antibody anti-zebrafish p53 (rabbit polyclonal)  (AnaSpec)

Journal: eLife

Article Title: Opposing p53 and mTOR/AKT promote an in vivo switch from apoptosis to senescence upon telomere shortening in zebrafish

doi: 10.7554/eLife.54935

Figure Lengend Snippet: ( A-B ) Western blot analysis of DNA damage and senescence-associated proteins in gut and testis of 3 month ( A ) or 9-month-old ( B ) of WT and tert-/- siblings (N >= 5 fish). Representative western blot (left panel) and corresponding quantification (right panel) showing induction of DNA Damage Response (H2A.X-P and p53) in 3-month-old and senescence (p15/16) in 9-month-old tert-/- zebrafish. ( C ) RT-qPCR analysis of senescence associated genes p15/16 and p21. RT-qPCR graphs are representing mean ± SEM mRNA fold increase after normalisation by rpl13a gene expression levels (* p-value<0.05; ** p-value<0.01, using the Mann-Whitney test). Figure 2—source data 1. Western Blot quantifications, as plotted in . Figure 2—source data 2. Real-time qPCR data of p15/16 and p21, as plotted in .

Article Snippet: Antibody , anti-zebrafish p53 (rabbit polyclonal) , Anaspec , #55342; RRID: AB_2287635 , WB (1:1000).

Techniques: Western Blot, Quantitative RT-PCR, Gene Expression, MANN-WHITNEY

Journal: eLife

Article Title: Opposing p53 and mTOR/AKT promote an in vivo switch from apoptosis to senescence upon telomere shortening in zebrafish

doi: 10.7554/eLife.54935

Figure Lengend Snippet: ( A and E ) Representative haematoxylin and eosin-stained sections of gut ( A ) (scale bar: 40 µm) and testis ( E ) (scale bar: 25 µm) from 6-month-old WT, tert-/-, tp53-/ - and tert-/- tp53-/ - siblings (N = 3 fish each);. Mutation of tp53 in tert-/- fish rescues short-telomere induced tissue defects. ( B and F ) Representative western blot analysis of AKT-p and SOD2 in gut ( B ) and testis ( F ) (N = 2 fish each). Mutation of tp53 in tert-/- fish prevents phosphorylation of AKT and downstream downregulation of SOD2 leading to a rescue of increased ROS levels (C and G; N = 3 fish per genotype). ( D and H ) Representative images of SA-β-GAL staining of gut (scale bar: 40 µm) ( D ) and testis (scale bar: 25 µm) ( H ) from 6 month-old WT, tert-/-, p53-/ - and tert-/- p53-/ - siblings (N = 3 fish). Data are represented as mean ± SEM (** p-value<0.01, using t-test). Figure 5—source data 1. ROS levels measurements, as plotted in .

Article Snippet: Antibody , anti-zebrafish p53 (rabbit polyclonal) , Anaspec , #55342; RRID: AB_2287635 , WB (1:1000).

Techniques: Staining, Mutagenesis, Western Blot, Phospho-proteomics

Journal: eLife

Article Title: Opposing p53 and mTOR/AKT promote an in vivo switch from apoptosis to senescence upon telomere shortening in zebrafish

doi: 10.7554/eLife.54935

Figure Lengend Snippet: Early telomere shortening triggers p53-dependent apoptosis and inhibition of cell proliferation. At early age, apoptosis is the predominant cell fate and it mostly counterbalanced by compensatory proliferation of neighboring cells. However, inhibition of cell proliferation results in a progressive loss of tissue cellularity, eventually leading to tissue damage. As age progresses, loss of tissue homeostasis triggers the pro-proliferative mTOR/AKT pathway. Akt phosphorylates FoxO, inducing its translocation from the nucleus to the cytoplasm. Loss of FoxO transcriptional activity reduces mitochondrial SOD2 expression generating mitochondria oxidative stress through increased ROS levels. Mitochondrial dysfunction eventually triggers p15/16 expression and senescence becomes the predominant cell fate.

Article Snippet: Antibody , anti-zebrafish p53 (rabbit polyclonal) , Anaspec , #55342; RRID: AB_2287635 , WB (1:1000).

Techniques: Inhibition, Translocation Assay, Activity Assay, Expressing

Journal: eLife

Article Title: Opposing p53 and mTOR/AKT promote an in vivo switch from apoptosis to senescence upon telomere shortening in zebrafish

doi: 10.7554/eLife.54935

Figure Lengend Snippet:

Article Snippet: Antibody , anti-zebrafish p53 (rabbit polyclonal) , Anaspec , #55342; RRID: AB_2287635 , WB (1:1000).

