Figure 5.

The ROD-1 β-Propeller Suppresses Ubiquitous and Complete Oligomerization of ROD-1 into Filaments

(A) Schematic of the protein complexes generated by the expression of RNAi-resistant mCherry::rod-1 transgenes integrated in single copy on chromosome II. Note that the actual RZZ complex is a dimer of the three-subunit assembly depicted here, which means mCherry::ROD-1(Δ1-372) is able to dimerize with endogenous ROD-1.

(B) Selected images from time-lapse sequences in early embryos co-expressing mCherry::ROD-1(Δ1-372), GFP::histone H2B, and GFP::γ-tubulin. mCherry::ROD-1(Δ1-372) oligomerizes into filaments when endogenous ROD-1 is depleted (see also Video S3). This causes defects in chromosome segregation (arrow), because RZZ is titrated away from kinetochores. Dashed lines outline the embryos. Scale bar, 10 μm.

(C) Embryonic viability assay showing that embryos expressing mCherry::ROD-1(Δ1-372) are only viable when endogenous ROD-1 is present. Values are plotted as mean ± 95% confidence interval, and n indicates the number of mothers whose progeny was counted.

(D) Immunofluorescence image of an isolated oocyte-producing gonad, showing that mCherry::ROD-1(Δ1-372) forms filaments ubiquitously in oocytes and embryos. Scale bar, 25 μm.

(E) Immunofluorescence image of maturing oocytes showing co-localization of mCherry::ROD-1(Δ1-372) with 3×FLAG-tagged CZW-1^Zw10 on filaments. Scale bar, 10 μm.

(F) Immunofluorescence image showing that mCherry::ROD-1(Δ1-372) filaments do not cluster at spindle poles during mitosis. Dashed lines outline the embryos. Scale bar, 5 μm.

(G) Immunofluorescence images showing co-localization of KNL-1 and BUB-1 with mCherry::ROD-1(Δ1-372) filaments in meiosis I embryos. Dashed lines outline the embryos. Scale bars, 5 μm.

(H) Fluorescence images of live oocytes showing that the formation of mCherry::ROD-1(Δ1-372) filaments depends on CZW-1^Zw10, but not ZWL-1^Zwilch. Scale bar, 10 μm.

See also Figure S5.