Picture from Phillips CM et al. (2006) Dev Cell "A family of zinc-finger proteins is required for chromosome-specific pairing ...."

Figure 5. ZIM Proteins Localize to Autosomal Pairing Centers(A-D) ZIM-1 colocalizes the left ends of chromosomes II and III. Two arrows point to the two ZIM-1 foci in a single nucleus and to the chromosomes II and III FISH probes, which colocalize with them.(E-H) ZIM-2 colocalizes with a FISH probe to the right end of chromosome V. A particularly clear example of this colocalization is indicated bythe arrow.(I-P) ZIM-3 colocalizes with FISH probes to the right end of chromosome I (I-L) and the left end of chromosome IV (M-P). Arrows point to clear examples. (A, E, I, and M) Late transition zone or early pachytene nuclei stained with DAPI. (B, F, J, and N) Immunolocalization of ZIM-1, ZIM-2,ZIM-3, and ZIM-3, respectively. (C, G, K, and O) FISH signals corresponding to IIL, IIIL, VR, IR, and IVL. (D, H, L, and P) Merged images.(Q) Correspondence of ZIM-1, -2, -3, and HIM-8 to the PC regions of the six C. elegans chromosomes. The region to which the PCs have been mapped genetically are demarcated in blue. Scale bars represent 5 um.

Picture from Phillips CM et al. (2006) Dev Cell "A family of zinc-finger proteins is required for chromosome-specific pairing ...."

Figure 4. ZIM Proteins Localize to Discrete Foci at the Nuclear Envelope during Early Meiotic Prophase(A) Wild-type gonad stained with DAPI and antibodies to ZIM-1 and ZIM-2. Foci are most apparent in the transition zone of the gonad (indicated by white bar). Some signals can be seen outside the transition zone/early pachytene region, but most of these do not correspond to DAPI-stained regions and thus are likely nonspecific background.(B and C) In the latter portion of the transition zone, most nuclei have two foci of ZIM-1 (B, green) and a single focus of ZIM-2 (C, red).(D) In early pachytene, two foci of ZIM-3 (orange) can been visualized in each nucleus.(E-G) Optical sections showing wild-type pachytene nuclei stained with antibodies against nuclear lamin/LMN-1 (white), ZIM-1 (E, green), ZIM-2(F, red), and ZIM-3 (G, orange). These cross-sections reveal the close juxtaposition of the ZIM proteins with the lamina more clearly than projections through the entire nuclear volume, but not all ZIM-1 and ZIM-3 foci are visible within the shallow focal plane. Scale bars represent 5 um.

Picture from Phillips CM et al. (2005) Cell "HIM-8 binds to the X chromosome pairing center and mediates chromosome-specific ...."

Figure S1. HIM-8 Localizes near the Nuclear Envelope. The larger image displays a cross-section through a field from the pachytene region of a him-8(me4) hermaphrodite stained with anti-LMN-1 (lamin) and anti-HIM-8 antibodies. DNA is stained with DAPI. Because the close association of HIM-8 foci with the nuclear lamina is not always apparent in 3D projections such as those shown in Figure 6, this image shows a single optical section from a deconvolved 3D data stack. Consequently, not all HIM-8 foci are visible in this plane. The DAPI (blue), anti-HIM-8 (green), and anti-LMN-1(red) images are displayed in separate RGB channels rather than the mixed pseudocolors used in Figure 6 so that intensity values in each channel could be analyzed separately. The inset shows a line-scan of intensities through one nucleus (outlined in the larger image), which was selected for display here because both of the unpaired HIM-8 foci could be detected in the same optical section. A horizontal line that traverses the peaks of both HIM-8 foci (shown in white) was scanned in each wavelength. Intensity traces are shown in white (DAPI), red (lamin), and green (HIM-8). Note that the DAPI intensity is bounded by and internal to the major peaks of lamin intensity, but the HIM-8 foci correspond closely in their peak positions to the outer lamin peaks.

