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Spectraplakins are difficult to address functionally, since they are large proteins
encoded by long transcripts with a variety of isoforms that challenge traditional
cloning and transgenic techniques. Moreover, their function relies on the presence
of specific functional domains that are cell type and context dependent. Fortunately,
genome-editing approaches offer new possibilities to study them, including in
zebrafish, where macf1 loss of function does not cause embryonic lethality like in
mammals (Kodama et al. 2003 ). Thus, zebrafish oocytes offer a unique possibility
to interrogate the requirement of Macf1 single modules in cell polarity. Such studies
will help to understand the relationship between the cytoskeleton, the Bb and the
RNA localization machinery that together drive AV polarity establishment during
early oogenesis.
An interesting question is how the translocation of the Bb to the future vegetal
pole is coordinated with the simultaneous growth of the oocyte. Between the point
when the mature Bb is first detected and the time the Bb reaches the future vegetal
cortex, docks, and disassembles, the oocyte is at least twofold larger (Fig. 5.2). This
growth involves increasing the cytoplasmic volume and expanding the cytoplasmic
membrane. How these changes affect the organization of cytoskeletal or other cel-
lular features that lead to Bb vegetal localization and disassembly is unknown.
Understanding the developmental context of oocyte differentiation will be impor-
tant to dissecting its polarization mechanisms, including tracing early stages of Bb
formation as well as analyzing its translocation, docking, and disassembly at the
future vegetal pole.
5.6 The Animal Pole Localization of RNAs
At the animal pole, the egg is fertilized in frogs and fish and the blastodisc forms in
zebrafish. Like at the vegetal pole, RNAs localize to the animal pole during oogen-
esis, though they localize after the Bb disassembles, suggesting that vegetal pole
identity is acquired first. In zebrafish, RNAs like cyclin B1 (cycB) and pou5f3 (pre-
viously called pou2) localize to the animal pole in stage II and early stage III,
respectively, remaining there in a cortical and tight distribution throughout oocyte
maturation (Fig. 5.2c) (Howley and Ho 2000 ). Interestingly, Vg1 RNA in zebrafish
localizes to the animal pole right below the micropyle (Marlow and Mullins 2008 ),
unlike in frogs where Vg1 localizes to the vegetal pole (Mowry and Melton 1992 ).
As stated earlier, the lack of vegetal pole identity in zebrafish buc mutants causes
the formation of multiple micropyles and a radial animal identity with no recogniz-
able blastodisc in the egg (Dosch et al. 2004 ; Marlow and Mullins 2008 ). Similarly,
pou5f3 and cycB RNAs show a radial localization in buc mutant stage III oocytes,
indicating that Buc defines the egg AV axis by also restricting the localization of
animal RNAs in the oocyte.
In stage IV, zebrafish oocytes undergo maturation. Through this process, oocytes
progress from prophase of meiosis I (MI) to metaphase of meiosis II (MII), extrud-
ing the first polar body at the animal pole. Several changes occur during this pro-
M. Escobar-Aguirre et al.