Mitotic Spindle Assembly and Function 145
act with MTs (Burgess and Northcote 1967; Bajer and Molè-Bajer 1969). Since
plants lack centrosomes or spindle pole bodies, it has been suggested that the
nuclear surface acts as an MTOC (Schmit 2002). Abundant evidence supports
the role of the nucleus as an MTOC: (1) isolated nuclei are able to nucleate MT
growth from their surfaces in vitro (Stoppin et al. 1994); (2) gamma tubulin,
the universal MT nucleator localizes to the NE (Liu et al. 1993; Erhardt et al.
2002); (3) spc98, a key centrosomal component is found in plants and local-
izes to the nuclear surface (Erhardt et al. 2002); (4) adding antibodies to spc98
inhibits the ability of nuclei to nucleate MTs (Erhardt et al. 2002); (5) anti-
bodies to centrosomal components frequently localize to the NE, and; (6) MTs
reform on the NE after recovery from depolymerization with drugs (Falconer
et al. 1988; Galatis and Apostolakos 1991).
2.4
Formation of the Prometaphase and Metaphase Spindle
Prometaphase begins at nuclear envelope breakdown (NEB) and lasts until
a clear metaphase plate is formed. During prometaphase, MTs congress in-
ward from the poles to the equator and immediately begin forming connec-
tions with kinetochores (forming k-fibers) or with other MTs from the op-
posite pole (forming interpolar bundles). K-fiber formation is asynchronous
between sister kinetochores (Bajer 1987), and corresponds with the initiation
of chromosome movements. K-fibers mediate the congression of chromo-
somes toward the metaphase plate and the separation of sister chromatids
during anaphase, while interpolar MTs serve to stabilize the spindle along
its length and width, and participate in anaphase spindle elongation. In
most cases, the metaphase spindle lacks well-defined poles (as seen in ani-
mal cells), even if the preceding prophase spindle did contain focused poles
(Fowke 1993). Metaphase spindles instead exhibit a barrel-shaped morph-
ology, in contrast to animal spindles, which maintain a fusiform morph-
ology throughout mitosis, even in the absence of centrosomes (Khodjakov
et al. 2000).
Early EM studies showed that prior to NEB, MTs may penetrate the NE in
the polar regions (Pickett-Heaps and Northcote 1966) and also associate with
deformations of the prophase nucleus (Bajer and Molè-Bajer 1969). During
NEB inHaemanthusendosperm, the NE appears wavy, giving the impres-
sion of “boiling”, and subsequently begins fragmentation at the polar regions,
with large invaginations of membrane apparently being pushed in by bundles
or sheets of 50–100 MTs (Bajer and Molè-Bajer 1969). The NE fragments are
later degraded, or they may persist within the spindle throughout mitosis, be-
coming associated with chromatin, and in some cases, becoming reincorpo-
rated into the NE of the daughter nuclei (Bajer and Molè-Bajer 1969). Electron
micrographs show that most MTs invade the nucleus in the above-mentioned
bundles before becoming attached to kinetochores or opposing MTs. These