DNA loop organization in glioblastoma T98G cells at their different functional states
The loop domain organization of chromatin, which plays an important role in transcription regulation, may depend on the cell functional state. Aim. To investigate DNA loop reorganization upon functional transitions in the glioblastoma T98G cells. Methods. Single cell gel electrophoresis (a comet ass...
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| Опубліковано в: : | Вiopolymers and Cell |
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Інститут молекулярної біології і генетики НАН України
2018
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| Цитувати: | DNA loop organization in glioblastoma T98G cells at their different functional states / K.S. Afanasieva, A.Y. Semenova, L.L. Lukash, A.V. Sivolob // Вiopolymers and Cell. — 2018. — Т. 34, № 6. — С. 426-434. — Бібліогр.: 22 назв. — англ. |
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Digital Library of Periodicals of National Academy of Sciences of Ukraine| _version_ | 1859605723666710528 |
|---|---|
| author | Afanasieva, K.S. Semenova, A.Y. Lukash, L.L. Sivolob, A.V. |
| author_facet | Afanasieva, K.S. Semenova, A.Y. Lukash, L.L. Sivolob, A.V. |
| citation_txt | DNA loop organization in glioblastoma T98G cells at their different functional states / K.S. Afanasieva, A.Y. Semenova, L.L. Lukash, A.V. Sivolob // Вiopolymers and Cell. — 2018. — Т. 34, № 6. — С. 426-434. — Бібліогр.: 22 назв. — англ. |
| collection | DSpace DC |
| container_title | Вiopolymers and Cell |
| description | The loop domain organization of chromatin, which plays an important role in transcription regulation, may depend on the cell functional state. Aim. To investigate DNA loop reorganization upon functional transitions in the glioblastoma T98G cells. Methods. Single cell gel electrophoresis (a comet assay) was used to analyze the kinetics of the DNA loop migration from the nucleoids obtained from the lysed cells. Results. The cells arrested in the G1 phase of the cell cycle were characterized by a relatively low amount of DNA in the comet tails due to a low content of DNA in the loops which may be resolved by the comet assay (up to ~300 kb). After cell reactivation, the contour length of the loops essentially increased in parallel with a decrease in the linear loop density along the genome. Conclusions. An increase in the loop size and a respective decrease in the loop density may be a general feature of activated cells as we earlier observed similar effects upon activation of human lymphocytes.
Організація петельних доменів хроматину, яка відіграє важливу роль у регуляції транскрипції, напевно може залежати від функціонального стану клітини. Мета. Дослідити можливу реорганізацію петель ДНК при функціональних переходах у гліобластомних клітинах T98G. Методи. Ми застосовували метод електрофорезу ДНК ізольованих клітин (кометний електрофорез) для аналізу кінетики міграції петель ДНК з нуклеоїдів, отриманих з лізованих клітин. Результати. Клітини, заарештовані на фазі G1 клітинного циклу, характеризуються порівняно низьким вмістом ДНК у хвостах комет внаслідок низького вмісту ДНК у складі петель, що знаходяться у межах роздільної здатності кометного електрофорезу (до ~300 кб). Після реактивації клітин контурна довжина петель суттєво зростає, паралельно зі зниженням лінійної щільності петель уздовж геному. Висновки. Оскільки подібні ефекти спостерігались нами раніше для активованих лімфоцитів, ми робимо висновок, що зростання розміру петель та відповідне зниження їхньої лінійної щільності може бути загальною характеристикою активованих клітин.
Организация петельных доменов хроматина, играющая важную роль в регуляции транскрипции, предположительно может зависеть от функционального состояния клетки. Цель. Исследовать возможной реорганизации петель ДНК при функциональных переходах в глиобластомных клетках T98G. Методы. Мы использовали электрофорез ДНК изолированных клеток (кометный электрофорез) для анализа кинетики миграции петель ДНК из нуклеоидов, полученных из лизированных клеток. Результаты. Клетки, арестованные на фазе G1 клеточного цикла, характеризуются сравнительно низким содержанием ДНК в хвостах комет из-за низкого содержания ДНК в составе петель, которые находятся в пределах разрешающей способности кометного электрофореза (до ~300 кб). После реактивации клеток контурная длина петель существенно возрастает, параллельно со снижением линейной плотности петель вдоль генома. Выводы. Поскольку аналогичные эффекты наблюдались нами ранее для активированных лимфоцитов, мы заключили, что возрастание размера петель и соответствующее снижение их линейной плотности может быть общей характеристикой активированных клеток.
