Some new data concerning the mutagenic action of DNA
It is shown that spermatozoa of Drosophila melanogaster can transfer molecules of exogenous DNA (or their fragments) from a solution of this DNA injected into the haemocel of an adult male into the ooplasm of the egg. As a result, mutations induced in the descendants arise both in the paternal and t...
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Інститут молекулярної біології і генетики НАН України
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irk-123456789-1547082019-06-16T01:32:33Z Some new data concerning the mutagenic action of DNA Gershenson, S.M. Alexandrov, Yu.N. It is shown that spermatozoa of Drosophila melanogaster can transfer molecules of exogenous DNA (or their fragments) from a solution of this DNA injected into the haemocel of an adult male into the ooplasm of the egg. As a result, mutations induced in the descendants arise both in the paternal and the maternal chromosomes obtained by them. The mutagenic effect of exogenous DNA has a prolonged character inducing mutations in the descendants during many cell generations after DNA-treatment of their father. The mechanism of such a prolongation of the mutagenic action of DNA is discussed. Выявлено, що сперматозоїди самця Drosophila melanogaster здатні переносити до ооплазми запліднюючого яйця молекули екзогенної ДНК (або їxнi фрагменти) з розчину цієї ДНК, що введений до гемоцелі самця; внаслідок цього індуковані ДНК мутації виникають у нащадків у хромосомах, одержаних як від батька, так i від матеpi. Показано, що мутагенна дія екзогенної ДНК має пролонгований характер i індукує у нащадків мутації протягом ряду клітинни поколінь. Обговорюється вірогідний механізм такої пролонгованої дії. Выявлено, что сперматозоиды самца Drosophila melanogaster способны переносить в ооплазмы оплодотворяющего яйца молекулы экзогенной ДНК (или иx фрагменты) из раствора другой ДНК, введен в гемоцели самца; вследствие этого индуцированные ДНК мутации возникают у потомков в хромосомах, полученных как от отца, так и от матеpи. Показано, что мутагенное действие экзогенной ДНК имеет пролонгированный характер и индуцирует у потомков мутации течение ряда клеточных поколений. обсуждается возможный механизм такой пролонгированного действия. 1994 Article Some new data concerning the mutagenic action of DNA / S.M. Gershenson, Yu.N. Alexandrov // Биополимеры и клетка. — 1994. — Т. 10, № 1. — С. 5-10. — Бібліогр.: 18 назв. — англ. 0233-7657 DOI: http://dx.doi.org/10.7124/bc.00039B http://dspace.nbuv.gov.ua/handle/123456789/154708 577.1 en Биополимеры и клетка Інститут молекулярної біології і генетики НАН України |
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It is shown that spermatozoa of Drosophila melanogaster can transfer molecules of exogenous DNA (or their fragments) from a solution of this DNA injected into the haemocel of an adult male into the ooplasm of the egg. As a result, mutations induced in the descendants arise both in the paternal and the maternal chromosomes obtained by them. The mutagenic effect of exogenous DNA has a prolonged character inducing mutations in the descendants during many cell generations after DNA-treatment of their father. The mechanism of such a prolongation of the mutagenic action of DNA is discussed. |
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Gershenson, S.M. Alexandrov, Yu.N. |
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Gershenson, S.M. Alexandrov, Yu.N. Some new data concerning the mutagenic action of DNA Биополимеры и клетка |
| author_facet |
Gershenson, S.M. Alexandrov, Yu.N. |
| author_sort |
Gershenson, S.M. |
| title |
Some new data concerning the mutagenic action of DNA |
| title_short |
Some new data concerning the mutagenic action of DNA |
| title_full |
Some new data concerning the mutagenic action of DNA |
| title_fullStr |
Some new data concerning the mutagenic action of DNA |
| title_full_unstemmed |
Some new data concerning the mutagenic action of DNA |
| title_sort |
some new data concerning the mutagenic action of dna |
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Інститут молекулярної біології і генетики НАН України |
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1994 |
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http://dspace.nbuv.gov.ua/handle/123456789/154708 |
| citation_txt |
Some new data concerning the mutagenic action of DNA / S.M. Gershenson, Yu.N. Alexandrov // Биополимеры и клетка. — 1994. — Т. 10, № 1. — С. 5-10. — Бібліогр.: 18 назв. — англ. |
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Биополимеры и клетка |
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2025-07-14T06:43:54Z |
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2025-07-14T06:43:54Z |
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1837603681208369152 |
| fulltext |
577.1
S. M. Gershenson. Yu. N. Alexandrov
SOME NEW DATA CONCERNING
THE MUTAGENIC ACTION OF DNA
// is shown that spermatozoa of Drosophila melanogaster can transfer molecules of exo
genous DNA (or their fragments) from a solution of this DNA injected into the haemo-
cel of an adult male into the ooplasm of the egg. As a result, mutations induced in the
descendants arise both in the paternal and the maternal chromosomes obtained by them.
