Studies on the recovery of crown gall tumor cells

Abstract

Our previous studies have shown that, while a persistent but potentially reversible suppression of the tumorous state appears to be a characteristic feature of the vegetative phase of teratoma shoot growth in the crown gafl disease of plants, a recovery from that state occurs during the reproductive phase. An analysis has now been made of the reproductive process in an attempt to define the precise stage at which recovery occurs. The results of this analysis have shown that diploid somatic cells of teratoma-derived flower parts such as those found in petals and filaments are inherently neoplastic. On the other hand, haploid cells of plants grown from anthers of the same flowers and diploid cells of F1 generation plants grown from teratoma-derived seed have, by generally accepted criteria, recovered from the tumorous state. These findings have been interpreted to mean that the loss of neoplastic properties occurs in crown gall teratoma cells during meiosis rather than during fertilization or later stages of the reproductive process.An analysis of more than 2000 teratoma-derived tumor shoots has shown, moreover, that a recovery from the tumorous state may also occur, although apparently as a very rare event, during the vegetative phase of teratoma shoot growth.Cloned lines of crown gall teratoma tissues of tobacco (Nicotiana tabacum L.) are characterized by a~capacity to organize tumor shoots when grown on a basic culture medium. When such teratoma-derived shoots are isolated and grafted at cambial level into appropriate morphologically distinct stock plants a broad spectrum of responses may be obtained (1). These responses range at the one extreme to a breakdown of the implanted shoot with a resulting development of typical disorganized teratomatous growths, while at the other extreme in this spectrum of examples are found shoots that develop quite normally, some of which ultimately flower and set viable seed. The leaves that develop from such teratoma shoots are composed of all of the specialized cell types found in normal leaves and they appear by all generally accepted criteria to be normal. It has been found, however, that when tissues are isolated from the teratoma-derived leaves and planted on a basic culture medium they again assume their neoplastic properties. formed by the T37 strain of the crown gall bacterium Agrobacterium tumefaciens. Procedures used in these studies for grafting teratoma shoots and for culturing cells have previously been described elsewhere (1). Briefly, shoots produced by teratoma tissue grown in liquid culture were grafted at cambial level to detopped stems of healthy tobacco plants of the morphologically distinguishable Turkish cultivar. Pieces of leaf, flower, and pith tissue derived from grafted teratoma and normal shoots were cultured on the agar-containing basic medium of Linsmaier and Skoog (2). The growth hormones naphthalene acetic acid (NAA) (0.5 mg/liter) and kinetin (6-furfurylaminopurine) (0.5 mg/liter) were added to the medium in certain cases as described in ...

Evidence for a cellular gene with potential oncogenic activity in plants.

“…Evidently, culturing the tissues triggers the persistent expression of the tumor phenotype. Similar conclusions may be drawn from studies of shoots regenerated from cloned lines of tobacco cells transformed with wild-type pTiT37 plasmids (41,42). Although these shoots appear to be composed of tumor cells (43), they often exhibit normal morphology and do not form tumors either spontaneously or in response to wounding.…”

“…Nevertheless, when normal-appearing tissues are cultured, they exhibit the tumor phenotype and form tumors when grafted on the host plant. There is good evidence that this culture-induced change depends on both the epigenetic state of the cell and the hormonal constitution of the culture medium (17,41,42).…”

Revertant seedlings from crown gall tumors retain a portion of the bacterial Ti plasmid DNA sequences

“…Some tissues from shoots regenerated from a teratoma still exhibited tumorous traits (35,36) and retained T-DNA (11,13,37). The parts of this shoot that had gone through meiosis no longer exhibited tumorous traits (36) and lacked detectable T-DNA (11,13,37).…”

“…Normal-appearing plants have been regenerated from both unorganized (31)(32)(33)(34) and teratoma-type (35,36) crown gall tumors. Some tissues from shoots regenerated from a teratoma still exhibited tumorous traits (35,36) and retained T-DNA (11,13,37).…”

A plausible model for reversal of neoplastic transformations in plants based on multiple steady states.

“…Most examples of reversal of the crown gall tumorous states studied (3)(4)(5)(6)(7)(8)(9), rarely capable of rooting and sensitive to surinfection by Agrobacterium, have turned out to be explained by either deletion (10)(11)(12)(13) or methylation of part or the totality of T-DNA sequences (14,15). Hairy root tumors induced by Agrobacterium strains carrying mannopine Ri plasmids can, however, spontaneously regenerate plants, albeit with altered morphology, that still contain and express T-DNA genes (16)(17)(18)(19).…”