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ZFIN ID: ZDB-FIG-170921-57
Gallagher et al., 2017 - Pnrc2 regulates 3'UTR-mediated decay of segmentation clock-associated transcripts during zebrafish segmentation. Developmental Biology   429(1):225-239 Full text @ Dev. Biol.
ADDITIONAL FIGURES
EXPRESSION / LABELING:
Genes:
Antibodies:
Fish:
Knockdown Reagent:
Anatomical Terms:
Stage: 14-19 somites
PHENOTYPE:
Fish:
Knockdown Reagent:
Observed In:
Stage: 14-19 somites

Fig. 7 Both reporter and endogenous cyclic transcripts accumulate in Pnrc2-depleted embryos, but protein expression appears normal. Embryos carrying the her1:her1-Venusbk15transgenic clock reporter were injected with pnrc2 splice-blocking morpholino (sbMO) and processed to detect Venus transcripts (A, B) and Venus protein (D, E) at mid-segmentation stages. Representative embryos are shown in A (n=21/21); B (n=18/18), D (n=32/32), and E (n=29/29). Venus immunofluorescence panels (D, E) are at slightly higher magnification than Venus in situ panels (A, B). Detection of her1 mRNA by in situ hybridization chain reaction (HCR-ISH) (C, F) is consistent with chromogenic NBT/BCIP-based in situ detection of endogenous her1 transcript in wild-type and pnrc2 mutant embryos (Fig. 2F, H’), with substantial cytoplasmic localization revealed by DAPI counter staining in 500X magnified view (C’, F’). Because relative intensity of her1 HCR-ISH in pnrc2 mutants to wild-type embryos is high, levels have been reduced in pnrc2 mutant panels (F-F’; see Fig S6 A’’’, B’’’). Misexpression of dlc and dld mRNA is detected throughout the presomitic mesoderm (PSM), formed somites and neurons in the expected one-quarter of embryos in a pnrc2oz22intercross, n=5/28 (Χ2=0.76, p=0.4) and n=7/40 (Χ2=1.2, p=0.3), respectively (G-J). In contrast, Dlc and Dld protein expression is indistinguishable among siblings of the same pnrc2oz22 heterozygote intercross (K-N). Dlc and Dld immunolabeled embryos were genotyped prior to imaging and a subset of wild-type and mutant siblings were imaged by confocal microscopy with representative embryos shown (K-N). Total genotyped individuals per representative panel: n=5 (K), n=6 (L), n=12 (M), n=4 (N). Scale bars=50 µm (D, F), 50 nm (F’), 100 µm (K).

Gene Expression Details
Gene Antibody Fish Conditions Stage Anatomy Assay
dlc AB standard conditions 14-19 somites neuron ISH
14-19 somites segmental plate ISH
14-19 somites somite ISH
pnrc2oz22/oz22(AB) standard conditions 14-19 somites nervous system ISH
14-19 somites segmental plate ISH
14-19 somites somite ISH
dld AB standard conditions 14-19 somites neuron ISH
Ab1-dld 14-19 somites neuron IHC
14-19 somites segmental plate ISH
Ab1-dld 14-19 somites segmental plate IHC
Ab1-dld 14-19 somites somite IHC
14-19 somites somite ISH
pnrc2oz22/oz22(AB) standard conditions 14-19 somites nervous system ISH
Ab1-dld 14-19 somites neuron IHC
Ab1-dld 14-19 somites segmental plate IHC
14-19 somites segmental plate ISH
Ab1-dld 14-19 somites somite IHC
14-19 somites somite ISH
Venus bk15Tg standard conditions 14-19 somites segmental plate IHC
14-19 somites segmental plate ISH
bk15Tg + MO2-pnrc2 standard conditions 14-19 somites segmental plate ISH
14-19 somites segmental plate IHC
Antibody Labeling Details
Antibody Assay Fish Conditions Stage Anatomy
Ab2-dlc IHC AB standard conditions 14-19 somites neuron
IHC 14-19 somites segmental plate
IHC 14-19 somites somite
IHC pnrc2oz22/oz22(AB) standard conditions 14-19 somites neuron
IHC 14-19 somites segmental plate
IHC 14-19 somites somite
Phenotype Details
Fish Conditions Stage Phenotype
bk15Tg + MO2-pnrc2 standard conditions 14-19 somites segmental plate Venus expression increased amount, abnormal
pnrc2oz22/oz22(AB) standard conditions 14-19 somites nervous system dld expression increased amount, abnormal
14-19 somites nervous system dlc expression increased amount, abnormal
14-19 somites segmental plate dlc expression increased amount, abnormal
14-19 somites segmental plate dld expression increased amount, abnormal
14-19 somites somite dld expression increased amount, abnormal
14-19 somites somite dlc expression increased amount, abnormal
Acknowledgments:
ZFIN wishes to thank the journal Developmental Biology for permission to reproduce figures from this article. Please note that this material may be protected by copyright.

Reprinted from Developmental Biology, 429(1), Gallagher, T.L., Tietz, K.T., Morrow, Z.T., McCammon, J.M., Goldrich, M.L., Derr, N.L., Amacher, S.L., Pnrc2 regulates 3'UTR-mediated decay of segmentation clock-associated transcripts during zebrafish segmentation, 225-239, Copyright (2017) with permission from Elsevier. Full text @ Dev. Biol.