Journal: Nature genetics

Article Title: Mutations in DZIP1L , which encodes a ciliary transition zone protein, cause autosomal recessive polycystic kidney disease

doi: 10.1038/ng.3871

Figure Lengend Snippet: Mapping of a new ARPKD locus on 3q22.1-q23 and identification of DZIP1L mutations. Genome-wide SNP analysis performed in two unrelated consanguineous multiplex pedigrees (shown at top) with a total of five children affected by ARPKD, resulted in identification of a single overlapping 7.5 Mb region of homozygosity on chromosome 3q22.1-q23. By various sequencing approaches, we identified different homozygous DZIP1L mutations in these and other consanguineous families with ARPKD (see text for details). In parallel, we identified an ENU-induced recessive mouse model (see Fig. 2) bearing the homozygous Dzip1l nonsense mutation c.1123C>T (p.Gln375*) (in blue).

Article Snippet: #17711-1-AP; 1:500 for IF); rabbit-anti-α-tubulin (Sigma-Aldrich T5192; 1:500 for IF); rabbit-anti-acetylated-α–tubulin (Cell Signalling Technology #5335; 1:800 for IF); mouse-anti-acetylated-α–tubulin (Sigma-Aldrich 6-11B-1; #T 6793; 1:500–700 for IF); mouse-anti-γ-tubulin (Sigma-Aldrich T6557; 1:500 for IF), rabbit-anti-DZIP1L (Sigma-Aldrich #HPA030404; 1:500 for IF); mouse-anti-DZIP1L (Abnova-MaxPab H00199221-B01P; 1:500 for IF; 1:1000 for western blot); rabbit-anti-DZIP1 (Proteintech Group, #13779-1-AP; 1:500 for IF); rabbit-anti-SEPT2 (Sigma-Aldrich, #HPA018481;1:2000 for western blot; 1:500 for IF); rabbit-anti-CEP164 (SAB3500022; Sigma), rabbit-anti-TCTN1 (ProteinTech Group, #15004-1-AP; 1:500 for IF); mouse-anti-Polycystin 1 (Abcam, #AB74115; Santa Cruz Biotechnology, # sc-130554; 1:200 for IF); rabbit-anti-Polycystin 2 (MV12; gift from R. Witzgall; 1:500 for IF); rabbit-anti-FPC (PKHD1) 9 (hAR-Np and hAR-c2P; gift from G. Wu; 1:10 and 1:20, respectively); rabbit-anti-SMO 51 was a kind gift of Rajat Rohatgi (Stanford University; 1:500 for IF); rabbit-anti-IFT88 (Proteintech 13967-1-AP; 1:300 for IF); anti-Aquaporin 2 antibody (Millipore; Cat # AB3274; 1:200 for IF); rabbit-anti-β-catenin antibody (Abcam, #AB6302; 1:500 for IF); goat-anti-HNF3β (M-20) (Santa-Cruz, #sc-6544; 1:50 for IF); goat-anti-SHH(N-19) (Santa-Cruz, #sc-1194; 1:100 for IF); the mouse monoclonal antibodies for Isl-1 (1:200 for IF), Pax6 (1:10 for IF) and Nkx2.2 (1:50 for IF) were obtained from the Developmental Studies Hybridoma Bank (developed under the auspices of the NICHD and maintained by The University of Iowa, Department of Biology, Iowa City, IA 52242.Bank); biotinylated Lotus Tetragonolobus Lectin (LTL) (Vector laboratories; Cat # B1325; 1:200).

Techniques: Genome Wide, Multiplex Assay, Sequencing, Mutagenesis

Journal: Nature genetics

Article Title: Mutations in DZIP1L , which encodes a ciliary transition zone protein, cause autosomal recessive polycystic kidney disease

doi: 10.1038/ng.3871

Figure Lengend Snippet: Mutations of DZIP1L in patients with ARPKD.

Article Snippet: #17711-1-AP; 1:500 for IF); rabbit-anti-α-tubulin (Sigma-Aldrich T5192; 1:500 for IF); rabbit-anti-acetylated-α–tubulin (Cell Signalling Technology #5335; 1:800 for IF); mouse-anti-acetylated-α–tubulin (Sigma-Aldrich 6-11B-1; #T 6793; 1:500–700 for IF); mouse-anti-γ-tubulin (Sigma-Aldrich T6557; 1:500 for IF), rabbit-anti-DZIP1L (Sigma-Aldrich #HPA030404; 1:500 for IF); mouse-anti-DZIP1L (Abnova-MaxPab H00199221-B01P; 1:500 for IF; 1:1000 for western blot); rabbit-anti-DZIP1 (Proteintech Group, #13779-1-AP; 1:500 for IF); rabbit-anti-SEPT2 (Sigma-Aldrich, #HPA018481;1:2000 for western blot; 1:500 for IF); rabbit-anti-CEP164 (SAB3500022; Sigma), rabbit-anti-TCTN1 (ProteinTech Group, #15004-1-AP; 1:500 for IF); mouse-anti-Polycystin 1 (Abcam, #AB74115; Santa Cruz Biotechnology, # sc-130554; 1:200 for IF); rabbit-anti-Polycystin 2 (MV12; gift from R. Witzgall; 1:500 for IF); rabbit-anti-FPC (PKHD1) 9 (hAR-Np and hAR-c2P; gift from G. Wu; 1:10 and 1:20, respectively); rabbit-anti-SMO 51 was a kind gift of Rajat Rohatgi (Stanford University; 1:500 for IF); rabbit-anti-IFT88 (Proteintech 13967-1-AP; 1:300 for IF); anti-Aquaporin 2 antibody (Millipore; Cat # AB3274; 1:200 for IF); rabbit-anti-β-catenin antibody (Abcam, #AB6302; 1:500 for IF); goat-anti-HNF3β (M-20) (Santa-Cruz, #sc-6544; 1:50 for IF); goat-anti-SHH(N-19) (Santa-Cruz, #sc-1194; 1:100 for IF); the mouse monoclonal antibodies for Isl-1 (1:200 for IF), Pax6 (1:10 for IF) and Nkx2.2 (1:50 for IF) were obtained from the Developmental Studies Hybridoma Bank (developed under the auspices of the NICHD and maintained by The University of Iowa, Department of Biology, Iowa City, IA 52242.Bank); biotinylated Lotus Tetragonolobus Lectin (LTL) (Vector laboratories; Cat # B1325; 1:200).

