Thioredoxin shapes the C. elegans sensory response to Pseudomonas produced nitric oxide
- Massachusetts General Hospital, United States
- Harvard Medical School, United States
- Center for Brain Science, Harvard University, United States
Figures
Figure 1
C. elegans avoids PA14, but not OP50 or ΔgacA.
(A) Foraging behavior of wild-type animals after 8 hr exposure to E. coli OP50 (top) and P. aeruginosa PA14 (bottom) lawns is shown. (B) Lawn occupancy of wild-type animals on OP50 and PA14 after 8 …
Figure 2
Bacterial NO is required for PA14 avoidance.
(A) The NO biosynthetic pathway in P. aeruginosa is shown. (B) Avoidance of PA14 ΔnirS lawns was dramatically reduced compared to wild-type PA14. By contrast, neither wild-type nor nirS mutants …
Figure 3
ASJ mediates both PA14 and NO avoidance behavior.
(A, B, C) The mutation tax-4(p678) abolished PA14 (A, B) and NO donor (C) avoidance. A transgene expressing tax-4 selectively in ASJ (A, C), but not in AWA/AWB/AWC or AQR/PQR/URX (B) rescued both …
Figure 4 with 1 supplement
The sensory neurons ASJ respond to NO.
(A, B) ASJ neurons respond to the onset and removal of NO with increased GCaMP6 signal (A) but do not respond when switched between control buffer (B). (C) Blocking synaptic transmission (in unc-13 …
Figure 4—figure supplement 1
NO response amplitudes for data in Figure 4.
(A, B) NO evoked ON (A) and OFF (B) response amplitudes (scatter plots and mean ± SEM) are shown for recordings in Figure 4. ON and OFF response amplitudes are defined in the Methods. For …
Figure 5 with 1 supplement
The rGCs DAF-11 and GCY-27 are required for ASJ responses to NO.
(A) The daf-11(m47) mutation abolished PA14 (left) and NO donor (right) avoidance, both of which were reinstated by a transgene that expresses daf-11 in the ASJ neurons. **p<0.01, and ****p<0.0001 …
Figure 5—figure supplement 1
NO response amplitudes for data in Figure 5.
(A, B) NO evoked ON (A) and OFF (B) response amplitudes (scatter plots and mean ± SEM) are shown for recordings in Figure 5. ON and OFF response amplitudes are defined in the Methods. For …
Figure 6 with 1 supplement
C. elegans senses NO as an external cue.
(A, B) osm-12(n1606), bbs-9(gk471) and daf-25(m362) mutants were defective for PA14 (A) and NO donor (B) avoidance. Expressing a wild-type daf-25 cDNA in ASJ partially rescued the daf-25 mutant PA14 …
Figure 6—figure supplement 1
NO response amplitudes for data in Figure 6.
(A, B) NO evoked ON (A) and OFF (B) response amplitudes (scatter plots and mean ± SEM) are shown for recordings in Figure 6. ON and OFF response amplitudes are defined in the Methods. For …
Figure 7 with 1 supplement
TRX-1/Thioredoxin regulates the ASJ response to NO.
(A) Domain organization of C. elegans thioredoxin TRX-1. Cysteines forming the de-nitrosylation active site (Cys38XXCys41) in TRX-1 are highlighted in red. Cys72, which is involved in …
Figure 7—figure supplement 1
NO response amplitudes for data in Figure 7.
(A, B) NO evoked ON (A) and OFF (B) response amplitudes (scatter plots and mean ± SEM) are shown for recordings in Figure 7. ON and OFF response amplitudes are defined in the Methods. For …
Figure 8
A model for TRX-1/Thioredoxin function in NO sensation.
