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Fig. 3. Localization of non-actin binding YFP-Myosin IIDN. The schematic
of myosin II structure shows the hexamer of two heavy chains (blue), two
essential light chains (red) and two regulatory light chains (yellow). The
schematic of mYFP-myosin IIDN shows the actin-binding head domain
of the heavy chain replaced with the YFP moiety (green), resulting in two
forms of YFP-containing myosin (heterodimer and homodimer) when expressed
alongside endogenous myosin II, with both forms compromised in their ability
to bind actin. (A,G) Cells undergoing cytokinesis are identified by antibody
staining for phosphohistone H3 (red), nuclei are stained with Hoechst (blue)
and myosin II (A) is stained with anti-myosin II antibody (green), whereas
mYFP-myosin IIDN (G) is stained with anti-GFP antibody (green).
Both myosin II and mYFP-myosinIIDN are localized to the cytokinetic
furrow of dividing cells (arrow). (B-F,H-L) Anti-NRT staining (red),
anti-myosin II staining (green in B-F) and anti-GFP staining (green in H-L).
Despite the tendency of mYFP-myosin IIDN to form aggregates
(asterisk in H), both the endogenous myosin II and the
mYFP-myosinIIDN localize to the basal cellularization front (arrow
in B,C,H,I), where levels later become greatly reduced in the ventral cells
(arrow in F,L). This reduction is, however, delayed (E,K) and patchy (F,L) for
mYFP-myosin IIDN. mYFP-myosin IIDN fails to localize to
the apical side of ventral cells during gastrulation (compare asterisks in D,E
with those in J,K).
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