Techniques: Mutagenesis

Journal: Nature genetics

Article Title: Mutations in DZIP1L , which encodes a ciliary transition zone protein, cause autosomal recessive polycystic kidney disease

doi: 10.1038/ng.3871

Figure Lengend Snippet: DZIP1L localization overlaps with basal body, centrosome and transition zone markers. (a,b) IF staining for DZIP1L (green) and acetylated-α-tubulin and γ-tubulin (both in magenta) in serum starved IMCD3 kidney cells. DZIP1L staining overlaps with the basal body in ciliated cells (a) and the centrosome in non-ciliated cells (b). (c–d) Co-localization of DZIP1L (magenta) with the distal (CEP164, green in c) and subdistal (ODF2, green in d) appendage proteins. (e–i) DZIP1L (green) tracks with the centrioles (magenta) throughout all stages of the cell cycle in IMCD3 cells. (j) TCTN1 (magenta) and DZIP1L (cyan) co-localize at the transition zone in human dermal fibroblasts. ARL13B-GFP was used to mark the ciliary membrane around the axoneme (green in j). (k) 3D-SIM superresolution microscopy on RPE-1 (human retinal pigment epithelial) cells confirms both DZIP1L (magenta) and TCTN1 (cyan) are closely associated at the transition zone. Cilium labeled with ARL13B-GFP (green). Nuclei are stained with DAPI (blue). Scale bars in a-I = 5µm; j = 1µm; k = 500 nm. DZIP1L stained with Sigma C-terminal antibody, except in panels c,d,j,k where the Abnova antibody was used.

Article Snippet: #17711-1-AP; 1:500 for IF); rabbit-anti-α-tubulin (Sigma-Aldrich T5192; 1:500 for IF); rabbit-anti-acetylated-α–tubulin (Cell Signalling Technology #5335; 1:800 for IF); mouse-anti-acetylated-α–tubulin (Sigma-Aldrich 6-11B-1; #T 6793; 1:500–700 for IF); mouse-anti-γ-tubulin (Sigma-Aldrich T6557; 1:500 for IF), rabbit-anti-DZIP1L (Sigma-Aldrich #HPA030404; 1:500 for IF); mouse-anti-DZIP1L (Abnova-MaxPab H00199221-B01P; 1:500 for IF; 1:1000 for western blot); rabbit-anti-DZIP1 (Proteintech Group, #13779-1-AP; 1:500 for IF); rabbit-anti-SEPT2 (Sigma-Aldrich, #HPA018481;1:2000 for western blot; 1:500 for IF); rabbit-anti-CEP164 (SAB3500022; Sigma), rabbit-anti-TCTN1 (ProteinTech Group, #15004-1-AP; 1:500 for IF); mouse-anti-Polycystin 1 (Abcam, #AB74115; Santa Cruz Biotechnology, # sc-130554; 1:200 for IF); rabbit-anti-Polycystin 2 (MV12; gift from R. Witzgall; 1:500 for IF); rabbit-anti-FPC (PKHD1) 9 (hAR-Np and hAR-c2P; gift from G. Wu; 1:10 and 1:20, respectively); rabbit-anti-SMO 51 was a kind gift of Rajat Rohatgi (Stanford University; 1:500 for IF); rabbit-anti-IFT88 (Proteintech 13967-1-AP; 1:300 for IF); anti-Aquaporin 2 antibody (Millipore; Cat # AB3274; 1:200 for IF); rabbit-anti-β-catenin antibody (Abcam, #AB6302; 1:500 for IF); goat-anti-HNF3β (M-20) (Santa-Cruz, #sc-6544; 1:50 for IF); goat-anti-SHH(N-19) (Santa-Cruz, #sc-1194; 1:100 for IF); the mouse monoclonal antibodies for Isl-1 (1:200 for IF), Pax6 (1:10 for IF) and Nkx2.2 (1:50 for IF) were obtained from the Developmental Studies Hybridoma Bank (developed under the auspices of the NICHD and maintained by The University of Iowa, Department of Biology, Iowa City, IA 52242.Bank); biotinylated Lotus Tetragonolobus Lectin (LTL) (Vector laboratories; Cat # B1325; 1:200).

Techniques: Staining, Microscopy, Labeling

Journal: Nature genetics

Article Title: Mutations in DZIP1L , which encodes a ciliary transition zone protein, cause autosomal recessive polycystic kidney disease

doi: 10.1038/ng.3871

Figure Lengend Snippet: Dzip1l is required for ciliary differentiation in the zebrafish kidney tubule. (a) Primary cilia (short arrows; myotome) and mono motile cilia (long arrows; pronephric duct) in a 24 hpf wild-type embryo. (b) No obvious difference in primary (short arrows) and mono motile cilia (long arrows) differentiation in a dzip1l splice morpholino (e5i5) injected embryo at 24 hpf (n=10). (c) MCC motile cilia bundles (arrows) in the pronephric duct of a wild-type embryo at 48 hpf (arrows). (d) MCC motile cilia bundles (arrows) are reduced in a 48 hpf maternal-zygotic dzip1l mutant zebrafish embryo (n=8). (e) Quantification of MCC cilia bundles in wild-type and maternal-zygotic dzip1l mutant zebrafish embryos. 8 embryos from each group were analyzed. **p=0.0017 based on unpaired Student’s t-test. Error bars show SEM. In all panels, cilia were stained with anti-acetylated tubulin antibody (magenta) and nuclei with DAPI (blue); in panels a,b,f-i cell membranes were stained with β-catenin antibodies (green). Scale bars in a,b,f-i = 10µm; c,d = 100µm.