Our data suggest that two enzymatic functions of TRX-1 endow ASJ neurons with a bi-phasic response to environmental NO. The trans-nitrosylation activity is proposed to be active during NO exposures. …
Tables
Key resources table
| Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| Strain (C. elegans) | N2 Bristol | http://www.wormbase.org | N2 | N/A |
| Strain (C. elegans) | tax-4(p678) | http://www.wormbase.org | PR678 | N/A |
| Strain (C. elegans) | tax-2(p671) | http://www.wormbase.org | PR671 | N/A |
| Strain (C. elegans) | tax-4(p678);nuEx1965[podr-3::TAX-4] | This paper | KP9653 | 10ng/ul KP#3587 injected |
| Strain (C. elegans) | tax-4(p678);nuEx1966[pgcy-36::TAX-4] | This paper | KP9654 | 10ng/ul KP#3588 injected |
| Strain (C. elegans) | tax-4(p678);nuEx1967[ptrx-1::TAX-4] | This paper | KP9655 | 10ng/ul KP#3589 injected |
| Strain (C. elegans) | nuIs556;nuEx1968[ptrx-1::EGL-1] | This paper | KP9656 | 10ng/ul KP#3590 injected |
| Strain (C. elegans) | nuIs556[ptrx-1::GCaMP6.0s] | This paper | KP9672 | KP#3311 (25 ng/ul), UV integrated |
| Strain (C. elegans) | unc-13(s69);nuIs556 | This paper | KP9673 | N/A |
| Strain (C. elegans) | tax-4(p678);nuIs556 | This paper | KP9657 | N/A |
| Strain (C. elegans) | tax-4(p678);nuIs556;nuSi211[ptrx-1::TAX-4] | This paper | KP9658 | nuSi211 single copy miniMos |
| Strain (C. elegans) | daf-11(m47) | http://www.wormbase.org | DR47 | N/A |
| Strain (C. elegans) | daf-11(ks67) | http://www.wormbase.org | FK183 | N/A |
| Strain (C. elegans) | daf-11(m47);nuSi196 [pssu-1::DAF-11] | This paper | KP9659 | nuSi196 single copy miniMos |
| Strain (C. elegans) | daf-11(m47);nuIs556 | This paper | KP9660 | N/A |
| Strain (C. elegans) | daf-11(m47);nuIs556;nuSi196 | This paper | KP9661 | N/A |
| Strain (C. elegans) | gcy-27(ok3653) | http://www.wormbase.org | RB2622 | N/A |
| Strain (C. elegans) | gcy-27(ok3653);nuIs556 | This paper | KP9662 | N/A |
| Strain (C. elegans) | daf-11(nu629 ΔHNOBA);nuIs556 | This paper | KP9663 | nu629 isolated by CRISPR; Sequence in Methods |
| Strain (C. elegans) | osm-12(n1606) | http://www.wormbase.org | MT3645 | N/A |
| Strain (C. elegans) | bbs-9(gk471) | http://www.wormbase.org | VC1062 | N/A |
| Strain (C. elegans) | daf-25(m362) | http://www.wormbase.org | DR2386 | N/A |
| Strain (C. elegans) | daf-25(m362);nuEx1969[pssu-1::DAF-25] | This paper | KP9664 | 10 ng/ul KP#3593 injected |
| Strain (C. elegans) | daf-25(m362);nuIs556 | This paper | KP9665 | N/A |
| Strain (C. elegans) | trx-1(jh127) | http://www.wormbase.org | KJ412 | N/A |
| Strain (C. elegans) | trx-1(jh127);nuIs556 | This paper | KP9666 | N/A |
| Strain (C. elegans) | trx-1(jh127);nuSi197 [ptrx-1::TRX-1] | This paper | KP9674 | nuSi197 single copy miniMos |
| Strain (C. elegans) | trx-1(jh127);nuIs556;nuSi197 | This paper | KP9667 | N/A |
| Strain (C. elegans) | trx-1(nu517 C38S) | This paper | KP9675 | nu517 isolated by CRISPR; Sequence in Methods |
| Strain (C. elegans) | trx-1(nu517 C38S);nuIs556 | This paper | KP9668 | N/A |
