run_metadata: 28783
This data as json
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| 28783 | SRR26639299 | SRX22339671 | SRS19389320 | SRP469870 | PRJNA1035005 | Loss of Nkd1 is dominant over loss of Axin2 in regulating Wnt signaling. | GSE246858 | Transcriptome Analysis | Wnt signaling is a crucial developmental pathway involved in early development as well as stem cell maintenance in adults and its misregulation leads to numerous diseases. Thus understanding the regulation of this pathway becomes vitally important. Axin2 and Nkd1 are widely utilized negative feedback regulators in Wnt signaling where Axin2 functions to destabilize cytoplasmic ß catenin and Nkd1 functions to inhibit the nuclear localization of ß catenin. Here we set out to further understand how Axin2 and Nkd1 regulate Wnt signaling by creating axin2 / nkd1 / single mutants and axin2 / ;nkd1 / double mutant zebrafish using sgRNA/Cas9. All three Wnt regulator mutants were viable and had impaired heart looping neuromast migration defects and behavior abnormalities in common but there were no signs of synergy in the axin2 / ;nkd1 / double mutants. Further Wnt target gene expression by qRT PCR and RNA seq analysis and protein expression by mass spec demonstrated that the double axin2 / ;nkd1 / mutant resembled the nkd1 / phenotype demonstrating that Axin functions upstream of Nkd1 and that loss of Nkd1 is dominant over the loss of Axin2. In support of this the data further demonstrates that Axin2 uniquely alters the properties of ß catenin dependent transcription having novel readouts of Wnt activity compared to nkd1 / or the axin2 / ;nkd1 / double mutant. We also tested the sensitivity of the Wnt regulator mutants to exacerbated Wnt signaling where the single mutants displayed characteristic heightened Wnt sensitivity resulting in an eyeless phenotype. Surprisingly this phenotype was rescued in the double mutant where we speculate that cross talk between Wnt/ß catenin and Wnt/Planar Cell Polarity pathways could lead to altered Wnt signaling in some scenarios. Collectively the data emphasizes both the commonality and the complexity in the feedback regulation of Wnt signaling. Overall design: To investigate Axin2 and Nkd1 regulation of Wnt signaling axin2 / and nkd1 / single knockout zebrafish were created using sgRNA/Cas9. These were crossed together to create axin2 / ;nkd1 / double knockout zebrafish. The nkd1 / zebrafish were made in the Tu genetic background and the axin2 / were made in the TL genetic background. The RNA seq samples were collected at 50% epiboly with biological replicates coming from the same parents but collected on different days. Gene expression analysis of the RNA seq data was used to determine that the nkd1 / single mutant was similar to axin2 / ;nkd1 / double mutant. Furthermore gene expression analysis was used to assess Wnt target gene expression in the mutants | Tu rep2 | GSM7880089 | source name:50% epiboly|tissue:50% epiboly|geo loc name:missing|collection date:missing | Tu rep2 | Rsubread v2.2.6 reads were aligned using the Rsubread align function to the Ensembl Genome Browser assembly ID: GRCz11 reads were filtered using EdgeR 3.30.3 filterByExpr function Assembly: GRCz11 Supplementary files format and content: csv for counts of all genotypes | 50% epiboly | 10 embryos at 50% epiboly were extracted using the FroggaBio GENEzol TriRNA pure kit following manufacturers instructions. NEBNext Ultra II DNA library prep kit for illumina New England BioLabs | tissue:50% epiboly | GSM7880089 | GSM7880089: Tu rep2; Danio rerio; RNA Seq | GSM7880089 r1 | GSM7880089 | 1 | 10 embryos at 50% epiboly were extracted using the FroggaBio GENEzol TriRNA pure kit following manufacturers instructions. NEBNext Ultra II DNA library prep kit for illumina New England BioLabs | RNA-Seq | TRANSCRIPTOMIC | cDNA | PAIRED | ILLUMINA | Illumina NovaSeq 6000 | SRP469870 | loader:fastq load.py | Tu_2_R1.fastq.gz Tu_2_R2.fastq.gz | fastq fastq | 18943158078.0 | 62725689.0 | GSM7880089 r1 | 0:151 1:151 | A:5140123698;C:4366290874;G:4396999060;T:5039656619;N:87827 | 151 | 151 | 5140123698 | 4366290874 | 4396999060 | 5039656619 | 87827 | SRX22339671 | SRS19389320 | SRA1744390 | MCB, University of Guelph | MCB, University of Guelph | 2 | 0.973 | 0.97483 | 0.07443 | 0.07095 | 0.7567 | 0.75599 | 0.48669 | 0.48759 | 151 | 151 | B | B | biological fallback assumption | illumina | novaseq_era | unknown | cdna_unspecified | nebnext | bulk | unknown | unknown | Canada | 2023-11-02 | Gastrula | Embryo | Embryo Imprecise | All anatomical structures |