature pollen grains.

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Section: The Aoprl-gus Fusion Protein Is Not Secretedmentioning

confidence: 99%

Isolation of an asparagus intracellular PR gene (AoPR1) wound‐responsive promoter by the inverse polymerase chain reaction and its characterization in transgenic tobacco

1993

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“…To solve this problem, we have used a primary culture system of mesophyll cells prepared from Asparagus officinalis L. Mechanically dispersed asparagus cells have relatively high viability and can vigorously proliferate in liquid medium (6,7).…”

Section: Tyr(so3h)-ile-tyr(so3h)-thr-oh) Psk-ac and Psk-p8 Werementioning

confidence: 99%

“…The precipitated single cells were washed with sterile water three times and used in the experiments. From 0.5 g of fresh cladodes, _ 107 cells were obtained.The basal medium used for cell culture, which contains 1.0 g of glutamine per liter, 10 g of sucrose per liter, 30 g of mannitol per liter, 1.0 mg of 1-naphthaleneacetic acid per liter, 0.3 mg of 6-benzyladenine per liter, inorganic salts, and trace elements, was prepared according to Paul et al (7). Dispersed single mesophyll cells were suspended in sterilized water, adjusted to twice the final cell density with the hemacytometer, and dispensed into 24-well microplates at a volume of 250 ,ul per well.…”

mentioning

confidence: 99%

Phytosulfokine, sulfated peptides that induce the proliferation of single mesophyll cells of Asparagus officinalis L.

Matsubayashi

Sakagami

1996

Proc. Natl. Acad. Sci. U.S.A.

437

293

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Proliferation of dispersed plant cells in culture is strictly dependent on cell density, and cells in a low-density culture can only grow in the presence of conditioned medium (CM). No known plant hormones have been able to substitute for CM. To quantify the mitogenic activity of CM, we examined conditions for the assay system using mechanically dispersed mesophyll cells ofAsparagus officinalis L. and established a highly sensitive bioassay method. By use of this method, the mitogenic activity of CM prepared from asparagus cells was characterized: it was heat-stable, susceptible to pronase digestion, and resistant to glycosidase treatment. On the basis of these results, the mitogenic activity in CM was purified 107-fold by column chromatography, and two factors named phytosulfokine-a and -13 (PSK-a and PSK-f8) were obtained. By amino acid sequence analysis and mass spectrometry, the structures of these two factors were determined to be sulfated pentapeptide (H-Tyr(SO3H)-IleTyr(SO3H)-Thr-Gln-OH) and sulfated tetrapeptide (HTyr(SO3H)-Ile-Tyr(SO3H)-Thr-OH). PSK-ac and PSK-p8 were prepared by chemical synthesis and enzymatic sulfation. The synthetic peptides exhibited the same activity as the natural factors, confirming the structure for PSK-a and PSK-p mentioned above. This is the first elucidation of the structure of a conditioned medium factor required for the growth of low-density plant cell cultures.The proliferation of plant cells in dispersed culture is strictly dependent on the initial cell density, and mitotic activity in low-density suspension cell cultures cannot be stimulated by supplementation with known plant hormones or defined nutrients. However, proliferation of plant cells at low density is induced by the addition of conditioned medium (CM) prepared from rapidly growing cells in culture (1). A possible explanation for this phenomenon is that a mitogenic factor or factors produced by the individual cells is essential for cell proliferation.Since the late 1980s, detection and characterization of mitogenic factors that are often called conditioned medium factors (CMFs) have been performed in a few culture systems: highly hydrophilic, neutral compounds in CM of maize cell line (2, 3); highly hydrophilic, relatively heat-stable compounds in CM of a carrot cell line (4); and pronase-resistant, relatively small compounds in CM of Pinus radiata cells (5) (Takii Shubyo, Japan) were planted on moist sterile soil and kept in a growth room at 25 ± 2°C under a daily 16-h light period [Toshiba (Tokyo) DR 400/T and Mitsubishi (Yokohama) MLRBOC 400 F-U lamps; -20,000 lux at plant level]. The cladodes of 40-to 60-day-old spears were used as the experimental material. Cell lines of Zea mays L. and Oriza sativa L. were maintained with weekly subculturing in Murashige and Skoog medium supplemented with 1.0 mg of 2,4-dichlorophenoxyacetic acid per liter and 30 g of sucrose per liter. Cultures were incubated at 25°C in the dark, with rotary shaking at 120 rpm.Bioassay. Approximately 0.5 g of cladodes were steri...

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“…Cell Isolation, Tissue Culture, Microscopic Observations, and Oxygen Evolution/Uptake Measurements Asparagus officinalis cells were mechanically isolated, cultured in the dark on sucrose-containing media, and cytologically examined as described previously by Paul et al (22). At appropriate time points, the contents of a Petri dish (4 x 106 viable starting cells in 10 mL) were pelleted, 6 mL of supernatant was removed, and the cells were resuspended in the remaining 4 mL of asparagus culture medium.…”

Section: Methodsmentioning

confidence: 99%

“…Mechanically isolated cells have been used in various studies of cellular physiology. For example, cells from asparagus (Asparagus officinalis) have been used to examine nitrogen metabolism (28), amino acid transport (4), photosynthesis (12), photoautotrophism in cultured cells (23), and cellular and molecular changes during dedifferentiation (10,22).…”

mentioning

confidence: 99%

Chloroplast Dedifferentiation in Mechanically Isolated Asparagus Cells during Culture Initiation

Harikrishna

Darby²,

Draper³

1992

Plant Physiol.

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Mechanically isolated asparagus (Asparagus officinalis) mesophyll cells dedifferentiate and divide when cultured in the dark in a medium containing sucrose. A strong correlation was observed between the onset of cell division and a loss of photosynthetic capacity. For the first 8 to 9 d of culture, there was no change in chloroplast size or morphology. However, following this period, the chloroplasts divided to form smaller proplastid-like structures. The gross chlorophyll content of the cell population did not change, suggesting that the loss of photosynthetic potential was not by senescence. Northern analysis showed that mRNA of the small subunit of ribulose 1,5-bisphosphate carboxylase/oxygenase was undetectable within 1 d postisolation, which was quicker than in dark-treated plants. The mRNA of the large subunit of ribulose 1,5-bisphosphate carboxylase/oxygenase decreased to low levels within 2 d of cell isolation. Both the large and small subunits of ribulose 1,5-bisphosphate carboxylase/oxygenase protein showed a gradual reduction in abundance, falling to basal levels by days 6 to 7, which coincided with the onset of rapid cell division. A similar trend was observed with chloroplast rRNA molecules, which decreased to basal levels by day 6 in culture.Wounding has been demonstrated to cause profound changes in metabolism, gene expression, and cellular morphology (3,6,15,18

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Dedifferentiation of Asparagus officinalis L. mesophyll cells during initiation of cell cultures

Cited by 16 publications

References 12 publications

Isolation of an asparagus intracellular PR gene (AoPR1) wound‐responsive promoter by the inverse polymerase chain reaction and its characterization in transgenic tobacco

Isolation of an asparagus intracellular PR gene (AoPR1) wound‐responsive promoter by the inverse polymerase chain reaction and its characterization in transgenic tobacco

Phytosulfokine, sulfated peptides that induce the proliferation of single mesophyll cells of Asparagus officinalis L.

Chloroplast Dedifferentiation in Mechanically Isolated Asparagus Cells during Culture Initiation

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