Article Snippet: #17711-1-AP; 1:500 for IF); rabbit-anti-α-tubulin (Sigma-Aldrich T5192; 1:500 for IF); rabbit-anti-acetylated-α–tubulin (Cell Signalling Technology #5335; 1:800 for IF); mouse-anti-acetylated-α–tubulin (Sigma-Aldrich 6-11B-1; #T 6793; 1:500–700 for IF); mouse-anti-γ-tubulin (Sigma-Aldrich T6557; 1:500 for IF), rabbit-anti-DZIP1L (Sigma-Aldrich #HPA030404; 1:500 for IF); mouse-anti-DZIP1L (Abnova-MaxPab H00199221-B01P; 1:500 for IF; 1:1000 for western blot); rabbit-anti-DZIP1 (Proteintech Group, #13779-1-AP; 1:500 for IF); rabbit-anti-SEPT2 (Sigma-Aldrich, #HPA018481;1:2000 for western blot; 1:500 for IF); rabbit-anti-CEP164 (SAB3500022; Sigma), rabbit-anti-TCTN1 (ProteinTech Group, #15004-1-AP; 1:500 for IF); mouse-anti-Polycystin 1 (Abcam, #AB74115; Santa Cruz Biotechnology, # sc-130554; 1:200 for IF); rabbit-anti-Polycystin 2 (MV12; gift from R. Witzgall; 1:500 for IF); rabbit-anti-FPC (PKHD1) 9 (hAR-Np and hAR-c2P; gift from G. Wu; 1:10 and 1:20, respectively); rabbit-anti-SMO 51 was a kind gift of Rajat Rohatgi (Stanford University; 1:500 for IF); rabbit-anti-IFT88 (Proteintech 13967-1-AP; 1:300 for IF); anti-Aquaporin 2 antibody (Millipore; Cat # AB3274; 1:200 for IF); rabbit-anti-β-catenin antibody (Abcam, #AB6302; 1:500 for IF); goat-anti-HNF3β (M-20) (Santa-Cruz, #sc-6544; 1:50 for IF); goat-anti-SHH(N-19) (Santa-Cruz, #sc-1194; 1:100 for IF); the mouse monoclonal antibodies for Isl-1 (1:200 for IF), Pax6 (1:10 for IF) and Nkx2.2 (1:50 for IF) were obtained from the Developmental Studies Hybridoma Bank (developed under the auspices of the NICHD and maintained by The University of Iowa, Department of Biology, Iowa City, IA 52242.Bank); biotinylated Lotus Tetragonolobus Lectin (LTL) (Vector laboratories; Cat # B1325; 1:200).

Techniques: Injection, Mutagenesis, Staining

Journal: Nature genetics

Article Title: Mutations in DZIP1L , which encodes a ciliary transition zone protein, cause autosomal recessive polycystic kidney disease

doi: 10.1038/ng.3871

Figure Lengend Snippet: DZIP1L associates with the ciliary transition zone protein SEPT2. (a) Schematic diagram of SEPT2, DZIP1L and a series of DZIP1L deletion mutants. (b) Interaction between SEPT2 (N-terminal FLAG) and DZIP1L and various DZIP1L deletion mutants (C-terminal HA) as determined by IP following transfection in HEK293T cells. (c,d) Endogenous interaction between DZIP1L and SEPT2 in RPE-1 cells. IP was performed with either the anti-DZIP1L antibody (Abnova; c) or the anti-SEPT2 antibody (d). (e) Expression of DZIP1L and SEPT2 in total cell lysate. (f) Co-localization of DZIP1L (cyan) and SEPT2 (magenta) in human dermal fibroblasts. ARL13B-GFP was used to mark the ciliary axoneme (green). (g,h) SEPT2 (green) localization to the transition zone was unaffected in DZIP1L mutant dermal fibroblasts from individual B155 (p.Gln155*). Ciliary axonemes and basal bodies were labelled with anti-acetylated tubulin and anti-γ-tubulin antibodies (magenta), respectively. Scale bars in f-h = 1µm. All co-IP experiments were repeated at least three times.

