Plant Science 160 2000 57 – 66
Inhibition of cytoplasmic DNA synthesis may be an initial signal for the induction of sexual reproduction in Closterium ehrenbergii
Machiko Imaizumi
a,
, Jin Hamada
b
a
Department of Biology, Shiga Uni6ersity of Medical Science, Ohtsu
520
-
2192
, Japan
b
Department of Community Medicine, Faculty of Medicine, Toyama Medical and Pharmaceutical Uni6ersity, Toyama
930
-
0152
, Japan Received 9 November 1999; received in revised form 8 August 2000; accepted 8 August 2000
Abstract
We investigated the effect that inhibitors of cytoplasmic DNA cytDNA andor nuclear DNA nDNA synthesis have on the frequency of zygote formation in Closterium ehrenbergii and whether these inhibitors correlate with arrest of pyrenoid production
and enlargement of pyrenoidal size accompanying pregametogenesis in the alga. CytDNA inhibitors, ethidium bromide EB, acridine orange and novobiocin NB, enhanced zygote formation in the presence of nitrate, and these chemicals correlated with
both a decrease in pyrenoid production and an increase in pyrenoidal size. By contrast, a nDNA inhibitor, aphidicolin APC, enhanced zygote formation in the presence and absence of nitrate, and correlated with only an increase in pyrenoidal size.
Meanwhile, inhibitors of both nDNA and cytDNA synthesis, mitomycin C MC, hydroxyurea HU and arabinosyl cytosine Ara-C, enhanced zygote formation in the presence andor absence of nitrate. In addition, MC and HU correlated with only an
increase in pyrenoidal size and Ara-C only a decrease in pyrenoid production. This result seemed to indicate that MC and HU functioned as selective inhibitors of nDNA synthesis, and Ara-C as a selective inhibitor of cytDNA synthesis. It is suggested that
differentiation into pregametes correlates with the inhibition of both nDNA and cytDNA synthesis, levels of which are probably low, the inhibition of cytDNA synthesis being the initial signal for pregametogenesis. © 2000 Elsevier Science Ireland Ltd. All
rights reserved.
Keywords
:
Closterium ehrenbergii; DNA synthesis inhibitor; Pyrenoid; Sexual reproduction; Zygote formation www.elsevier.comlocateplantsci
1. Introduction
Cells of Closterium ehrenbergii form zygotes ef- fectively when two heterothallic strains are mixed
under nitrogen source KNO
3
, CaNO
3 2
· 4H
2
O- depleted conditions [1,2]. The presence of a nitro-
gen source
lowers the
frequency of
zygote formation. This suggests that some pathway in
nitrogen metabolism regulates the induction of zygote formation sexual reproduction in this
alga. However, the regulatory mechanism remains largely unknown. In a preliminary study, we found
that the frequency of zygote formation in this alga is greatly increased when ethidium bromide EB,
which is an inhibitor of cytoplasmic DNA cytDNA synthesis, is added to the alga under
conditions in which nitrate is present. This result indicates that the inhibition of cytDNA synthesis
is a signal for the induction of zygote formation in the alga. Hamada [3] suggested, on the other
hand, that nuclear DNA nDNA duplication did not occur during gametogenesis in this alga and
that inhibition of nDNA synthesis was involved in the induction of zygote formation. It was reported
that nuclear and organellar DNAs replicated at different times in the cell cycles [4 – 7] and their
replication cycles were tightly coupled with each other [8]. All this evidence suggests that the inhibi-
Corresponding author. Tel.: + 81-77-5482123; fax: + 81-77- 5482048.
E-mail address
:
imaizumibelle.shiga-med.ac.jp M. Imaizumi. 0168-945200 - see front matter © 2000 Elsevier Science Ireland Ltd. All rights reserved.
PII: S 0 1 6 8 - 9 4 5 2 0 0 0 0 3 6 1 - 7
tion of nDNA andor cytDNA synthesis is closely involved in the induction of zygote formation.
We noticed that the pyrenoids in C. ehrenbergii were greatly enlarged during sexual reproduction
compared with those during asexual reproduction. This would suggest that regulation of pyrenoid
function occurs during the conversion from asex- ual to sexual reproduction in this alga. In the
present
study, we
investigated the
ratio of
pyrenoidcell number, and the pyrenoidal sizes of the alga throughout the asexual and sexual stages
of reproduction, and found that the modulation of pyrenoid production and function accompanied
the formation of pregametes pregametogenesis.
The present study investigated whether the inhi- bition of nDNA andor cytDNA synthesis is in-
volved in the induction of zygote formation in C. ehrenbergii and if so, which form of inhibition is an
initial signal for the induction. To this end, the effect of selective inhibitors of cytDNA or nDNA
synthesis on the frequency of zygote formation in the alga, as well as on the modulation of pyrenoid
production and pyrenoidal size during pregameto- genesis, was examined.
2. Materials and methods
