Biology Reference
In-Depth Information
interacts with Centrocortin, a protein that is required for cleavage furrow forma-
tion and is localized both to centrosomes and to cleavage furrows (Kao and
Megraw
2009
). It is therefore possible that the centrosome functions as a signaling
hub within the cell. At this signaling hub, proteins can interact in order to integrate
information and later move to other domains in the cell where they execute their
function (Alieva and Uzbekov
2008
; Wang et al.
2009
).
Another way by which to study centrosomes in early embryogenesis is to study
hypomorphic mutations of centrosomal proteins that are essential for centrosome
formation and produce a female that can mate. In this case, the studied protein is
partially functional and missing an activity that is essential specifically for cen-
trosome function during embryogenesis. An example of one such mutation is asl
1
,
in which the C-terminus of the essential centrosomal protein Asterless (Asl) is
truncated (Blachon et al.
2008
). While flies homozygous for the severe loss-
of-function allele asl
mecD
are unable to walk and mate, asl
1
generates viable
females that can lay eggs. Analysis of embryonic centrosomes generated from an
asl
1
female finds that they initially form asters, but these asters are not stable and
later fall apart ((Varmark et al.
2007
) and S. B. unpublished data). In addition,
pronuclei fusion does not take place and embryo development is arrested at the
zygote stage. Similar results were obtained when a hypomorphic mutation of the
Drosophila spd-2 gene was studied, while severe loss-of-function spd-2 mutants
are unable to walk and mate (Dix and Raff
2007
; Giansanti et al.
2008
).
One can also study centrosomal proteins that are essential for centrosome
formation and produce a female that can mate using genetic tricks. One way to do
so is to make germline clones that lack any particular essential centrosomal protein
from maternal contribution (Stevens et al.
2007
). This is done using the dominant
female sterile (DFS) technique (Chou and Perrimon
1996
), a variant of FLP-FRT
recombination. In this case, recombination is induced in larvae via a heat shock-
inducible flippase (FLP); as adults, the fly produces homozygous mutant oocytes
that lack the essential centrosomal protein. Study of oocytes that are mutant for
sas-4 or sas-6 after they are fertilized with a wild-type male finds that they possess
two centriolar structures (presumably the giant centriole and PCL) (Stevens et al.
2007
). These centriolar structures can nucleate centrosomes and can undergo few
nuclear cycles before embryogenesis is arrested presumably because the centro-
somes cannot duplicate. This suggests that the PCL can form an independent
centrosome and demonstrates that centrosomes are essential for syncytial blasto-
derm development (after their role in zygote pronuclei fusion).
An alternative method to study centrosomes in the syncytial blastoderm is to
inject it with an antibody for a particular centrosomal protein and observe the
consequence (Conduit et al.
2010
). A potential problem with this approach is that
when a centrosomal protein forms a complex, binding of an antibody may not only
inactivate its intended protein target, but may also inactivate other proteins found
in the complex.