Article Snippet: #17711-1-AP; 1:500 for IF); rabbit-anti-α-tubulin (Sigma-Aldrich T5192; 1:500 for IF); rabbit-anti-acetylated-α–tubulin (Cell Signalling Technology #5335; 1:800 for IF); mouse-anti-acetylated-α–tubulin (Sigma-Aldrich 6-11B-1; #T 6793; 1:500–700 for IF); mouse-anti-γ-tubulin (Sigma-Aldrich T6557; 1:500 for IF), rabbit-anti-DZIP1L (Sigma-Aldrich #HPA030404; 1:500 for IF); mouse-anti-DZIP1L (Abnova-MaxPab H00199221-B01P; 1:500 for IF; 1:1000 for western blot); rabbit-anti-DZIP1 (Proteintech Group, #13779-1-AP; 1:500 for IF); rabbit-anti-SEPT2 (Sigma-Aldrich, #HPA018481;1:2000 for western blot; 1:500 for IF); rabbit-anti-CEP164 (SAB3500022; Sigma), rabbit-anti-TCTN1 (ProteinTech Group, #15004-1-AP; 1:500 for IF); mouse-anti-Polycystin 1 (Abcam, #AB74115; Santa Cruz Biotechnology, # sc-130554; 1:200 for IF); rabbit-anti-Polycystin 2 (MV12; gift from R. Witzgall; 1:500 for IF); rabbit-anti-FPC (PKHD1) 9 (hAR-Np and hAR-c2P; gift from G. Wu; 1:10 and 1:20, respectively); rabbit-anti-SMO 51 was a kind gift of Rajat Rohatgi (Stanford University; 1:500 for IF); rabbit-anti-IFT88 (Proteintech 13967-1-AP; 1:300 for IF); anti-Aquaporin 2 antibody (Millipore; Cat # AB3274; 1:200 for IF); rabbit-anti-β-catenin antibody (Abcam, #AB6302; 1:500 for IF); goat-anti-HNF3β (M-20) (Santa-Cruz, #sc-6544; 1:50 for IF); goat-anti-SHH(N-19) (Santa-Cruz, #sc-1194; 1:100 for IF); the mouse monoclonal antibodies for Isl-1 (1:200 for IF), Pax6 (1:10 for IF) and Nkx2.2 (1:50 for IF) were obtained from the Developmental Studies Hybridoma Bank (developed under the auspices of the NICHD and maintained by The University of Iowa, Department of Biology, Iowa City, IA 52242.Bank); biotinylated Lotus Tetragonolobus Lectin (LTL) (Vector laboratories; Cat # B1325; 1:200).

Techniques: Transfection, Expressing, Mutagenesis, Co-Immunoprecipitation Assay

Journal: Nature genetics

Article Title: Mutations in DZIP1L , which encodes a ciliary transition zone protein, cause autosomal recessive polycystic kidney disease

doi: 10.1038/ng.3871

Figure Lengend Snippet: Loss of DZIP1L affects the localization of PC1 and PC2 to the ciliary membrane. (a) In most wild-type ciliated MEFs PC1 (green; 7e12 mAb, Abcam) staining was distributed along the length of the axoneme as marked with ARL13B (magenta). (b) By contrast PC1 staining was more often concentrated at the proximal end of the axoneme in Dzip1lwpy/wpy MEFs (c) Quantification of the percentage of ciliated cells with PC1 along the axoneme (n=6, derived from 3 MEF cell lines from independent embryos, each counted in two separate experiments; between 50–185 cells counted per replicate, total 737 Dzip1l+/+ and 649 Dzip1lwpy/wpy cells counted). (d,e) Decreased PC1 (green) localization in the ciliary membrane in human dermal fibroblasts from affected individual B155 (p.Gln155*). ARL13B (magenta) marks the ciliary axoneme. (g,h) Decreased PC2 localization (green) in the ciliary membrane in DZIP1L mutant human dermal fibroblasts. Acetylated-α–tubulin and γ-tubulin (magenta) mark the cilia. (f,i) Quantification of the percentage of ciliated cells with PC1 and PC2 along the axoneme, respectively, in control and DZIP1L mutant human dermal fibroblasts. For human cells, cilia were counted in three experiments, with cells from three independent coverslips counted for each experiment (approximately 100 ciliated cells counted on each coverslip). In all cases, quantification included cilia with or without detectable PC1 or PC2 staining. Statistical analyses based on unpaired Student’s t-test. Error bars show SEM. For PC1 staining on MEFs, ****p<0.0001; PC1 and PC2 staining on human dermal fibroblasts, ***p<0.001. Scale bars in a,b = 2µm; d,e,g,h = 1µm.

Article Snippet: #17711-1-AP; 1:500 for IF); rabbit-anti-α-tubulin (Sigma-Aldrich T5192; 1:500 for IF); rabbit-anti-acetylated-α–tubulin (Cell Signalling Technology #5335; 1:800 for IF); mouse-anti-acetylated-α–tubulin (Sigma-Aldrich 6-11B-1; #T 6793; 1:500–700 for IF); mouse-anti-γ-tubulin (Sigma-Aldrich T6557; 1:500 for IF), rabbit-anti-DZIP1L (Sigma-Aldrich #HPA030404; 1:500 for IF); mouse-anti-DZIP1L (Abnova-MaxPab H00199221-B01P; 1:500 for IF; 1:1000 for western blot); rabbit-anti-DZIP1 (Proteintech Group, #13779-1-AP; 1:500 for IF); rabbit-anti-SEPT2 (Sigma-Aldrich, #HPA018481;1:2000 for western blot; 1:500 for IF); rabbit-anti-CEP164 (SAB3500022; Sigma), rabbit-anti-TCTN1 (ProteinTech Group, #15004-1-AP; 1:500 for IF); mouse-anti-Polycystin 1 (Abcam, #AB74115; Santa Cruz Biotechnology, # sc-130554; 1:200 for IF); rabbit-anti-Polycystin 2 (MV12; gift from R. Witzgall; 1:500 for IF); rabbit-anti-FPC (PKHD1) 9 (hAR-Np and hAR-c2P; gift from G. Wu; 1:10 and 1:20, respectively); rabbit-anti-SMO 51 was a kind gift of Rajat Rohatgi (Stanford University; 1:500 for IF); rabbit-anti-IFT88 (Proteintech 13967-1-AP; 1:300 for IF); anti-Aquaporin 2 antibody (Millipore; Cat # AB3274; 1:200 for IF); rabbit-anti-β-catenin antibody (Abcam, #AB6302; 1:500 for IF); goat-anti-HNF3β (M-20) (Santa-Cruz, #sc-6544; 1:50 for IF); goat-anti-SHH(N-19) (Santa-Cruz, #sc-1194; 1:100 for IF); the mouse monoclonal antibodies for Isl-1 (1:200 for IF), Pax6 (1:10 for IF) and Nkx2.2 (1:50 for IF) were obtained from the Developmental Studies Hybridoma Bank (developed under the auspices of the NICHD and maintained by The University of Iowa, Department of Biology, Iowa City, IA 52242.Bank); biotinylated Lotus Tetragonolobus Lectin (LTL) (Vector laboratories; Cat # B1325; 1:200).