Picture from Phillips CM et al. (2005) Cell "HIM-8 binds to the X chromosome pairing center and mediates chromosome-specific ...."

Pachytene nuclei were stained with antibodies against HIM-8 (yellow) and anti-LMN-1 (red), which marks the nuclear lamina, or nuclear envelope. Scale bar represent 5 μm.

Picture from Phillips CM et al. (2005) Cell "HIM-8 binds to the X chromosome pairing center and mediates chromosome-specific ...."

Figure 4. The HIM-8 Protein Localizes to the PC Region of the X Chromosome. All images show projections through fields of pachytene-region nuclei from animals of the indicated genotypes. The entire nuclear volume is shown in each projection. All scale bars represent 5 μm.(A) Wild-type hermaphrodite, revealing a single HIM-8 focus in each nucleus due to close association between the two binding sites detected in premeiotic and very early meiotic nuclei (Figure 3).(B) Wild-type male, which displays a single (unpaired) HIM-8 focus in each nucleus. In contrast to hermaphrodites, males also have one HIM-8 focus per nucleus in the premeiotic region of the gonad (data not shown).(C) Hermaphrodite carrying two copies of the mnDp66 duplication of the X chromosome PC region. An additional HIM-8 focus is present in each nucleus, corresponding to the paired and synapsed duplication.(D) Hermaphrodite carrying both mnDp66 and the PC deficiency meDf2, which eliminates one of the two foci observed in (C).(E) HIM-8 immunostaining was performed in conjunction with FISH to two probes on the X chromosome, one from the left arm (1.5 Mb from the telomere) and one from a more medial position (7.4 Mb from the left end). The XL probe is closely associated with HIM-8 focus, while the XC probe is clearly more distant from the HIM-8 focus. Probes to the right end of the X and autosomal loci were also tested (data not shown) and did not correlate in their position with HIM-8 foci.

Picture from Phillips CM et al. (2005) Cell "HIM-8 binds to the X chromosome pairing center and mediates chromosome-specific ...."

Figure 3. HIM-8 is a C2H2 Zinc-Finger Protein that Localizes to Distinct Nuclear Foci during Meiosis. (A) Three-dimensional projection through a wild-type gonad stained with DAPI and antibodies against HIM-8. Subnuclear HIM-8 foci are present in all germline nuclei throughout the premeiotic, transition-zone, and pachytene region of the gonad. The transition-zone region outlined by the yellow box is magnified in (B).(B) Prior to meiotic entry, two HIM-8 foci (yellow) are observed in each nucleus. Examples of premeiotic nuclei in which two foci can be clearly observed are outlined in brown circles. Once nuclei have entered the transition zone, representing the leptotene/zygotene stages of meiosis, they usually reveal only a single HIM-8 focus or closely spaced pair of foci. The scale bar represents 5 μm.(C) Schematic representation of the HIM-8 protein. The diagram displays the location of two predicted zinc fingers as well as the sites of point mutations or deletions resulting from the four mutant alleles of him-8.

Picture from Phillips CM et al. (2005) Cell "HIM-8 binds to the X chromosome pairing center and mediates chromosome-specific ...."

The HIM-8 Protein Localizes to the PC Region of the X Chromosome. All images show projections through fields of pachytene-region nuclei from animals of the indicated genotypes. The entire nuclear volume is shown in each projection. All scale bars represent 5 μm.(A) Wild-type hermaphrodite, revealing a single HIM-8 focus in each nucleus due to close association between the two binding sites detected in premeiotic and very early meiotic nuclei (Figure 3).(B) Wild-type male, which displays a single (unpaired) HIM-8 focus in each nucleus. In contrast to hermaphrodites, males also have one HIM-8 focus per nucleus in the premeiotic region of the gonad (data not shown). (E) HIM-8 immunostaining was performed in conjunction with FISH to two probes on the X chromosome, one from the left arm (1.5 Mb from the telomere) and one from a more medial position (7.4 Mb from the left end). The XL probe is closely associated with HIM-8 focus, while the XC probe is clearly more distant from the HIM-8 focus. Probes to the right end of the X and autosomal loci were also tested (data not shown) and did not correlate in their position with HIM-8 foci.

