ZFIN ID: ZDB-FIG-180516-7
Zhang et al., 2017 - Rb1 promotes T cell maturation from premature apoptosis by inhibiting E2F1. Development (Cambridge, England)   145(1) Full text @ Development
ADDITIONAL FIGURES
EXPRESSION / LABELING:
Genes:
Fish:
Knockdown Reagent:
Anatomical Terms:
Stage: Day 5
PHENOTYPE:
Fish:
Knockdown Reagent:
Observed In:
Stage: Day 5

Fig. 5

The excessive apoptosis of early T lymphocytes in the thymus of rb1smu8/smu8 mutants is e2f1 mediated. (A) Relative expression of e2f1 in excised thymus and tail tissue from 5 dpf embryos by qPCR analysis (mean±s.e.m.; ***P<0.001; ns, not significant; n=30). (B) The zebrafish e2f1 gene structure. Exons are indicated by grey boxes. Location and sequence of the splice morpholino (MOsp) target site for the e2f1 gene. FP/RP, e2f1 qPCR forward/reverse primers. Relative expression of e2f1 in controls and e2f1 morphants at 5 dpf by qPCR analysis (mean±s.e.m.; **P<0.01; n=30). (C) Confocal images of rag2:DsRed cells in the thymus of wild-type embryos injected with double distilled H2O or e2f1 MOsp at the one-cell stage with rag2:DsRed+ cell number measured at 5 dpf. Scale bars: 50 μm. Quantification of rag2-DsRed+ cells (right panel) (mean±s.e.m.; ***P<0.001; n=13). (D) Confocal images of T cells (red) exhibiting cell apoptosis (overlap of TUNEL staining) in siblings and rb1smu8/smu8 mutant embryos injected with double distilled H2O or e2f1 MOsp at 5 dpf. Scale bars: 50 μm. (E) Percentages of T cells (red) exhibiting cell apoptosis (overlap of TUNEL staining) in siblings and rb1smu8/smu8 mutant embryos injected with double distilled H2O or e2f1 MOsp at 5 dpf (mean±s.e.m.; ***P<0.001; n=20). (F) Quantification of rag2-DsRed+ cell shown in D (mean±s.e.m.; ***P<0.001; n=10). (G) qPCR analysis of e2f1, rag2, tcra, tcrb, cd3 and casp3 expression in mutants injected with double distilled H2O or e2f1 MOsp at 5 dpf (mean±s.e.m.; ***P<0.001, **P<0.01 *P<0.05; n=30).

Gene Expression Details
Gene Antibody Fish Conditions Stage Anatomy Assay
casp3a AB + MO1-e2f1 control Day 5 whole organism RTPCR
rb1sm11/sm11 control Day 5 whole organism RTPCR
rb1sm11/sm11 + MO1-e2f1 control Day 5 whole organism RTPCR
cd247 AB + MO1-e2f1 control Day 5 whole organism RTPCR
rb1sm11/sm11 control Day 5 whole organism RTPCR
rb1sm11/sm11 + MO1-e2f1 control Day 5 whole organism RTPCR
DsRed rb1sm11/sm11 ; zf411Tg control Day 5 thymus T cell IHC
rb1sm11/sm11 ; zf411Tg + MO1-e2f1 control Day 5 thymus T cell IHC
zf411Tg control Day 5 thymus T cell IHC
Day 5 thymus T cell IFL
zf411Tg + MO1-e2f1 control Day 5 thymus T cell IFL
Day 5 thymus T cell IHC
e2f1 AB + MO1-e2f1 control Day 5 whole organism RTPCR
rb1sm11/sm11 control Day 5 thymus anatomical region RTPCR
Day 5 whole organism RTPCR
rb1sm11/sm11 + MO1-e2f1 control Day 5 whole organism RTPCR
linc-tcrb AB + MO1-e2f1 control Day 5 whole organism RTPCR
rb1sm11/sm11 control Day 5 whole organism RTPCR
rb1sm11/sm11 + MO1-e2f1 control Day 5 whole organism RTPCR
rag2 AB + MO1-e2f1 control Day 5 whole organism RTPCR
rb1sm11/sm11 control Day 5 whole organism RTPCR
rb1sm11/sm11 + MO1-e2f1 control Day 5 whole organism RTPCR
trac AB + MO1-e2f1 control Day 5 whole organism RTPCR
rb1sm11/sm11 control Day 5 whole organism RTPCR
rb1sm11/sm11 + MO1-e2f1 control Day 5 whole organism RTPCR
Antibody Labeling Details No data available
Phenotype Details
Fish Conditions Stage Phenotype
AB + MO1-e2f1 control Day 5 whole organism casp3a expression decreased amount, abnormal
Day 5 whole organism cd247 expression increased amount, abnormal
Day 5 whole organism e2f1 expression decreased amount, abnormal
Day 5 whole organism rag2 expression increased amount, abnormal
Day 5 whole organism trac expression increased amount, abnormal
Day 5 whole organism linc-tcrb expression increased amount, abnormal
rb1sm11/sm11 control Day 5 thymus anatomical region e2f1 expression increased amount, abnormal
Day 5 whole organism casp3a expression increased amount, abnormal
Day 5 whole organism cd247 expression decreased amount, abnormal
Day 5 whole organism e2f1 expression increased amount, abnormal
Day 5 whole organism rag2 expression decreased amount, abnormal
Day 5 whole organism trac expression decreased amount, abnormal
Day 5 whole organism linc-tcrb expression decreased amount, abnormal
rb1sm11/sm11 + MO1-e2f1 control Day 5 whole organism casp3a expression amount, ameliorated
Day 5 whole organism cd247 expression amount, ameliorated
Day 5 whole organism e2f1 expression amount, ameliorated
Day 5 whole organism rag2 expression amount, ameliorated
Day 5 whole organism trac expression amount, ameliorated
Day 5 whole organism linc-tcrb expression amount, ameliorated
rb1sm11/sm11 ; zf411Tg control Day 5 T cell apoptotic process increased occurrence, abnormal
Day 5 thymus T cell decreased amount, abnormal
rb1sm11/sm11 ; zf411Tg + MO1-e2f1 control Day 5 T cell apoptotic process occurrence, ameliorated
Day 5 thymus T cell decreased amount, ameliorated
zf411Tg + MO1-e2f1 control Day 5 thymus T cell increased amount, abnormal
Acknowledgments:
ZFIN wishes to thank the journal Development (Cambridge, England) for permission to reproduce figures from this article. Please note that this material may be protected by copyright. Full text @ Development