tive in inducing synchronization in plant cells; either the majority of the methods were found to be only
partially effective or the agents that cause synchrony were found to have toxic effects on cell metabolism.
Among the DNA synthesis inhibitors, aphidicolin is found to be the most effective in inducing syn-
chronous growth in suspension cultures as well as in differentiated tissues. However, because of endoge-
nous aphidicolin-inactivating activity in plant cells, which varies between cell types and plants, the con-
centration of aphidicolin and length of the incubation should be determined empirically in each case.
Treatment of cells with aphidicolin, a mycotoxin that specifically blocks nuclear DNA replication by in-
hibiting DNA polymerase , causes accumulation of cells at the G 1 /S boundary of the cell cycle [323].
The effect of this inhibitor is reversible, hence removal of aphidicolin from the medium results in syn-
chronous resumption of DNA synthesis. In several plant cells aphidicolin was shown to arrest about
80–95% of cells in G 1 , which were found to move synchronously through the first round of mitosis after
G 1 /S arrest [148,322]. The tobacco Bright Yellow (BY-2) cell line is one of the most well-characterized
cell culture systems and can be synchronized efficiently using these inhibitors [324].
In suspension cultures of Catharanthus roseus, double phosphate starvation effectively induces syn-
chrony [321]. This system is already helping to identify some of the phase-specific changes in mRNA and
proteins [234,325]. In suspension cultures of Datura, hydroxyurea, another inhibitor of DNA synthesis,
reversibly arrested the cells at the G 1 /S boundary [321].
Other cell cycle inhibitors have been found that block specific stages of the cell cycle by inhibiting
cell cycle proteins. Olomoucine, a purine analogue that inhibits Cdks at micomolar concentrations while
having little effect on other protein kinases, inhibits both the G 1 /S and G 2 /M transitions in plants [116].
Two structurally modified olomoucine-like molecules, bohemine and roscovitine, inhibit Cdks 10 to 100-
fold better than olomoucine [117]. Roscovitine was found to block the cell cycle prior to entry into S and
M phases [310,326].
Synchronization of plant cells with the preceding methods coupled with flow cytometry should
greatly expedite the progress in cell cycle research in plants [327]. During the last 10 years flow cytom-
etry has been increasingly used in analyzing plant cells. Protoplasts and isolated nuclei are amenable to
flow cytometry. However, when protoplasts are used some modifications in methods and instrumentation
are necessary because of their large size (20–75 m) [327,328]. Developments in the use of flow cytom-
etry for plant protoplasts have opened new avenues to analyze cell cycle regulatory proteins. Using mul-
tiparameter analysis, one could monitor the levels of two or more desired proteins during different phases
of the cell cycle [329].
VIII. CELL CYCLE IN PLANT DEVELOPMENT
Cell division is one of the primary determinants of various aspects of development in multicellular eu-
karyotic organisms. The regulatory mechanisms that determine various aspects of the cell cycle (e.g.,
which of the cells in an organism should undergo cell division, the timing and the plane of cell division
in these cells, and which cells should remain quiescent and reenter the cell cycle) play a critical role in
plant developmental processes such as embryogenesis, seed germination, and flowering. Hence, investi-
gating these regulatory mechanisms will not only help us understand cell cycle regulation but also enable
us to elucidate developmental programming in plants. Various developmental processes that involve the
cell cycle are unique to plants. Unlike that in animals, cell division in higher plants is restricted to meris-
tematic regions (shoot apical meristem, root apical meristem, and lateral meristem). The primary meris-
tems such as shoot and root apical meristems continuously divide and contribute to the production of new
organs and growth of the plants. Furthermore, shoot apical meristem can lose its indeterminate vegetative
growth to become determinate floral meristem. The transition from vegetative meristem to floral meris-
tem involves shortening of the cell cycle time as well as synchronization of the cell cycle [5]. In plants,
during the course of normal development, quiescent cells become proliferative. For instance, lateral
meristems (pericycle and cambium), auxillary buds, and cambium retain their ability to undergo cell di-
vision and enter into the cell cycle in response to some developmental cues. The root apex in plants con-
tains, in addition to dividing cells, a group of cells called the quiescent center, which do not normally un-
dergo cell division. However, if the root meristem is damaged, cells in the quiescent center reenter the cell
cycle and form new meristem. In addition, if the cells from the quiescent center are cultured in vitro in
250 REDDY AND DAY