The mutagenic effect of exogenous DNA has a prolonged character inducing mutations
in the descendants during many cell generations after DNA-treatment of their father.
The mechanism of such a prolongation of the mutagenic action of DNA is discussed.
In our previous experiments [1, 2] on the induction of recessive lethal
mutations in the 2nd chromosome of Drosophila melanogaster by injec
tions of a solution of exogenous DNA into the haemocel of adult males
we found that the frequency of induced lethal in the progeny which deve
loped from eggs laid by the female fertilized by a DNA-treated male du
ring the first three days after copulation was in most cases only slightly
different from the frequency of such mutations in the progeny which de
veloped from eggs laid later. This surprised us as it meant that DNA
can induce mutations not only in the dividing premeiotic germ cells of
the males but also in their completely formed ripe spermatozoa. This was
later confirmed by the results of an experiment in which DNA-treated
males were immediately after their first copulation separated from the
females with which they have copulated, so that we could be certain that
the eggs laid by the females were fertilized by spermatozoa which were
in contact with exogenous DNA already being ripe. The frequency of
lethals induced by DNA was here again significantly higher than in the
untreated control.
In seems utterly unbelievable that very large DNA molecules present
in the solution injected into the haemocel of the male can penetrate into
the tightly compressed nucleus forming the head of the spermatozoan.
Much more probable is that DNA molecules are adsorbed on the surface
of the spermatozoan and thus are purely mechanically transported and
introduced into the ooplasm of the fertilized egg where they reach both.
the male and female pronuclei and eventually the paternal and maternal
chromosomes in them.
If this supposition is correct then exogenous DNA injected into the
male should induce mutations in chromosomes received by the descen
dants both form their treated father and their untreated mother. In fa
vour of this supposition speaks the experiment described in 1971 by Brac-
kett et al. [3]; it showed that mammalian spermatozoa can introduce into
the egg DNA molecules adsorbed by them from the surrounding fluid.
Recently this has been confirmed by Lavitrano et al. [4] and by Siracuso
et al. [5]; their experiments showed, for example, that murine spermato
zoa, incubated in an isotopic buffer containing an exogenous DNA (the
plasmid pSV2 CAT) are introduced into eggs fertilized by them. Embry
os thus obtained were implanted into pseudo-pregnant females and among
250 descendants of these females about a third contained in their genomic
DNA nucleotide sequences of the above-mentioned plasmid. And when
© S. M. Gershenson, Yu. N. /Uexandrov, 1994
ISSN 0233-7657. БИОПОЛИМЕРЫ И КЛЕТКА. 1994. Т. 10. № 2 5
transgenic females were crossed with CD1 males the gene CAT was ex
pressed in the bones and tails of their descendants. In all, these authors
had proved that such a transport of exogenous DNA into the ooplasm
can be achieved by spermatozoa of mice, bulls and man; and it is quite
probable ihat this can place in insects.
To check this hypothesis we carried out an experiment using as mar
kers the following mutant genes located in the 2nd chromosome of D.
melanogaster — Curly (Cy), Bristle (Bl), Lobe2 (L2), black (b) and cin
nabar (en). Calf thymus DNA was dissolved in saline and injected (ca.