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| first_indexed | 2025-11-28T03:50:04Z |
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K. S. Afanasieva, A. Y. Semenova, L. L. Lukash
© 2018 K. S. Afanasieva et al.; Published by the Institute of Molecular Biology and Genetics, NAS of Ukraine on behalf of Bio-
polymers and Cell. This is an Open Access article distributed under the terms of the Creative Commons Attribution License
(http://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium,
provided the original work is properly cited
UDC 577.323:576.08
DNA loop organization in glioblastoma T98G cells at their different
functional states
K. S. Afanasieva1, A. Y. Semenova1, L. L. Lukash2, A. V. Sivolob1
1 ESC "Institute of Biology and Medicine", Taras Shevchenko National University of Kyiv
64/13, Volodymyrska Str., Kyiv, Ukraine, 01601
2 Institute of Molecular Biology and Genetics, NAS of Ukraine
150, Akademika Zabolotnoho Str., Kyiv, Ukraine, 03143
aphon@ukr.net
The loop domain organization of chromatin, which plays an important role in transcription
regulation, may depend on the cell functional state. The aim of this work was to investigate
DNA loop reorganization upon functional transitions in the glioblastoma T98G cells.
Methods. Single cell gel electrophoresis (a comet assay) was used to analyze the kinetics of
the DNA loop migration from the nucleoids obtained from the lysed cells. Results. The cells
arrested in the G1 phase of the cell cycle were characterized by a relatively low amount of
DNA in the comet tails due to a low content of DNA in the loops which may be resolved by
the comet assay (up to ~300 kb). After cell reactivation, the contour length of the loops es-
sentially increased in parallel with a decrease in the linear loop density along the genome.
Conclusions. An increase in the loop size and a respective decrease in the loop density may
be a general feature of activated cells as we earlier observed similar effects upon activation of
human lymphocytes.
K e y w o r d s: DNA loops, T98G cell line, comet assay, cell functional state.
Introduction
Loop domains are known to be the key ele-
ments of a higher order chromatin structure
[1–5]. The loops govern gene regulation and
other functional processes in chromatin, and
thus the loop organization is thought to vary
depending on cell functional states [6, 7].
Despite the comprehensive understanding of
general principles of the chromatin loop forma-
tion [5, 8–12] the reorganization of the loops
upon functional transitions remains far from
being studied in details.
Single-cell gel electrophoresis (the comet
assay) is a well-known technique which may
be applied to investigate the properties of nu-
cleoids obtained after a cell lysis. The assay
starts from the cells embedded in a thin layer
of agarose on a microscope slide and then
lysed. In the electric field the nucleoid DNA
Structure and Function
of Biopolymers
ISSN 1993-6842 (on-line); ISSN 0233-7657 (print)
Biopolymers and Cell. 2018. Vol. 34. N 6. P 426–434
doi: http://dx.doi.org/10.7124/bc.00098D
427
DNA loop organization in glioblastoma T98G cells at their different functional states
migrates towards the anode forming an elec-
trophoretic track, which resembles a comet tail
and can be visualized by fluorescent micros-
copy [13, 14].
In our previous works [15–21], measuring
the kinetics of DNA exit during electropho-
resis, we have shown that the nucleoid struc-
ture reflects some important features of the
loop organization in vivo. Moreover, we have
argued that some large-scale features of the
loop domain organization (and re-organiza-
tion) are preserved in nucleoids after cell
lysis and hence may be detected due to a
relatively simple technique, the comet assay
[20, 21].
Most of our previous experiments were
done using intact human lymphocytes.
However, in our recent work [20] we have
shown that the reorganization of DNA loops
occurs upon the lymphocyte activation by in-
terleukin 2, and that the loops in glioblastoma
T98G cells are organized differently with re-
spect to lymphocytes. In this article we studied
these cancer cells in more details.