Techniques: Staining, Derivative Assay, Mutagenesis

Journal: Pediatric Nephrology (Berlin, Germany)

Article Title: Detection of DZIP1L mutations by whole-exome sequencing in consanguineous families with polycystic kidney disease

doi: 10.1007/s00467-022-05441-4

Figure Lengend Snippet: Pedigrees of the families. A : Family 1 . The two affected children (V:3 and V:6) are both homozygous for the DZIP1L sequence variant: c.216C > G; p.(Cys72Trp). a : Ultrasound scan of the left kidney in V:3 and b : Ultrasound scan of the left kidney in V:6. B . Family 2. The proband (III:3) is homozygous for the same sequence variant as identified in family 1 : c.216C > G; p.(Cys72Trp). C. Family 3 . The proband (VI:2) is homozygous for the DZIP1L sequence variant: c.193T > C; p.(Cys65Arg). Filled symbols indicate homozygous individuals, and half-filled symbols indicate heterozygous relatives. a : MR-scan of the kidneys from VI:2 showing enlarged kidneys with multiple cysts. b : Histological sections from the right kidney in HE staining (VI:2): Cortical and medulla areas with cysts. Subcapsular, a small area shows a more normal structure. For details, see text. Arrows indicate the probands

Article Snippet: Cells were washed in PBS and incubated with mouse polyclonal anti-human DZIP1L protein antibody (1:500, H00199221-B01P, Abnova) overnight at 4 °C.

Techniques: Sequencing, Variant Assay, Staining

Journal: Pediatric Nephrology (Berlin, Germany)

Article Title: Detection of DZIP1L mutations by whole-exome sequencing in consanguineous families with polycystic kidney disease

doi: 10.1007/s00467-022-05441-4

Figure Lengend Snippet: A HEK293 cells transiently transfected with GFP-DZIP1L WT, Cys72Trp and mGFP were labeled with an anti-DZIP1L antibody. The anti-DZIP1L antibody only labeled cells that were transfected with the DZIP1L constructs. Scale bar: 20 µm. B Normal adult human kidney samples were stained with the same anti-DZIP1L antibody. Arrows indicate strong punctuate staining observed in select cells. Arrowheads indicate weaker and more diffuse staining, mainly found in collecting duct cells. C Double staining with anti-DZIP1L (red) and AQP2 (green) shows that the strong punctuate staining is not restricted to AQP2 expressing cells in the collecting system

Article Snippet: Cells were washed in PBS and incubated with mouse polyclonal anti-human DZIP1L protein antibody (1:500, H00199221-B01P, Abnova) overnight at 4 °C.

Techniques: Transfection, Labeling, Construct, Staining, Double Staining, Expressing

Journal: Pediatric Nephrology (Berlin, Germany)

Article Title: Detection of DZIP1L mutations by whole-exome sequencing in consanguineous families with polycystic kidney disease

doi: 10.1007/s00467-022-05441-4

Figure Lengend Snippet: The six DZIP1L sequence variants reported to date. All detected in homozygous form

Article Snippet: Cells were washed in PBS and incubated with mouse polyclonal anti-human DZIP1L protein antibody (1:500, H00199221-B01P, Abnova) overnight at 4 °C.

Techniques: Sequencing

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: The ARPKD Protein DZIP1L Regulates Ciliary Protein Entry by Modulating the Architecture and Function of Ciliary Transition Fibers.

doi: 10.1002/advs.202308820

Figure Lengend Snippet: Figure 1. C. elegans DZIP-1 is an evolutionarily conserved basal body protein. A) The sequence alignment of two evolutionarily conserved domain (DZIP- like domain and the C2H2 zine finger domain) within C. elegans DZIP-1, H. sapiens DZIP1/DZIP1L and M. musculus DZIP1/DZIP1L. The Dzip-like_N domain (red line) is the most conserved domain in DZIP1/DZIP1L proteins. The ZnF_C2H2 domain is indicated by an orange line. And the highly conserved region is enclosed in the purple box. Asterisks are used to indicate the residues that, when mutated, can cause human ARPKD. B) DZIP-1 localizes to the ciliary base in C. elegans. The schematic diagram of phasmid cilia is depicted on the left, GFP-tagged DZIP-1 specifically localizes to

Article Snippet: Protein localization was confirmed by immunofluorescence imaging. gRNA sequence was as follows: DZIP1L gRNA#1, 5′- TGATCATCTTGCGACGCCGG-3′; DZIP1L gRNA#2, 5′- CAGTCCATGCTATCATGGCG-3′; ANKRD26 gRNA#1, 5′-GGGTAGCTCACAATCCTCTG-3′; ANKRD26 gRNA#2, 5′- AAATTCTTGTAACTAGAGTG-3′; Antibodies: The following primary antibodies were used in this study: ANKRD26 (rabbit, GeneTex, GTX128255, 1:500), DZIP1L (rabbit, Proteintech, 17474-1-AP, 1:500), FBF1 (rabbit, Proteintech, 11531-1-AP, 1:500), ARL13B (rabbit, Proteintech, 17711-1-AP, 1:500), KIAA0586/TALPID3 (rabbit, Proteintech, 24421-1-AP, 1:500), SCLT1 (rabbit, Proteintech, 14875- 1-AP, 1:500), CEP164 (rabbit, Proteintech, 22227-1-AP, 1:500), CEP89 (rabbit, Abcam, ab204410, 1:500), ODF2 (rabbit, Abcam, ab43840, 1:500), ODF2 (mouse, Abnova, H00004957-M01, 1:500), γ-tublin (mouse, Sigma-Aldrich, T6557, 1:1,000), FAM92A (rabbit, Proteintech, 24803-1- AP, 1:500), CBY1 (rabbit, Proteintech, 12239-1-AP, 1:500), GFP(mouse, Roche, 11814460001,1:200), IFT140 (rabbit, Proteintech, 17460-1-AP, 1:500), GT335(mouse, adipogen life science, A40251903), GLI3 (AF3690, R&D Systems), PC2 (Baltimore Polycystic Kidney Disease (PKD) Research and Clinical Core Center), Ac-tubulin (mouse, sigma, T7451, 1:300), Flag (mouse, sigma, F1804, 1:300), CEP290 (rabbit, abcam, ab84870).

