Environmental and Molecular Mutagenesis - Legacy Tobacco ...

218
CYTOGENETIC ACTION OF METHYLMERCURY CHLORIDE IN HUMAN LYMPHO-
CYTES,
Helena Groot de Restrepo and Luz A . Carvajal de Gil . Laboratorio de Genbtica
Hurnana, Universidad de los Andes, A .A . 4976, Bogot3, Colombia .
An in vitro study was carried out to analyse the effects of inethylmercury chloride
(MM) in hurnan lymphocyte txaltures . Analyses of cell cycle kinetics, chromosome
aberrations and sister chromatid exchange (SCE) were performed . The results
show a dose effect relationship in tlie cell proliferative processes . No
statistically difference was observed for the frequency of SCE in the cultures exposed
to different concentrations of MM (dose-response curve) non in the exposed
and control cultures (blood from 10 donors) . However the dose-response curve
at one of the concentrations of MM used (1 .26 ug/ml) show a significant increase
of SCE and a higher replication index (RI) . A significant increase in the frequency
of chromatid type aberrations was found in the MM exposed group .
219
INSERTIONAL MUTAGENESIS OF HUMAN CELLS USING RETROVIRUS SHUTTLE VECTORS
Andrew J . Grosovskyt, Linda S . Rosst, Barry W. Glickmanz and Adonis Skandalis2, tUniversity of
California, Riverside, California 92521 and 2York University, Toronto, Ontario M3J 1P3
Insertional mutagenesis is a powerful approach for identifying novel genes with an identifiable phenotype
because genes of interest may be simultaneously inactivated and tagged for cloning as a consequence of the
insertion . Although this strategy has been widely used in prokaryotes and lower eukaryotes, its applicability
in mammalian systems has been limited by the lack of suitable insertion elements . Retrovirus based shuttle
vectors are, however, well suited for this role since they infect a broad range of host cells at high efficiency,
integrate into the host genome at low copy number and at random, and encode selectable markers within the
vector genome.
We present here a characterization of insertional mutagenesis of the human B lymphoblastoid cell line
TK6 following infection with the retrovirus shuttle vecto pZipNeo . TK6 cultures were exposed to pZipNeo
infection for periods ranging from 12 to 72 hours . A population which contained stably integrated provirus
was selected by exposing the infected population to G418 ; G418 resistance is encoded by the neo glne carried
in the vector genome . The infection efficiency in the exposed population, as estimated by determining the
frequency of G418 resistance in a clonal assay, ranged up to 3% in cells exposed for 72 hours. Insertion
mutation frequency was monitored using several endogenous selectable markers available in TK6 cells (aprt,
hpri, tk, Na'/K- ATPase) . An exposure time dependant induction vyas seen for the first three markers .
Induction ranged from 6 to 30 fold following a 72 hour infection period . A small induction was also
observed at the Na'/K" ATPase following 24 hours of infection ; this may be attributable to effects on gene
expression or gene amplification associated with retroviral integration . '
Experiments are now being conducted to determine the percentage of mutants recovered following
retrovirus exposure which are directly attributable to disruption of the coding sequence
by a provirus, and to determine the provirus copy number after various infection periods .
220
G---7CTOXIC :T'_ Or FESTICL0:S N•TD P :J :a'T_^ SY31'»2ii
: . :;, Gt~)v:: .2, School of Life Sciences, 3uru Nanak -'ev University,
q,-,rit ;ar-143005, In3ia .
Several short term assays encompassing all the maior tax>nomic groups
hav' been rsaommen3e3. Although plant 3ystems are being use3
successfull_v by a ntmtber of workers, yet the scepticism about it
Persists . A comparative account of the cp_notnxicity of 20 pestici3es
workz3 out by cytogenetic assays in root meristems and pollen mother
cells in &li_~rn ceDa/Hor3_e_tlm W19 ra , Ln
chromosomal aberration
and microzuclel assays in bone marrow calls in rats an .9 histidine
r=version assa :• in j~1lmonel_l:a_ tyaUmu~j