The T98G cell line originates from the T98
line derived from a human glioblastoma mul-
tiform tumor [22, 23]. Like many other cancer
cells, the T98G cells may be cultivated in
suspension. But, in contrast to most of cancer
cell lines, the T98G cells can be arrested at G1
phase of the cell cycle when serum is absent
in the medium [22, 23]. At the same time,
after serum addition, the cells may be reacti-
vated to proliferation [22]. We have used these
features to investigate a possible reorganiza-
tion of DNA loops upon functional transitions
in the T98G cells. Our results show that, in-
deed, the loop organization varies in different
functional states of cells.
Materials and Methods
Sample preparation. The T98G cells were
cultivated at 37°C in Dulbecco’s medium,
which contained 10 % fetal bovine serum and
antibiotics. In order to synchronise the culture
and stimulate its arrest in G1 phase, the cells
were put in the same medium without serum
and incubated for 48 hours. For reactivation
to proliferation the cells were precipitated by
centrifugation, put in Dulbecco’s medium with
10 % fetal bovine serum and cultivated for
8 hours. The cells (either G1-arrested or reac-
tivated) were collected by centrifugation and
washed twice with PBS buffer (137 мМ NaCl,
2.7 мМ KСl, 10 мМ Na2HPO4, 2 мМ КH2PO4,
рН 7.4). In some cases nuclei were isolated
from the cells as described [19].
The comet assay was performed as de-
scribed earlier [15–20]. Briefly, the cells (or
nuclei) were embedded in the 0.67 % agarose
gel on the surface of a microscope slide. Slides
were treated with ice-cold lysis solution (2.5 M
NaCl, 100 mM EDTA, 10 mM Tris-HCl
(pH 8.0), 1 % Triton X-100 (Ferak, Germany))
for several hours. Then the slides were washed
twice by TBE buffer (89 мМ Tris-borat, 2 мМ
EDTA, рН 7.5) and electrophoresed in the
same buffer. In some cases chloroquine
(Sigma, USA) was added to the electrophore-
sis buffer. Several slides, simultaneously pre-
pared in the same way, were placed into the
electrophoresis tank, and then were taken out
every 10 minutes of electrophoresis. After
electrophoresis the slides were stained with
DAPI and immediately analyzed with a fluo-
rescent microscope. In total 100–150 random-
ly chosen nucleoids on each slide were exam-
ined using image analysis software CometScore
(TriTec, USA) to determine the relative amount
428
K. S. Afanasieva, A. Y. Semenova, L. L. Lukash et al.
of DNA in the tails and the tail length. The
relative amount of DNA in the tail was deter-
mined as the ratio of the tail fluorescence in-
tensity to the total intensity of the comet. The
tail length was defined as the distance from the
center of mass of the comet head to [the] dis-
tal end of the tail. Taking the contour length
of the loop to be roughly two times longer than
the extended loop, the tail length was multi-
plied by two and divided by 0.34 nm (the
distance between the adjacent base pairs) to
convert it in the contour length (in base pairs)
of the longest loops.
Data analysis. To compare experimental
kinetic plots the p-value was calculated using
a statistical permutation test [24, 25].
Kinetic plots (the relative amount of DNA
in the tail F versus electrophoresis time t) were
fitted according to two models. The first one,
which can be called “one-step”, corresponds
to the standard equation of monomolecular
kinetics:
F = Fm(1 – exp(–kt)), (1)
where Fm is the maximum relative amount of
DNA that can exit, k is the rate constant. The
second, “two-step” model takes into account
a two-step behavior of the kinetic plots [17].
According to this model, the plots were fitted
with the equation:
F = Fm [1– 1+ γLm( )exp(–γLm )]
F = A1 1– exp(– k1t)( ) + A2
1+ exp k2 t0 – t( )( )
F = Fm(1 – exp(–kt))
, (2)
where A1 and A2 are the maximum amplitudes
of the two components (A1 + A2 = Fm), k1 and
k2 are the rate constants, and t0 is the transition
half-time. The first term in Eq. 2 describes the
first rapid phase of the DNA exit; the second
term, which obeys the sigmoidal Boltzmann
equation, corresponds to the second retarded
phase.