Ю.07—0.09 micrograms of DNA in 0.25 ml of fluid per male) into the
haemocel of heterozygous males having one of its 2nd chromosomes mar
ked with Cy and Bl and the other with en (Cy Bl/cn). These males were
crossed with virgin heterozygous females having one of its 2nd chromo
somes marked with Cy and the other with b(Cy/b). The Fi from these
crosses consisted of flies including two Curly classes, Cy/cn and Cy/b
which can be easily distinguished one from another. Each of the Fi Cy
Bl/b males was individually crossed with a virgin Cy/L2 female and their
F2 descendants were inbred, this allowing to detect in the F3 recessive
lethal mutations which had arisen in the 2nd chromosome of their untre
ated grandmother (in this case no black flies will be present in the F3).
Likewise, each of the Fi Cy/cn males was individually crossed with a vir
gin Cy/L2 female and their F2 descendants were inbred, this allowing to
jdetect in the F3 recessive lethal mutations which had arise in the 2nd
chromosome of their DNA-treated grandfather (in this case no cinnabar
flies will be present in the F3).
The results of this experiment is presented in Table 1. As shown in
this table, the frequency of recessive lethal mutations in the paternal and
the maternal chromosome is about the same, exceeding about ten times
the frequency of such mutations which spontaneously arose in the untre
ated control. This allows us to be rather certain that the induced mutati
ons arose not in ripe spermatozoa but in the chromosomes of both pro
nuclei (paternal and maternal) contained in the fertilized egg. (A similar
experiment was performed on lethals induced in the 2nd chromosome by
the synthetic polyribonucleotide poly (A, U). This experiment also showen
that the mutagen can be carried by spermatozoa to the female pronucleus.
The number of lethals induced in the paternal (5.73 %f) and the maternal
(4.50 %) chromosome of Fi flies was nearly equal.)
However, as this experiment was carried out on a relatively modest
scale this conclusion is only preliminary and needs a repetition on a lar
ger scale.
In one of the experiments of Fahmy and Fahmy [6].. in which exoge
nous DNA was injected into adult Drosophila males, a statistically sig
nificant increase was observed among Fi females of the mutability of the
garnet locus in the X chromosome received from their mother and not
from the DNA-treated father. These authors assume that here the exo
genous DNA injected into the father was passively transmitted by the
spermatozoa to his daughters. We are inclined to regard this case as
speaking for the correctness of our conclusion based on the data presented
in Table 1.
T a b l e i
Recessive lethal mutations induced in the 2nd chromosome of Drosophila melanogaster
in experiments on transportation of DNA molecules by spermatozoa into the egg
Source of chromosomes
Number of
chromosomes
tested
Number
of induced
lethals
Percentage of lethals
Paternal 204 8 3.9±1.4 and 1 miniature
mutation
Maternal 178 6 3.3±1.34
Control (no treatment) 235 1 0.4±0.4
6 ISSN 0233-7657. БИОПОЛИМЕРЫ И КЛЕТКА. 1994. Т. 10. № 2
Already in our first experiments [7] on the induction of mutations
in D. melanogaster by exogenous DNA it was found that the mutagenic
action of DNA is often not immediate but considerably delayed so that
many mutations in the Fi of DNA-treated males appear as mosaic indi
viduals. As was shown by these experiments and numerous later ones,
some gene mutations induced by exogenous DNA added to the food of
D. melanogaster larvae or injected into the haemocel of adult males ap
pear in the Fi as buches of mutants (this showing that the arose in pre-
meiotic germ cells of the male) or as whole (non-mosaic) individuals
but many of them, usually more than 50 per cent, appear as mosaic in
which only 1/2, 1/4 or a lesser part of their body consists of mutant tis
sues. For example, in the Fi of some such experiments among 13 247 flies
27 visible gene mutations were found twenty of which appeared as mosa
ics (chiefly 1/2 or 1/4 mosaics). This group included flies manifesting
phenotypical mosaicism (e. g. with one normal and one miniature wings)
which had mosaic gonads; and also it included some phenotypically whole
mutant (non-mosaic flies which had mosaic gonads [8]). On Table 2 are
shown the results of crossing such Fi mosaic males with virgin attached-X
females. Other mosaic males in this and other similar experiments gave
analogous results. To the 20 mosaic flies in this experimental series a
number of phenotypically mosaic flies probably should be added which
gave no offspring or gave only wild-type descendants and therefore were
not registered as mutants.