Techniques: Sequencing

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: The ARPKD Protein DZIP1L Regulates Ciliary Protein Entry by Modulating the Architecture and Function of Ciliary Transition Fibers.

doi: 10.1002/advs.202308820

Figure Lengend Snippet: Figure 6. The precise localization of DZIP1L in human RPE cells. Spatial localization of DZIP1L was examined using a combination of Ultrastructure expansion microscopy (ExM) and confocal microscopy. As our DZIP1L polyclonal antibody is derived from rabbits, and most of our antibodies for TF and TZ markers are also polyclonal antibodies from rabbit, co-labeling them with antibodies is not feasible. To assess their co-localization, we employed transgenic FLAG-DZIP1L and utilized a mouse monoclonal FLAG antibody (IgG1 isotype) to label DZIP1L. While cilia were marked using anti-Ac-tubulin, a mouse monoclonal antibody with IgG2b isotype. A) FLAG-DZIP1L was localized beneath the classical TZ component CEP290 in both ciliated and non-

Article Snippet: Protein localization was confirmed by immunofluorescence imaging. gRNA sequence was as follows: DZIP1L gRNA#1, 5′- TGATCATCTTGCGACGCCGG-3′; DZIP1L gRNA#2, 5′- CAGTCCATGCTATCATGGCG-3′; ANKRD26 gRNA#1, 5′-GGGTAGCTCACAATCCTCTG-3′; ANKRD26 gRNA#2, 5′- AAATTCTTGTAACTAGAGTG-3′; Antibodies: The following primary antibodies were used in this study: ANKRD26 (rabbit, GeneTex, GTX128255, 1:500), DZIP1L (rabbit, Proteintech, 17474-1-AP, 1:500), FBF1 (rabbit, Proteintech, 11531-1-AP, 1:500), ARL13B (rabbit, Proteintech, 17711-1-AP, 1:500), KIAA0586/TALPID3 (rabbit, Proteintech, 24421-1-AP, 1:500), SCLT1 (rabbit, Proteintech, 14875- 1-AP, 1:500), CEP164 (rabbit, Proteintech, 22227-1-AP, 1:500), CEP89 (rabbit, Abcam, ab204410, 1:500), ODF2 (rabbit, Abcam, ab43840, 1:500), ODF2 (mouse, Abnova, H00004957-M01, 1:500), γ-tublin (mouse, Sigma-Aldrich, T6557, 1:1,000), FAM92A (rabbit, Proteintech, 24803-1- AP, 1:500), CBY1 (rabbit, Proteintech, 12239-1-AP, 1:500), GFP(mouse, Roche, 11814460001,1:200), IFT140 (rabbit, Proteintech, 17460-1-AP, 1:500), GT335(mouse, adipogen life science, A40251903), GLI3 (AF3690, R&D Systems), PC2 (Baltimore Polycystic Kidney Disease (PKD) Research and Clinical Core Center), Ac-tubulin (mouse, sigma, T7451, 1:300), Flag (mouse, sigma, F1804, 1:300), CEP290 (rabbit, abcam, ab84870).

Techniques: Microscopy, Confocal Microscopy, Derivative Assay, Labeling, Transgenic Assay

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: The ARPKD Protein DZIP1L Regulates Ciliary Protein Entry by Modulating the Architecture and Function of Ciliary Transition Fibers.

doi: 10.1002/advs.202308820

Figure Lengend Snippet: Figure 7. Conserved role of DZIP1L-ANKRD26 module in cilia gating in human RPE cells. A) The genetic interaction between DZIP1L and ANKRD26 in cilia formation is conserved in mammalian cells. The knockouts of either DZIP1L or ANKRD26 led to a moderate decrease in ciliation ratio. However, when both DZIP1L and ANKRD26 were knocked out, cilia formation was nearly completely blocked. The ciliation ratio was shown in the lower panel. Data are presented as mean ± SEM. n > 300 cells from three independent experiments. Significant differences were determined by two tailed t-test analysis. Scale bars: 5μm. B) Ciliary entry of the IFT component IFT140 was compromised in double mutants of DZIP1L and ANKRD26, compared to either of