Picture from Phillips CM et al. (2005) Cell "HIM-8 binds to the X chromosome pairing center and mediates chromosome-specific ...."

Figure 6. HIM-8 Foci Associate with Nuclear-Envelope Components in Both Wild-Type and him-8(me4) Animals. (A and B) Pachytene nuclei were stained with antibodies against HIM-8 (yellow) and anti-LMN-1 (red), which marks the nuclear lamina, or nuclear envelope. All scale bars represent 5 μm. See Figure S1 for an optical section through a similar data set, where nuclear-envelope association is somewhat more evident.(C) A possible role for nuclear-envelope association of meiotic chromosomes is the reduction of homology search from a 3D to a 2D problem. As discussed in the text, our analysis of the him-8(me4) mutant indicates that the requirement for HIM-8 is likely to extend beyond such a role.

Picture from Phillips BT et al. (2007) Proc Natl Acad Sci U S A "Reciprocal asymmetry of SYS-1/beta-catenin and POP-1/TCF controls asymmetric ...."

SYS-1 asymmetry during embryonic and larval development. (A-E) Nomarski (Left) and fluorescent (Right) images of same embryo or animal. (A) Early embryo. Dotted lines outline nuclei. For clarity, nuclear fluorescence is shown separately in the Inset. Images were treated identically for comparison. (B) Mid-stage embryo. SYS-1 asymmetry was observed in several daughter pairs (two are indicated). Anterior is to the left. (C) L1 larva. VNS::SYS-1 was more abundant in the posterior T cell daughter (large arrowhead) than its anterior sister (small arrowhead). (D) Developing vulva. Vulval precursors and their divisions are marked. Carats show SYS-1 asymmetry, with open end toward daughter with more SYS-1. (E) VNS::SYS-1 is enriched at a position typical of centrosomes during cell division.

Picture from Phillips CM et al. (2014) Curr Biol "MUT-14 and SMUT-1 DEAD box RNA helicases have overlapping roles in germline RNAi ...."

Figure S3. Related to Figure 4.(A) Localization of MUT-16::GFP in adult animals. The germline is outlined in magenta. Mutator foci in the germline and gut cells in theintestine are indicated.(B) Localization of MUT-7::GFP in the vulva of an adult animal. Two vulval cells are indicated.(C) Diagram of protein interactions identified in a yeast two-hybrid screen. Arrows face away from the bait. Bidirectional arrows indicate aninteraction was detected with either protein as bait.(D) Western blot analysis of MUT-16::GFP and mCherry::EGO-1 from cell lysates (in) and anti-GFP co-immunoprecipitates (IP) from rrf-1wild type (rrf-1 +/+) or mutant (rrf-1 -/-) animals. A faint mCherry::EGO-1 band was observed in the mCherry::ego-1; mut-16::GFP; rrf-1-/- IPlane after overexposing the blot to film.(E) In vitro assay for siRNA synthesis. Western blot analysis of FLAG::RRF-1 from cell lysates (in) and anti-FLAG co-immunoprecipitates(IP) from mut-16 wild type (mut-16 +/+) and mutant (mut-16 -/-) animals (top panel). Western blot analysis of MUT-14::GFP and MUT14DKAD::GFPfrom mut-14 smut-1 mutant animal cell lysates (in) and anti-GFP co-immunoprecipitates (IP) (middle panel). mut-16prom::GFPis included as a negative control. In vitro siRNA synthesis (bottom panel). Immunoprecipitated protein complexes containing FLAG::RRF-1,wild type MUT-14 (MUT-14::GFP), or catalytically dead MUT-14 (MUT-14DKAD::GFP) were incubated with a mixture of radiolabeled and nonlabelednucleotides.