Environmental and Molecular Mutagenesis - Legacy Tobacco ...

154 1989 EMS Abstracts 445
http://legacy.library.ucsf.edu/tid/clb93d00/pdf
NOteS._ , .,; . ROLE OB Ai.TW4~D HEPATIC FOCI IN THE STAGES OF CARCINOGENESIS. n P't
~ Yvonne Dt9t~an, Mark Pyron, Chris Laufer, and Tahir Rlzvi
.
. McArdle Laboratory Research, UmverWof Wisconsin, Madison, Wisconsin 53706
Hepatoaltrcittogettrsis in the rat can be divided into the stages of initiation, promotion, and progression
. Initiadon by a DNA-alkylating chemical results in the appcarance of individual hepatocytes
and their progeny exhibiting altered gene expression of specific tnarker(s) . The stage of promotion is
induced by exo~enous and/or endogenous chemicals, usually non-DNA-reactive, which stimulate
both the re licatton rate and reversibly altered gene expression of clones of initiated cells . However,
different chemical classes of promoting agents induce populations of altered hepatic foci (AHF)
which, in general, exhibit different phenotypic characterls 'ttcs, unique to the class of chemical promoting
agents . Furthetmore, when multiple markers are utilized to score the phenotypes of the
clonally expanded inidated cell populations, all possible phenotypic combinations within the constraints
of the action of the promoting agent itself are represented in the AHF populadons . The stage
of progression, when not induced simultaneously with~ge of initiation, may be heralded by the
appearance of phenotypic heterogeneity within individual AHF in contrast to the phenotyp ic homogeneity
of most or all AHF during the stage of promotion . It is in the stage of progression that karyotypic
changes, proto-oncogene activation, and malignant neoplasia occur either predominantly or
exclusively. Progressor agents capable of inducing this stage of neoplastic development are characterized
ubiquitously by their clastogenic effects . Based on this rtwdel, point mutations and clastogenic
effects appear to play critical but different roles at the sta es of initiation and progression in the
development of hepatic neoplasia respectively. (Supported ~y grants from the National Cancer
Institute: CA-07175, -22484, -43700 and a contract from the National Toxicology Program : ES-3-
5024.)
446
DOUBLE-STRAND GAP PLASMIDS DO NOT ALTER THE RATE OF MUTATION OR GENE
CONVERSION IN Saccharornyres cerrvisiac . MJ . Plewa, S . 7>tylor, A. Kumar and R. Carr. Institute for
Environmental Studies and the Department of Microbiology, University of Illinois, Urbana, 61801 USA .
We are developing a system to locate and recover information from specific mutant sites in eukaryotic
chromosomes . We are studying the mutational spectrum of forward mutation at CANl by the doublestrand
gap repair of a series of yeast shuttle vectors . The advantage of this approach is that the
information copied onto the gapped plasmid occurs after the mutational event on the chromosome . We
constructed a progenitor shuttle vector - pMPS - which contains amp' . E. coli Ori, LEUZ GlNI, and
the yeast 2µ Ori. Yeast that are resistant to canavanine (canl') become sensitive to canavanine after
transformation with pMPS. However, it is important to determine if linear, double-strand gap plasmid can
modify the rate of forward mutation at GlNl on chromosome V or gene conversion within the yeast
genome. Linear pMHIS8 (a vector used in the construction of pMP5) at concentrations of 0, 1, 2 .5. 5
and 10 µg were added to competent yeast cells (strain XY729) under transforming conditions . Forward
mutation at CANI and cell toxicity were measured . There was no difference in the frequency of canl'
cells among the groups with a mean value (t SE) of 39 t 4 .0 mutants per 3.5 x 10' competent cells
plated . The same concentrations of linear pMH15S were added to competent D7 cells and gene conversion
was measured between trpS-12 and apS-27 heteroalleles . Again there was no difference in the gene
conversion frequency at dpS with a mean value of 750 s 20 .3 convertants per 3 .4 x 10' competent cells
plated . These controls indicate that the use of double-strand gap shuttle vectors does not affect the
processes of mutation or gene conversion in competent cells .
447
THE AC'ITVATION OF PROMUTAGENS BY PLANT CELL SYSTEMS. Michael J. Plewa . Institute
for Environmental Studies, University of Illinois, Urbana, IL 61801 USA .
Plant activation is the process by which promutagenic agents are activated into mutagens by plant
systems. Many promutagens are activated by the familiar mammalian microsomal monooxygenase systems
as well as by plants . However, several environmentally important agents are preferentially activated by
plant cells. Plants have become a reservoir for the deposition and accumulation of environmental
xenobiotics . With the widespread use of agricultural chemicals on crop plants and with the global exposure
of plants to pollutants, the possibility that plant-activated agents may be introduced into the human food
chain is a cause of concern . Due to these and other concerns, envitonmentaliy relevant agents should be
evaluated with plant assays. The plant celUmicrobe coincubation assay uses cultured plant cell suspensions
as the activating system and bacteria or yeast cells as the genetic indicator organism . After a treatment
time, the microbes are plated on selective medium. In this way the activation system and the genetic
system can be independently studied. In addition the viability of the plant cells and the microbial cells can
be independently determined so that the toxicity of a test agent can be evaluated . We have employed
cultured tobacco, cotton, carrot, maize and 7Yadescanlia cells to study the activation of test agents and
complex environmental mixtures . In addition to screening, this assay is being used in basic research to
elucidate the biochemical mechanisms of plant activation. Data using model monocyclic and polycyclic
aromatic amines will be used to identify developmental and biochemical pathways involved in plant
activation. This research supported by an OSWR grant and USAF grant N88-AF P-0511 .
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