The dependences of the relative amount of
DNA in the tail F on the contour length Lm of
the longest loops in the tail were fitted with
the equation derived in [20]:
F = Fm [1– 1+ γLm( )exp(–γLm )]
F = A1 1– exp(– k1t)( ) + A2
1+ exp k2 t0 – t( )( )
F = Fm(1 – exp(–kt))
, (3)
where γ is the linear loop density (the number
of loops per 1 kb).
Results and Discussion
As it was mentioned in the Introduction, the
T98G cells can be arrested at G1 phase of the
cell cycle. The first part of our experiments
was done with these G1-arrested cells.
Fig. 1A,B shows the kinetics of the comet tail
formation during the comet assay. In com-
parison with human lymphocytes, for which
the relative amount of DNA in the tails reached
21 % after a long-time electrophoresis [17–20],
for [the] T98G cells the saturation level of
DNA in the tails is much lower. It may be
suspected that a lower DNA exit might be
related to the less effective lysis of these cells.
However, the same maximum amount of DNA
exit into the tail was observed for the nuclei
isolated from the T98G cells (Fig. 1A, B).
Hence, the lysis procedure was quite effective.
As it was shown in our previous works for
human lymphocytes at different cell cycle
phases [17, 20], the kinetic plots of DNA exit
always have a two-step shape. The analysis of
our previous results allowed us to conclude
that the first step may be attributed to DNA on
the nucleoid surface whereas the second
429
DNA loop organization in glioblastoma T98G cells at their different functional states
step — to inner supercoiled loops, the exit of
which is retarded and cooperative [17]. In the
case of the T98G cells the two-step shape does
not seem so obvious. In principle, the plot can
be approximated by two models described in
the Materials and Methods: the “one-step”
model that obeys the standard equation (Eq. 1)
of monomolecular kinetics (Fig. 1A, C); and
the “two-step” model (Eq. 2) that includes the
second cooperative step described by the sig-
moidal Boltzmann equation (Fig. 1B, D).
However, the comparison of residuals and χ2
values (Fig. 1C, D) clearly shows that the two-
step model fits the experimental data much
better.
An additional evidence of the two-step be-
havior was obtained in the experiments with
chloroquine. As it was shown in our previous
works [15–17], the migration rate of the inner
loops is very sensitive to supercoiling: the
Fig. 1. The average relative amount (F) of DNA in the comet tails as a function of electrophoresis time (A, B) and
residuals (r) between experimental points and theoretical curves (C, D) for nucleoids obtained from T98G cells ar-
rested at G1 phase. Data for nucleoids from isolated cell nuclei (▲) are also presented in panels A and B. The theo-
retical curves are the results of fitting according to Eq. 1 (A, C) and Eq. 2 (B, D). The χ2 values for experimental points
relative to theoretical curves are 2·10–3 (A, C) and 8·10–5 (B, D).
A
C
B
D
430
K. S. Afanasieva, A. Y. Semenova, L. L. Lukash et al.
migration is accelerated when the loops are
relaxed due to partial DNA unwinding upon
intercalation of chloroquine at the concentra-
tion ~25 μg/ml. In total agreement with those
observations, the second step of the DNA exit
from T98G-derived nucleoids was also accel-
erated at the same chloroquine concentration
(Fig. 2).
Thus, similarly to the lymphocyte-derived
nucleoids, the comet tail of the T98G-derived
nucleoids is formed by two types of the loops:
surface loops that migrate rapidly and inner
supercoiled loops, the migration of which is
retarded. However, while the amplitude of the
first step of migration (A1 in Eq. 2) is the same
for both cell types (A1 = 0.07±0.01), the am-
plitude of the second step is very low for the
T98G cells (A2 = 0.05±0.01 against 0.14±0.01
for lymphocytes [17]). In other words, an es-
sential decrease in the relative DNA amount
in the tails in T98G-derived nucleoids is due
to the decreased number of the inner loops that
can be resolved by the comet assay.
