ZFIN ID: ZDB-FIG-140416-36
Elks et al., 2013 - Hypoxia inducible factor signaling modulates susceptibility to mycobacterial infection via a nitric oxide dependent mechanism. PLoS pathogens   9(12):e1003789 Full text @ PLoS Pathog.
ADDITIONAL FIGURES
EXPRESSION / LABELING:
Genes:
Antibody:
Fish:
Condition:
Knockdown Reagent:
Anatomical Term:
Stage: Long-pec
PHENOTYPE:
Fish:
Condition:
Observed In:
Stage: Long-pec

Fig. 5

Hif-1α mediated anti-mycobacterial effect is iNOS dependent.

(A) Fluorescent confocal micrographs of DAF-FM DA stained embryos at 2 dpf in the absence of infection. Neutrophils are identified by lyz:dsRed expression. DAF-FM DA staining between the timepoint of infection and 1 dpi produced varying levels of background and stained cells of the central nervous system (neurons and notochord) as well as having leukocyte-associated staining. The line of staining at the top of each image is DAF-FM DA staining in the notochord. Punctae of DAF-FM DA show upregulation of NO signaling Dominant active hif-1αb (DA1) embryos had more punctae than phenol red (PR) controls. nos2a morpholino (NOS2MO) reduced the punctae in both the PR and DA1 background. (B) Fluorescent confocal micrographs of DAF-FM DA stained embryos at 2 dpf in the presence of Mm infection. In phenol red (PR) controls DAF-FM DA punctae are increased. DAF-FM DA staining is not specific for iNOS (it is a pan-NOS probe), and the nos2a morpholino (NOS2MO) was not able to downregulate all of the DAF-FM DA staining after Mm infection, although punctae number were reduced. The number of punctae was also reduced in the dominant active hif-1αb (DA1) injected embryos after Mm infection. Dominant negative hif-1αb (DN1) caused no change in punctae in the presence of Mm infection compared to PR controls. (C) Fluorescent confocal micrographs of iNOS antibody staining in mpx:GFP embryos. Phenol red (PR) injected controls had very low levels of anti-iNOS antibody staining. Dominant active hif-1αb (DA1) had increased levels of anti-iNOS antibody, a stain which was mainly neutrophil specific. (D) Bacterial burden at 4 dpi after injection of DA hif-1αb (DA1) or phenol red control (PR) and treatment with the pan-NOS inhibitor L-NAME. Data shown are mean ± SEM, n = 62–89 as accumulated from 3 independent experiments. (E) Bacterial burden at 4 dpi after injection of DA hif-1αb (DA1) and treatment with the iNOS inhibitor L-NIL. Data shown are mean ± SEM, n = 60–87 as accumulated from 3 independent experiments. (F) Bacterial burden at 4 dpi after co-injection of DA hif-1αb and the nos2a morpholino, using the standard control (SCMO) morpholino as a negative control. Data shown are mean ± SEM, n = 109–116 as accumulated from 4 independent experiments.

Gene Expression Details
Gene Antibody Fish Conditions Stage Anatomy Assay
DsRed2 nz50Tg control Long-pec neutrophil IHC
nz50Tg bacterial treatment Long-pec neutrophil IHC
nz50Tg + MO3-nos2a standard conditions Long-pec neutrophil IHC
nz50Tg + MO3-nos2a bacterial treatment Long-pec neutrophil IHC
GFP i114Tg control Long-pec neutrophil IHC
Antibody Labeling Details
Antibody Assay Fish Conditions Stage Anatomy
Ab2-nos IHC i114Tg control Long-pec neutrophil
Phenotype Details
Fish Conditions Stage Phenotype
nz50Tg bacterial treatment Long-pec leukocyte nitric-oxide synthase activity increased process quality, abnormal
Acknowledgments:
ZFIN wishes to thank the journal PLoS pathogens for permission to reproduce figures from this article. Please note that this material may be protected by copyright. Full text @ PLoS Pathog.