Article Snippet: Protein localization was confirmed by immunofluorescence imaging. gRNA sequence was as follows: DZIP1L gRNA#1, 5′- TGATCATCTTGCGACGCCGG-3′; DZIP1L gRNA#2, 5′- CAGTCCATGCTATCATGGCG-3′; ANKRD26 gRNA#1, 5′-GGGTAGCTCACAATCCTCTG-3′; ANKRD26 gRNA#2, 5′- AAATTCTTGTAACTAGAGTG-3′; Antibodies: The following primary antibodies were used in this study: ANKRD26 (rabbit, GeneTex, GTX128255, 1:500), DZIP1L (rabbit, Proteintech, 17474-1-AP, 1:500), FBF1 (rabbit, Proteintech, 11531-1-AP, 1:500), ARL13B (rabbit, Proteintech, 17711-1-AP, 1:500), KIAA0586/TALPID3 (rabbit, Proteintech, 24421-1-AP, 1:500), SCLT1 (rabbit, Proteintech, 14875- 1-AP, 1:500), CEP164 (rabbit, Proteintech, 22227-1-AP, 1:500), CEP89 (rabbit, Abcam, ab204410, 1:500), ODF2 (rabbit, Abcam, ab43840, 1:500), ODF2 (mouse, Abnova, H00004957-M01, 1:500), γ-tublin (mouse, Sigma-Aldrich, T6557, 1:1,000), FAM92A (rabbit, Proteintech, 24803-1- AP, 1:500), CBY1 (rabbit, Proteintech, 12239-1-AP, 1:500), GFP(mouse, Roche, 11814460001,1:200), IFT140 (rabbit, Proteintech, 17460-1-AP, 1:500), GT335(mouse, adipogen life science, A40251903), GLI3 (AF3690, R&D Systems), PC2 (Baltimore Polycystic Kidney Disease (PKD) Research and Clinical Core Center), Ac-tubulin (mouse, sigma, T7451, 1:300), Flag (mouse, sigma, F1804, 1:300), CEP290 (rabbit, abcam, ab84870).

Techniques: Two Tailed Test

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: The ARPKD Protein DZIP1L Regulates Ciliary Protein Entry by Modulating the Architecture and Function of Ciliary Transition Fibers.

doi: 10.1002/advs.202308820

Figure Lengend Snippet: Figure 7. Conserved role of DZIP1L-ANKRD26 module in cilia gating in human RPE cells. A) The genetic interaction between DZIP1L and ANKRD26 in cilia formation is conserved in mammalian cells. The knockouts of either DZIP1L or ANKRD26 led to a moderate decrease in ciliation ratio. However, when both DZIP1L and ANKRD26 were knocked out, cilia formation was nearly completely blocked. The ciliation ratio was shown in the lower panel. Data are presented as mean ± SEM. n > 300 cells from three independent experiments. Significant differences were determined by two tailed t-test analysis. Scale bars: 5μm. B) Ciliary entry of the IFT component IFT140 was compromised in double mutants of DZIP1L and ANKRD26, compared to either of

Article Snippet: Protein localization was confirmed by immunofluorescence imaging. gRNA sequence was as follows: DZIP1L gRNA#1, 5′- TGATCATCTTGCGACGCCGG-3′; DZIP1L gRNA#2, 5′- CAGTCCATGCTATCATGGCG-3′; ANKRD26 gRNA#1, 5′-GGGTAGCTCACAATCCTCTG-3′; ANKRD26 gRNA#2, 5′- AAATTCTTGTAACTAGAGTG-3′; Antibodies: The following primary antibodies were used in this study: ANKRD26 (rabbit, GeneTex, GTX128255, 1:500), DZIP1L (rabbit, Proteintech, 17474-1-AP, 1:500), FBF1 (rabbit, Proteintech, 11531-1-AP, 1:500), ARL13B (rabbit, Proteintech, 17711-1-AP, 1:500), KIAA0586/TALPID3 (rabbit, Proteintech, 24421-1-AP, 1:500), SCLT1 (rabbit, Proteintech, 14875- 1-AP, 1:500), CEP164 (rabbit, Proteintech, 22227-1-AP, 1:500), CEP89 (rabbit, Abcam, ab204410, 1:500), ODF2 (rabbit, Abcam, ab43840, 1:500), ODF2 (mouse, Abnova, H00004957-M01, 1:500), γ-tublin (mouse, Sigma-Aldrich, T6557, 1:1,000), FAM92A (rabbit, Proteintech, 24803-1- AP, 1:500), CBY1 (rabbit, Proteintech, 12239-1-AP, 1:500), GFP(mouse, Roche, 11814460001,1:200), IFT140 (rabbit, Proteintech, 17460-1-AP, 1:500), GT335(mouse, adipogen life science, A40251903), GLI3 (AF3690, R&D Systems), PC2 (Baltimore Polycystic Kidney Disease (PKD) Research and Clinical Core Center), Ac-tubulin (mouse, sigma, T7451, 1:300), Flag (mouse, sigma, F1804, 1:300), CEP290 (rabbit, abcam, ab84870).

Techniques: Two Tailed Test

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: The ARPKD Protein DZIP1L Regulates Ciliary Protein Entry by Modulating the Architecture and Function of Ciliary Transition Fibers.

doi: 10.1002/advs.202308820

Figure Lengend Snippet: Figure 7. Conserved role of DZIP1L-ANKRD26 module in cilia gating in human RPE cells. A) The genetic interaction between DZIP1L and ANKRD26 in cilia formation is conserved in mammalian cells. The knockouts of either DZIP1L or ANKRD26 led to a moderate decrease in ciliation ratio. However, when both DZIP1L and ANKRD26 were knocked out, cilia formation was nearly completely blocked. The ciliation ratio was shown in the lower panel. Data are presented as mean ± SEM. n > 300 cells from three independent experiments. Significant differences were determined by two tailed t-test analysis. Scale bars: 5μm. B) Ciliary entry of the IFT component IFT140 was compromised in double mutants of DZIP1L and ANKRD26, compared to either of