As it was noted above, the T98G cells con-
sidered so far were arrested at G1 phase. These
cells can be reactivated: proliferation of these
cells is renewed after addition of serum in the
medium [22]. Such reactivation should obvi-
ously be accompanied by an increase in the
transcriptional activity. To examine possible
changes in the DNA loop organization upon
the reactivation we have investigated the kinet-
ics of DNA exit from nucleoids, derived from
the reactivated cells (Fig. 3). In comparison
with the G1-arrested cells, there were the fol-
lowing differences. First, an increase in the
DNA amount in the tails was observed at both
steps of the migration (A1 = 0.09 ± 0.01,
A2 = 0.08 ± 0.02). Second, an additional retar-
dation was observed for the second step: the
transition half-time (see Eq. 2) t0 = 41 ± 10
min against 27 ± 8 min for [the] nucleoids
Fig. 2. The average relative amount (F) of DNA in the
comet tails as a function of electrophoresis time for nucle-
oids obtained from T98G cells arrested at G1 phase in the
absence (○) and in the presence chloroquine at the con-
centration of 25 μg/ml (□). p = 0.019 (permutation test).
Fig. 3. The average relative amount (F) of DNA in the
comet tails as a function of electrophoresis time for nu-
cleoids obtained from arrested at G1 phase (○) and reac-
tivated (Δ) T98G cells. p = 0.003 (permutation test).
431
DNA loop organization in glioblastoma T98G cells at their different functional states
derived from the G1-arrested cells. Thus, the
reactivation of the T98G cells was accompa-
nied by an increase of DNA contained in both
the surface and inner loops that can be resolved
by the comet assay.
We have analyzed also the kinetics of the
tail length during the comet assay, the param-
eter that gives an estimation of the contour
length of the longest loops in the tail. Fig. 4
shows that the size of the longest loops was
essentially higher for the reactivated T98G
cells, especially for a long duration of electro-
phoresis. Such increase in the size may be a
reason for both the increase in the DNA amount
in the tails and the retardation of the DNA exit.
The correlations between the length of the
longest loops and the relative amount of DNA
in the tail for G1-arrested and reactivated cells
are presented in Fig. 5. These dependences,
which are proportional to the cumulative prob-
abilities of the loops below some size to be
present in the tail [20], allow one to estimate
the loop density (γ in Eq. 3), the main param-
eter of the exponential distribution of the loop
length [20]. Fitting Eq. 3 to dependences of
Fig. 5 gives γ = 0.059 ± 0.007 kb–1 for the
G1-arrested cells and γ = 0.025 ± 0.002 kb–1
for the reactivated T98G cells. Note that the
loop density estimated is related only to the
loops, the sizes of which are within the resolu-
tion of the comet assay (not larger than
~300 kb). A decrease in the density of these
loops upon reactivation clearly occurs in favor
of the larger loops that cannot be resolved by
the comet assay. The same effect (a decrease
of the loop density) was observed earlier for
Fig. 4. The contour length of the longest loops in the tails (Lm) as a function of electrophoresis time for nucleoids
obtained from arrested at G1 phase (A) and reactivated (B) T98G cells.
A B
432
K. S. Afanasieva, A. Y. Semenova, L. L. Lukash et al.
the lymphocytes activated by interleukin 2
[20]. In other words, an increase in the loop
size (which is in parallel with an increase of
the largest loops within the resolution of the
comet assay, Fig. 4) seems to be a general
feature of activated cells.
Conclusions
The results of our analysis of the kinetics of
electrophoretic track formation for the nucle-
oids derived from glioblastoma T98G cells can
be summarized as follows. (1) The cells ar-
rested at G1 phase of the cell cycle are char-
acterized by a relatively low amount of DNA
in the tails after a long-time comet assay. The
main reason of this is a low content of DNA
in the loops, the sizes of which are within the
resolution of the comet assay (up to ~300 kb).
(2) The reactivation of the T98G cells is ac-
companied by a redistribution of the loops: the
contour length of the loops resolved by the
comet assay essentially increases, in parallel
with a decrease in the linear density of these
loops along the genome. The results of this
work, together with our results obtained ear-
lier for activated lymphocytes, suggest that an
increase in the loop size may be a general
feature of the activated cells.
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434
K. S. Afanasieva, A. Y. Semenova, L. L. Lukash et al.
Організація петельних доменів ДНК
в гліобластомних клітинах T98G за їх різних
функціональних станів
К. С. Афанасьєва, A. Ю. Семенова, Л. Л. Лукаш,
A. В. Сиволоб
Резюме. Організація петельних доменів хроматину,
яка відіграє важливу роль у регуляції транскрипції,
напевно може залежати від функціонального стану
клітини. Мета роботи полягала у дослідженні мож-
ливої реорганізації петель ДНК при функціональних
переходах у гліобластомних клітинах T98G. Методи.