| Strain (C. elegans) | trx-1(jh127);nuSi198 [ptrx-1::TRX-1(C72S)] | This paper | KP9676 | nuSi198 single copy miniMos |
| Strain (C. elegans) | trx-1(jh127);nuIs556;nuSi198 | This paper | KP9669 | N/A |
| Strain (C. elegans) | gsnor-1(nu518) | This paper | KP9670 | nu518 isolated by CRISPR; Sequence in Methods |
| Strain (C. elegans) | gsnor-1(nu518);nuIs556 | This paper | KP9671 | N/A |
| Strain (E. coli) | OP50 | (Brenner, 1974) | OP50 | N/A |
| Strain (P. aeruginosa) | PA14 | Fred Ausubel lab | PA14 | N/A |
| Strain (P. aeruginosa) | PA14 gacA | Fred Ausubel lab | PA14 gacA | N/A |
| Strain (P. aeruginosa) | PA14 nirS | Constantine Haidaris lab | PA14 nirS | (Van Alst et al., 2009) |
| Recombinant DNA reagent | pmyo-2::NLS-mCherry | Kaplan lab | KP#1480 | N/A |
| Recombinant DNA reagent | punc-122::mCherry | This paper | KP#2186 | 796bp unc-122 promoter |
| Recombinant DNA reagent | podr-3::TAX-4 | This paper | KP#3587 | 3kb odr-3 promoter; tax-4 cDNA from C. Bargmann |
| Recombinant DNA reagent | pgcy-36::TAX-4 | This paper | KP#3588 | gcy-36 promoter from C. Bargmann |
| Recombinant DNA reagent | ptrx-1::TAX-4 | This paper | KP#3589 | 1028bp trx-1 promoter |
| Recombinant DNA reagent | ptrx-1::EGL-1 | This paper | KP#3590 | 321bp egl-1 cDNA |
| Recombinant DNA reagent | ptrx-1::GCaMP6.0s | This paper | KP#3311 | GCaMP6.0s from Jihong Bai |
| Recombinant DNA reagent | ptrx-1::TAX-4 miniMos | This paper | KP#3591 | N/A |
| Recombinant DNA reagent | pssu-1::DAF-11 miniMos | This paper | KP#3592 | 543bp ssu-1 promoter and 3234bp daf-11 cDNA |
| Recombinant DNA reagent | pssu-1::DAF-25 | This paper | KP#3593 | 543 bp ssu-1 promoter; 1167bp daf-25 cDNA |
| Recombinant DNA reagent | ptrx-1::TRX-1 miniMos | This paper | KP#3594 | 1028bp trx-1 promoter; 723bp trx-1b genomic fragment |
| Recombinant DNA reagent | ptrx-1::TRX-1(C72S) miniMos | This paper | KP#3595 | created by Site-Directed Mutagenesis |
| Commercial assay or kit | QIAprep Spin Miniprep Kit | QIAGEN | 27106 | N/A |
| Commercial assay or kit | QIAquick Gel Extraction Kit | QIAGEN | 28706 | N/A |
| Commercial assay or kit | HiSpreed Plasmid Midi Kit | QIAGEN | 12643 | N/A |
| Commercial assay or kit | Q5 Site-Directed Mutagenesis Kit | NEW ENGLAND BIOLABS INC | E0552S | N/A |
| Chemical compound, drug | DPTA NONOate | Cayman Chemical | 82110 | N/A |
| Chemical compound, drug | MAHMA NONOate | Sigma | M1555 | N/A |
| Chemical compound, drug | BDM (2,3-Butanedione monoxide) | Sigma | B0753 | N/A |
| Software, algorithm | Metamorph 7.1 | Molecular Devices | N/A | N/A |
| Software, algorithm | Fiji | https://fiji.sc/ | N/A | N/A |
| Software, algorithm | Prism 6 | https://www.graphpad.com/scientific-software/prism/ | N/A | N/A |
| Software, algorithm | IBM SPSS statistics 25 | https://www.ibm.com/products/spss-statistics | N/A | N/A |
| Other | olfactory microfluidic chips | (Chronis et al., 2007) | N/A | N/A |
Additional files
-
Transparent reporting form
- https://doi.org/10.7554/eLife.36833.015
