84 P.A. Sabelli · B.A. Larkins
Recent work and previous findings indicate that endoreduplicated chromo-
somes in maize endosperm are completely replicated with a loose polytene
structure (Kowles and Phillips 1985; Kowles and Phillips 1988; Kowles et al.
1990) in which they appear to be tightly associated at the centromere and,
unexpectedly, at the highly heterochromatic knob regions (Bauer and Birch-
ler 2006). This observation suggested that during endoreduplication specific
chromatin binding proteins, perhaps of the cohesion (cohesin) class, may
be in place to ensure chromosome association with a higher concentration
at centromeres and knobs. A polytene structure of chromosomes is likely
to involve extensive chromatin reorganization (Zhao and Grafi 2000), al-
though this aspect of endosperm development has been poorly investigated.
However, this view is supported by the observation that interploidy crosses,
which are known to perturb caryopsis development and the endoreduplica-
tion pattern, result in dramatic alteration of chromatin organization (Bauer
and Birchler 2006). Chromatin reorganization has also been observed during
the endoreduplication phase in the endosperm of durum wheat, but per-
haps surprisingly, in this system endoreduplication seems to entail increased
chromatin condensation, potentially resulting in methylation-induced gene
silencing (Polizzi et al. 1998).
During the early endoreduplication period of maize endosperm develop-
ment, DNA replication appears to proceed extremely rapidly. Each round of
DNA synthesis is estimated to take 22 – 24 h, with the last round requiring an
average DNA synthesis of 916 μm/second/nucleus, which is equivalent to the
incorporation of 2.75× 106 bases pairs/second/nucleus (Kowles and Phillips
1988).
Early experiments on the analysis of CDK activity typically associated with
S-phase or M-phase revealed a peak in M-phase-associated CDK activity at
10 DAP, coincident with a peak in the frequency of mitotic figures (Kowles
and Phillips 1988), and a peak in S-phase-associated CDK activity at 16 DAP,
which is when most endosperm cells are engaged in endoreduplication (Grafi
and Larkins 1995). Thus, similarly to what was observed in animal systems,
a switch from a mitotic to an endoreduplication cell cycle in developing endo-
sperm appears to involve a downregulation of M-phase-associated CDK and
a sustained or upregulated S-phase CDK activity. In agreement with this view,
the characterization of A- and B-type CDKs has shown that whereas CDKA
protein expression is relatively constant in developing endosperm, expression
of the M-phase-associated CDKB decreases in endoreduplicating endosperm
(Dante 2005).
CDKA, on the other hand, plays a key role in endoreduplication. CDKA
activity was specifically downregulated in transgenic endosperms by overex-
pressing a dominant-negative mutant form of CDKA (CDKA-DN), in which
an Asp146Asn mutation abolished kinase activity, but did not interfere with
cyclin binding (Leiva-Neto et al. 2004). Because CDKA-DN expression was
driven by the strong 27 -kDγ-zein promoter throughout the starchy endo-