Techniques: Sequencing, Control, In Situ, Cell Viability Assay, Staining

Journal: Journal of Applied Genetics

Article Title: Splice-acceptor site mutation in p53 gene of hu888 zebrafish line

doi: 10.1007/s13353-014-0239-4

Figure Lengend Snippet: Scheme of the human ( a ) and the zebrafish ( b ) p53 gene structure: an N-terminal transactivation domain (TAD), a proline-rich domain (PRD), a large DNA binding domain (DBD), a tetramerization domain (4D), a C-terminal regulatory domain (CTD), a nuclear export signal (NES), a nuclear localization signal (NLS), a bipartite nuclear localization signal (BNLS). Amino acid numbers for functional domains are indicated. ( c ) The p53 hu888 zebrafish line has a G to T point mutation at the splice-acceptor site in the intron 8 of p53 gene. Numbers of constitutive exons are indicated

Article Snippet: The membrane was then incubated overnight at 4 °C with rabbit anti-zebrafish p53 antibody (#55342, Anaspec, San Jose; CA), and subsequently incubated with HRP-conjugated goat anti-rabbit IgG antibody (Cell Signaling; USA) for 1 h at room temperature.

Techniques: Binding Assay, Functional Assay, Mutagenesis

Journal: Journal of Applied Genetics

Article Title: Splice-acceptor site mutation in p53 gene of hu888 zebrafish line

doi: 10.1007/s13353-014-0239-4

Figure Lengend Snippet: Expression of p53 splicing variants in hu888 zebrafish line. ( a ) Electropherograms of DNA sequences spanning regions of the G/T mutation (K). ( b ) Agarose gel electrophoresis of RT-PCR amplification of mRNA from liver of two wild type zebrafish line and p53 hu888 zebrafish mutant (heterozygous). Primers Drp53- 7266F and Dr-p53-7502R were used. Lane “T” p53 : amplicon of the mRNA expressed from the liver of wild type Tübingen zebrafish strain, lane p53 : amplicons of the mRNA expressed from the liver of wild type Sanger zebrafish strain, lane p53 hu888/+ : amplicons of the mRNA expressed from the liver of p53hu888 zebrafish mutant (heterozygous). Normally spliced mRNA yielded a 164-bp amplicon, whereas the alternative splicing resulted in a 152-bp amplicon. ( c ) Expression intensities of p53 splice isoforms in liver of two wild type zebrafish line and p53 hu888/+ zebrafish (blue peaks); size standard GS 500LIZ (orange peaks). ( d ) Splicing mechanism using a normal splice site (above). The resulting product corresponds to the bands of 164 bp. Alternative splicing mechanism using a cryptic splice sites (below), owing to mutations at splice-acceptor site G>T (marked in red). In case of this abnormal splicing pathway 12 nt are deleted from downstream of exon 9. The resulting product corresponds to the bands of 152 bp

Article Snippet: The membrane was then incubated overnight at 4 °C with rabbit anti-zebrafish p53 antibody (#55342, Anaspec, San Jose; CA), and subsequently incubated with HRP-conjugated goat anti-rabbit IgG antibody (Cell Signaling; USA) for 1 h at room temperature.

Techniques: Expressing, Mutagenesis, Agarose Gel Electrophoresis, Reverse Transcription Polymerase Chain Reaction, Amplification, Alternative Splicing

Journal: Journal of Applied Genetics

Article Title: Splice-acceptor site mutation in p53 gene of hu888 zebrafish line

doi: 10.1007/s13353-014-0239-4

Figure Lengend Snippet: Clustal X alignments of two p53 splicing products obtained in this study. The nucleotide sequences of p53 amplicons from both wild type ( p53 +/+ ) and p53 hu888 zebrafish. The RT-PCR primers are indicated by arrows. The amino acid sequence of zebrafish is shown (NCBI: NP_001258749.1)

Article Snippet: The membrane was then incubated overnight at 4 °C with rabbit anti-zebrafish p53 antibody (#55342, Anaspec, San Jose; CA), and subsequently incubated with HRP-conjugated goat anti-rabbit IgG antibody (Cell Signaling; USA) for 1 h at room temperature.

Techniques: Reverse Transcription Polymerase Chain Reaction, Sequencing

Journal: Journal of Applied Genetics

Article Title: Splice-acceptor site mutation in p53 gene of hu888 zebrafish line

doi: 10.1007/s13353-014-0239-4

Figure Lengend Snippet: Representative Western blot results from Western blot analysis of p53 expression in two wild type zebrafish lines and p53 hu888/+ mutant. Actin was used as the loading control. “T” p53 : wild type Tübingen zebrafish strain; p53 : wild type Sanger zebrafish strain; p53 hu888/+ : p53 hu888 zebrafish mutant (heterozygous)

Article Snippet: The membrane was then incubated overnight at 4 °C with rabbit anti-zebrafish p53 antibody (#55342, Anaspec, San Jose; CA), and subsequently incubated with HRP-conjugated goat anti-rabbit IgG antibody (Cell Signaling; USA) for 1 h at room temperature.

Techniques: Western Blot, Expressing, Mutagenesis, Control