Ми застосовували метод електрофорезу ДНК ізольо-
ваних клітин (кометний електрофорез) для аналізу
кінетики міграції петель ДНК з нуклеоїдів, отриманих
з лізованих клітин. Результати. Клітини, заарешто-
вані на фазі G1 клітинного циклу, характеризуються
порівняно низьким вмістом ДНК у хвостах комет
внаслідок низького вмісту ДНК у складі петель, що
знаходяться у межах роздільної здатності кометного
електрофорезу (до ~300 кб). Після реактивації клітин
контурна довжина петель суттєво зростає, паралель-
но зі зниженням лінійної щільності петель уздовж
геному. Висновки. Оскільки подібні ефекти спосте-
рігались нами раніше для активованих лімфоцитів,
ми робимо висновок, що зростання розміру петель
та відповідне зниження їхньої лінійної щільності
може бути загальною характеристикою активованих
клітин.
К л юч ов і с л ов а: петлі ДНК, клітинна лінія
T98G, кометний електрофорез, функціональні стани
клітин.
Организация петельных доменов ДНК
в глиобластомных клетках T98G при их разных
функциональных состояниях
К. С. Афанасьева, A. Ю. Семенова, Л. Л. Лукаш,
A. В. Сиволоб
Резюме. Организация петельных доменов хроматина,
играющая важную роль в регуляции транскрипции,
предположительно может зависеть от функционально-
го состояния клетки. Цель работы заключалась в ис-
следовании возможной реорганизации петель ДНК при
функциональных переходах в глиобластомных клетках
T98G. Методы. Мы использовали электрофорез ДНК
изолированных клеток (кометный электрофорез) для
анализа кинетики миграции петель ДНК из нуклеоидов,
полученных из лизированных клеток. Результаты.
Клетки, арестованные на фазе G1 клеточного цикла,
характеризуются сравнительно низким содержанием
ДНК в хвостах комет из-за низкого содержания ДНК в
составе петель, которые находятся в пределах разре-
шающей способности кометного электрофореза (до
~300 кб). После реактивации клеток контурная длина
петель существенно возрастает, параллельно со сниже-
нием линейной плотности петель вдоль генома.
Выводы. Поскольку аналогичные эффекты наблюда-
лись нами ранее для активированных лимфоцитов, мы
заключили, что возрастание размера петель и соответ-
ствующее снижение их линейной плотности может
быть общей характеристикой активированных клеток.
К л юч е в ы е с л ов а: петли ДНК, клеточная линия
T98G, кометный электрофорез, функциональные со-
стояния клеток.
Received 10.10.2018
|
| id | nasplib_isofts_kiev_ua-123456789-154372 |
| institution | Digital Library of Periodicals of National Academy of Sciences of Ukraine |
| issn | 0233-7657 |
| language | English |
| last_indexed | 2025-11-28T03:50:04Z |
| publishDate | 2018 |
| publisher | Інститут молекулярної біології і генетики НАН України |
| record_format | dspace |
| spelling | Afanasieva, K.S. Semenova, A.Y. Lukash, L.L. Sivolob, A.V. 2019-06-15T14:39:20Z 2019-06-15T14:39:20Z 2018 DNA loop organization in glioblastoma T98G cells at their different functional states / K.S. Afanasieva, A.Y. Semenova, L.L. Lukash, A.V. Sivolob // Вiopolymers and Cell. — 2018. — Т. 34, № 6. — С. 426-434. — Бібліогр.: 22 назв. — англ. 0233-7657 DOI: http://dx.doi.org/10.7124/bc.00098D https://nasplib.isofts.kiev.ua/handle/123456789/154372 577.323:576.08 The loop domain organization of chromatin, which plays an important role in transcription regulation, may depend on the cell functional state. Aim. To investigate DNA loop reorganization upon functional transitions in the glioblastoma T98G cells. Methods. Single cell gel electrophoresis (a comet assay) was used to analyze the kinetics of the DNA loop migration from the nucleoids obtained from the lysed cells. Results. The cells arrested in the G1 phase of the cell cycle were characterized by a relatively low amount of DNA in the comet tails due to a low content of DNA in the loops which may be resolved by the comet assay (up to ~300 kb). After cell reactivation, the contour length of the loops essentially increased in parallel with a decrease in the linear loop density along the genome. Conclusions. An increase in the loop size and a respective