Article Snippet: Protein localization was confirmed by immunofluorescence imaging. gRNA sequence was as follows: DZIP1L gRNA#1, 5′- TGATCATCTTGCGACGCCGG-3′; DZIP1L gRNA#2, 5′- CAGTCCATGCTATCATGGCG-3′; ANKRD26 gRNA#1, 5′-GGGTAGCTCACAATCCTCTG-3′; ANKRD26 gRNA#2, 5′- AAATTCTTGTAACTAGAGTG-3′; Antibodies: The following primary antibodies were used in this study: ANKRD26 (rabbit, GeneTex, GTX128255, 1:500), DZIP1L (rabbit, Proteintech, 17474-1-AP, 1:500), FBF1 (rabbit, Proteintech, 11531-1-AP, 1:500), ARL13B (rabbit, Proteintech, 17711-1-AP, 1:500), KIAA0586/TALPID3 (rabbit, Proteintech, 24421-1-AP, 1:500), SCLT1 (rabbit, Proteintech, 14875- 1-AP, 1:500), CEP164 (rabbit, Proteintech, 22227-1-AP, 1:500), CEP89 (rabbit, Abcam, ab204410, 1:500), ODF2 (rabbit, Abcam, ab43840, 1:500), ODF2 (mouse, Abnova, H00004957-M01, 1:500), γ-tublin (mouse, Sigma-Aldrich, T6557, 1:1,000), FAM92A (rabbit, Proteintech, 24803-1- AP, 1:500), CBY1 (rabbit, Proteintech, 12239-1-AP, 1:500), GFP(mouse, Roche, 11814460001,1:200), IFT140 (rabbit, Proteintech, 17460-1-AP, 1:500), GT335(mouse, adipogen life science, A40251903), GLI3 (AF3690, R&D Systems), PC2 (Baltimore Polycystic Kidney Disease (PKD) Research and Clinical Core Center), Ac-tubulin (mouse, sigma, T7451, 1:300), Flag (mouse, sigma, F1804, 1:300), CEP290 (rabbit, abcam, ab84870).

Techniques: Two Tailed Test

Journal: Nature genetics

Article Title: Mutations in DZIP1L , which encodes a ciliary transition zone protein, cause autosomal recessive polycystic kidney disease

doi: 10.1038/ng.3871

Figure Lengend Snippet: DZIP1L localization overlaps with basal body, centrosome and transition zone markers. (a,b) IF staining for DZIP1L (green) and acetylated-α-tubulin and γ-tubulin (both in magenta) in serum starved IMCD3 kidney cells. DZIP1L staining overlaps with the basal body in ciliated cells (a) and the centrosome in non-ciliated cells (b). (c–d) Co-localization of DZIP1L (magenta) with the distal (CEP164, green in c) and subdistal (ODF2, green in d) appendage proteins. (e–i) DZIP1L (green) tracks with the centrioles (magenta) throughout all stages of the cell cycle in IMCD3 cells. (j) TCTN1 (magenta) and DZIP1L (cyan) co-localize at the transition zone in human dermal fibroblasts. ARL13B-GFP was used to mark the ciliary membrane around the axoneme (green in j). (k) 3D-SIM superresolution microscopy on RPE-1 (human retinal pigment epithelial) cells confirms both DZIP1L (magenta) and TCTN1 (cyan) are closely associated at the transition zone. Cilium labeled with ARL13B-GFP (green). Nuclei are stained with DAPI (blue). Scale bars in a-I = 5µm; j = 1µm; k = 500 nm. DZIP1L stained with Sigma C-terminal antibody, except in panels c,d,j,k where the Abnova antibody was used.

Article Snippet: #17711-1-AP; 1:500 for IF); rabbit-anti-α-tubulin (Sigma-Aldrich T5192; 1:500 for IF); rabbit-anti-acetylated-α–tubulin (Cell Signalling Technology #5335; 1:800 for IF); mouse-anti-acetylated-α–tubulin (Sigma-Aldrich 6-11B-1; #T 6793; 1:500–700 for IF); mouse-anti-γ-tubulin (Sigma-Aldrich T6557; 1:500 for IF), rabbit-anti-DZIP1L (Sigma-Aldrich #HPA030404; 1:500 for IF); mouse-anti-DZIP1L (Abnova-MaxPab H00199221-B01P; 1:500 for IF; 1:1000 for western blot); rabbit-anti-DZIP1 (Proteintech Group, #13779-1-AP; 1:500 for IF); rabbit-anti-SEPT2 (Sigma-Aldrich, #HPA018481;1:2000 for western blot; 1:500 for IF); rabbit-anti-CEP164 (SAB3500022; Sigma), rabbit-anti-TCTN1 (ProteinTech Group, #15004-1-AP; 1:500 for IF); mouse-anti-Polycystin 1 (Abcam, #AB74115; Santa Cruz Biotechnology, # sc-130554; 1:200 for IF); rabbit-anti-Polycystin 2 (MV12; gift from R. Witzgall; 1:500 for IF); rabbit-anti-FPC (PKHD1) 9 (hAR-Np and hAR-c2P; gift from G. Wu; 1:10 and 1:20, respectively); rabbit-anti-SMO 51 was a kind gift of Rajat Rohatgi (Stanford University; 1:500 for IF); rabbit-anti-IFT88 (Proteintech 13967-1-AP; 1:300 for IF); anti-Aquaporin 2 antibody (Millipore; Cat # AB3274; 1:200 for IF); rabbit-anti-β-catenin antibody (Abcam, #AB6302; 1:500 for IF); goat-anti-HNF3β (M-20) (Santa-Cruz, #sc-6544; 1:50 for IF); goat-anti-SHH(N-19) (Santa-Cruz, #sc-1194; 1:100 for IF); the mouse monoclonal antibodies for Isl-1 (1:200 for IF), Pax6 (1:10 for IF) and Nkx2.2 (1:50 for IF) were obtained from the Developmental Studies Hybridoma Bank (developed under the auspices of the NICHD and maintained by The University of Iowa, Department of Biology, Iowa City, IA 52242.Bank); biotinylated Lotus Tetragonolobus Lectin (LTL) (Vector laboratories; Cat # B1325; 1:200).

Techniques: Staining, Microscopy, Labeling