A preponderance of mosaic mutants over whole ones was characteris
tic not only of sex-linked visible mutations but also of autosomal ones
•and was observed in our experiments not only in the Fi but in the F2
and F3 as well.
Many Fi mosaics in which «the mutant part was ca 1/4 of the body
|or less must have arisen as a result of mutation which took place two
or more cell generations after the direct treatment of DNA had stopped.
But the mutagenic effect of DNA may be manifested even later. We have
shown in our experiments that the mutagenic effect of DNA is evident
not only in the Fi but also in the F2 and the F3 though in the latter it is
somewhat weaker; so it is still present several dozens of cell generations
after the cessation of the treatment.
In some cases the same gene mutation arose in different generations
of descendants originating from a single treated male. Thus, in one of
our experiments a mosaic miniature male was found in the Fi; this male
was crossed with an attached-X female and some of their sons were mi
niature and other had wild-type wings, this being a result of mosaicism
of the father's gonads. One of the wild-type sons had deformed eyes and
in order to analyse this deformity the male was crossed with an atta
ched-X female. The deformed eye proved to be non-hereditary but among
the offspring of this cross a single mosaic miniature male again appeared
from which a pure miniature stock was later established. A similar pic
ture was observed in the progeny of a mosaic rudimentary male and ru
dimentary mosaics and whole rudimentary males continued to appear un-
till the F6. A pedigree of this family is given in Fig.
In another of our experiments an Fi male was a fused mutant. In
the offspring of its wild-type male brother again a fused mutant was fo-
T a b l e 2
Offspring of mosaic mutants of Drosophila melanogaster; the mutations were induced
by exogenous DNA
Mutation Phenotype of father
Offspring
s? Mutant Wild- type
Miniature Both wings mutant 101 86 24
Miniature One wing mutant 27 4 38
Small-wing Both wings mutant 70 3 25
ISSN 0233-7657. БИОПОЛИМЕРЫ И КЛЕТКА. 1994. Т. 10. № 2 7
und; it was a mosaic both in respect of wings and gonads. Such cases
show that treatment with exogenous DNA leads to the appearance of a
transmittable unstability of certain genes.
In one case the mutant nature of an F{ fly could be ascertained only
because it was a mosaic. This fly, a male, carried a dominant autosomal
mutations which we named Beaded-crossveinless (its phenotype remin
ded both the well known Beaded and the crossveinless mutations of D.
melanogaster). When crossed with an attached-X female it gave a pro
geny consisting of 67 flies among which 9 females and 11 males were
Part of a pedigree in which rudimentary mutations appeared among the descendants
of a mutant male in which the mutation was induced by exogenous DNA
Beaded-crossveinless. Both saxes of these mutants had underdeveloped
gonads and were completely steril. Evidently, their mutant father has
been a mosaic the fertility of which was due to the presence in his body
of normal non-mutant tissues.
We did not study mosaicism among recessive lethal mutations indu
ced by DNA but Mathew [9] and Khan and Alderson [10] used a gene
tic technique which allowed to detect in such experiments not only com
plete (non-mosaic) lethals but mosaic ones as well. The proportion of
•mosaic was here even higher than in our experiments on the induction
of visible mutations. In many treated lines Fi mosaics again produced
mosaics in the F2 and so on, up to F5 (in the experiments of Khan and
Alderson) and even to F9 (in experiments of Mathew). The extension of
the mutagenic effect of DNA to such late generations strongly supports
the supposition made above about a transmittable destabilization by exo
genous DNA of certain genes.
Fahmy and Fahmy [6, 11] found mosaics among visible recessive
mutations and Minutes induced by injection into adult D. melanogaster
8 ISSN 0233-7657. БИОПОЛИМЕРЫ И КЛЕТКА. 1994. Т. 10. № 2
males of DNA isolated from larvae of the same species and from
rat liver.