decrease in the loop density may be a general feature of activated cells as we earlier observed similar effects upon activation of human lymphocytes. Організація петельних доменів хроматину, яка відіграє важливу роль у регуляції транскрипції, напевно може залежати від функціонального стану клітини. Мета. Дослідити можливу реорганізацію петель ДНК при функціональних переходах у гліобластомних клітинах T98G. Методи. Ми застосовували метод електрофорезу ДНК ізольованих клітин (кометний електрофорез) для аналізу кінетики міграції петель ДНК з нуклеоїдів, отриманих з лізованих клітин. Результати. Клітини, заарештовані на фазі G1 клітинного циклу, характеризуються порівняно низьким вмістом ДНК у хвостах комет внаслідок низького вмісту ДНК у складі петель, що знаходяться у межах роздільної здатності кометного електрофорезу (до ~300 кб). Після реактивації клітин контурна довжина петель суттєво зростає, паралельно зі зниженням лінійної щільності петель уздовж геному. Висновки. Оскільки подібні ефекти спостерігались нами раніше для активованих лімфоцитів, ми робимо висновок, що зростання розміру петель та відповідне зниження їхньої лінійної щільності може бути загальною характеристикою активованих клітин. Организация петельных доменов хроматина, играющая важную роль в регуляции транскрипции, предположительно может зависеть от функционального состояния клетки. Цель. Исследовать возможной реорганизации петель ДНК при функциональных переходах в глиобластомных клетках T98G. Методы. Мы использовали электрофорез ДНК изолированных клеток (кометный электрофорез) для анализа кинетики миграции петель ДНК из нуклеоидов, полученных из лизированных клеток. Результаты. Клетки, арестованные на фазе G1 клеточного цикла, характеризуются сравнительно низким содержанием ДНК в хвостах комет из-за низкого содержания ДНК в составе петель, которые находятся в пределах разрешающей способности кометного электрофореза (до ~300 кб). После реактивации клеток контурная длина петель существенно возрастает, параллельно со снижением линейной плотности петель вдоль генома. Выводы. Поскольку аналогичные эффекты наблюдались нами ранее для активированных лимфоцитов, мы заключили, что возрастание размера петель и соответствующее снижение их линейной плотности может быть общей характеристикой активированных клеток. en Інститут молекулярної біології і генетики НАН України Вiopolymers and Cell Structure and Function of Biopolymers DNA loop organization in glioblastoma T98G cells at their different functional states Організація петельних доменів ДНК в гліобластомних клітинах T98G за їх різних функціональних станів Организация петельных доменов ДНК в глиобластомных клетках T98G при их разных функциональных состояниях Article published earlier |
| spellingShingle | DNA loop organization in glioblastoma T98G cells at their different functional states Afanasieva, K.S. Semenova, A.Y. Lukash, L.L. Sivolob, A.V. Structure and Function of Biopolymers |
| title | DNA loop organization in glioblastoma T98G cells at their different functional states |
| title_alt | Організація петельних доменів ДНК в гліобластомних клітинах T98G за їх різних функціональних станів Организация петельных доменов ДНК в глиобластомных клетках T98G при их разных функциональных состояниях |
| title_full | DNA loop organization in glioblastoma T98G cells at their different functional states |
| title_fullStr | DNA loop organization in glioblastoma T98G cells at their different functional states |
| title_full_unstemmed | DNA loop organization in glioblastoma T98G cells at their different functional states |
| title_short | DNA loop organization in glioblastoma T98G cells at their different functional states |
| title_sort | dna loop organization in glioblastoma t98g cells at their different functional states |
| topic | Structure and Function of Biopolymers |
| topic_facet | Structure and Function of Biopolymers |
| url | https://nasplib.isofts.kiev.ua/handle/123456789/154372 |
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