The data obtained in our previous work the induction of visible and
lethal mutations in D. melanogaster by exogenous DNAs showed that
these mutations in many important aspects closely resemble mutations in
duced by spontaneous insertions into the chromosomes of the recipient
of mobile genetic element of several kinds [12—15] and many other. It
is most unlikely that this parallelism of the peculiarities of the mutagenic
effects of spontaneous insertions of mobile genetic elements and of injec
tions of exogenous DNA is accidental. It seems much more probable that
fragments of molecules of exogenous DNA act like transpositions of mo
bile genetic elements becoming inserted into chromosomes and selecti
vely altering or destabilizing certain genes. If this hypothesis is correct
it explains the prolonged mutagenic action of exogenous DNA. Fragments
of this DNA for a time remain in a free state, like episomes, in the cells
of the recipient and are thus transmitted from one cell generation to
another; or they can be inserted into the DNA of chromosomes this cau
sing gene mutations or inducing minor chromosome rearrangement this
behaviour resembling that of some mobile genetic elements, e. g. copia
(Flavell and Ish-Horowicz [16], Shiba and Saigo [17], Yamafumi et al.
[18] and other authors).
The reversions to wild-type observed by us of some unstable muta
tions induced in Drosophila by exogenous DNA may be caused by an ex
cision of the inserted fragment of foreign DNA or by a change of its ori
entation within the chromosome.
С. М. Гершензон, Ю. М. Александров
ДЕЯК.1 HOBI ДАН1 СТОСОВНО МУТАГЕННОТ ДН ДНК
Р е з ю м е
Выявлено, що сперматозоТди самця Drosophila melanogaster здатш переносити до
ооплазми заплцшюючого яйця молекули екзогенно!" ДНК (або i'xHi фрагменты) з роз-
чину in'e'i ДНК, що введений до гемоцел1 самця; внаслщок цього шдуковаш ДНК му-
тацп виникають у нащадюв у хромосомах, одержаних як в'щ батька, так i В1'д мате-
pi. Показано, що мутагенна д1я екзогенно'1 ДНК мае пролонгований характер i шду-
куе у нащадшв мутацп протягом ряду юптинних поколшь. Обговорюеться в1роп'дний
мехашзм тако!' пролонговано!' да.
REFERENCES
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http://Abs.tr
11. Fahmy О. G., Fahmy M. J. Induction of mutations by deoxyribonucleic acid in Dro-
sophila melanogaster//Ibid.— 1961.— 191, 4790.—P. 776—779.
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инсерционном мутагенезе у дрозофилы // Стабильность и изменчивость генома.—
М. : Наука, 1985.—С. 26—38.
13. Green M. M. Mobile DNA element and spontaneous gene mutations//Eukaryotic
transposable elements as mutagenic agents.— New York : Cold. Spring Harbor Lab.,
1988.—P. 41—50.
14. Rubin G. M. Dispersed repetitive DNA in Drosophila//Mobile Genetic Elements.—
New York: Acad, press, 1983.—P. 329—361.
15. Terracol R. Transcription of rDNA insertions in bobbed mutants of Drosophila me
lanogaster J/ Genet. Res.—1986.—48, N 3.—P. 167—174.
16. Flavell A. J., Ish-Horowicz D. The origin of extrachromosomal circular elements//
Cell.— 1986.—34.—P. 415—427.
17. Shibo Т., Saigo V. Retrovirus-like particles containing RNA-homologous to transpo
sable elements copia in Drosophila melanogaster 11 Nature.— 1983.— 302.— P. 119—124.
18. Yamafumi E., Shiba T. et al. The nucleotide sequence in сорш-related DNA in Dro
sophila virus particles//Ibid.—1985.—316.—P. 771—776.
Institute of Plant Physiology and Genetics, 20.10.93
Academy of Sciences of the Ukraine, Kiev
10 ISSN 0233-7657. БИОПОЛИМЕРЫ И КЛЕТКА. 1994. Т. 10. № 2
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