5 figures and 1 table

Figures

Figure 1 with 3 supplements
mRNA levels of act-5 and act-3 in WT and mutant alleles.

qPCR analysis of act-5 (A) and act-3 (B) mRNA levels in WT and act-5(ptc), act-5(Δ1), and act-5(Δ2) mutants. act-3 mRNA levels are upregulated when act-5 mutant mRNA levels are reduced (i.e., only in the act-5(ptc) allele). WT expression levels are set at 1. Data are mean ± S.E.M.; average dCt values are shown in Figure 1—source data 1. Two-tailed Student’s t-test was used to calculate P values.

Figure 1—source data 1

Average dCt values from qPCR analysis of act-5 and act-3 mRNA levels in WT and act-5 mutants.

https://cdn.elifesciences.org/articles/50014/elife-50014-fig1-data1-v1.pdf
Figure 1—source data 2

Average dCt values from qPCR analysis of act-1, act-2 and act-4 mRNA levels in WT and act-5(ptc) mutants.

https://cdn.elifesciences.org/articles/50014/elife-50014-fig1-data2-v1.pdf
Figure 1—figure supplement 1
Organization of act-5 locus.

(A) Schematic of all known act-5 isoforms and alleles used in this study. Black boxes indicate the deletion and nonsense alleles used in this study; black arrows point to the position of the qPCR primers. The act-5(ok1397) isoform was only identified in the act-5(Δ2) allele. (B) Partial sequence of act-5 (WT, ptc and Δ1 alleles). The PTC in the act-5(ptc) allele is 264 nucleotides from the next exon-intron junction and 888 nucleotides from the stop codon. Red indicates the mutation; PTC is underlined; ‘nnn’ and ‘---’ indicate deleted nucleotides in the act-5(Δ1) allele. The dt2017 deletion leads to a PTC which is located in the last exon (153 bases from the stop codon) and is thus predicted not to trigger NMD (Kashima et al., 2010; Lindeboom et al., 2016). act-5(dt2019) = act-5(ptc); act-5(dt2017) = act-5(Δ1); act-5(ok1397) = act-5(Δ2).

Figure 1—figure supplement 2
mRNA levels of act-1 (A), act-2 (B) and act-4 (C) in WT and act-5(ptc) mutants.

WT expression levels are set at 1. Data are mean ± S.E.M.; average dCt values are shown in Figure 1—source data 2. Two-tailed Student’s t-test was used to calculate P values.

Figure 1—figure supplement 3
Pre-mRNA levels of act-3 in WT and act-5(ptc) mutants.

(A) qPCR analysis of act-3 pre-mRNA levels in WT and act-5(ptc) mutants. (B) Average dCt values from qPCR analysis of act-3 pre-mRNA levels in WT and act-5(ptc) mutants. WT expression levels are set at 1. Data are mean ± S.E.M.. Two-tailed Student’s t-test was used to calculate P values.

Figure 2 with 1 supplement
Extrachromosomal reporter expression in WT and mutant alleles.

(A) act-5p::rfp extrachromosomal reporter expression was observed in the intestine in 153 of 300 WT animals. (B) act-3p::rfp extrachromosomal reporter expression was observed in the pharynx in 182 of 400 WT animals. (C) act-3p::rfp extrachromosomal reporter expression was observed in the pharynx and intestine in 138 of 320 act-5(ptc) mutants.

Figure 2—figure supplement 1
act-5p::rfp extrachromosomal reporter expression.

act-5p::rfp extrachromosomal reporter expression was observed in the pharynx and intestine in 148 of 300 act-5(ptc) mutants.

Figure 3 with 3 supplements
mRNA levels of unc-89 and sax-3 in WT and mutant alleles.

qPCR analysis of unc-89 (C) and sax-3 (D) mRNA levels in WT and unc-89(ptc1), unc-89(ptc2), unc-89(ptc3), and unc-89(Δ) mutants. sax-3 mRNA levels in unc-89 alleles are upregulated when unc-89 mutant mRNA levels are reduced, except in the deletion allele. WT expression levels are set at 1. Data are mean ± S.E.M.; average dCt values are shown in Figure 3—source data 2. Two-tailed Student’s t-test was used to calculate P values.

Figure 3—source data 1

List of ttn-1 paralogous genes based on WormBase release WS266.

https://cdn.elifesciences.org/articles/50014/elife-50014-fig3-data1-v1.pdf
Figure 3—source data 2

Average dCt values from qPCR analysis of unc-89 and sax-3 mRNA levels in WT and unc-89 mutants.

https://cdn.elifesciences.org/articles/50014/elife-50014-fig3-data2-v1.pdf
Figure 3—source data 3

Distance, in nucleotides, from each PTC to the next exon-intron junction and to the stop codon in each unc-89 isoform in the unc-89(ptc1), unc-89(ptc2), and unc-89(ptc3) alleles.

‘-’ indicates the absence of the PTC from the isoform.

https://cdn.elifesciences.org/articles/50014/elife-50014-fig3-data3-v1.pdf
Figure 3—figure supplement 1
Organization of unc-89 locus.

Schematic of a subset of the 16 known unc-89 isoforms as well as the deletion and nonsense alleles used in this study (black boxes). Black arrows point to the position of the qPCR primers. (B) Partial sequence of the longest unc-89 isoform with the single nucleotide change causing PTCs. Red indicates the mutations; PTCs are underlined. The distance from each PTC to the next exon-intron junction and to the stop codon is shown in Figure 3—source data 3. unc-89 (gk469156) = unc-89(ptc1); unc-89 (gk506355) = unc-89(ptc2); unc-89 (gk509355) = unc-89(ptc3); unc-89(bns7000) = unc-89(Δ).

Figure 3—figure supplement 2
Pre-mRNA levels of sax-3 in WT and unc-89(ptc1), unc-89(ptc2), unc-89(ptc3) mutants.

(A) qPCR analysis of sax-3 pre-mRNA levels in WT and unc-89(ptc1), unc-89(ptc2), unc-89(ptc3) mutants. (B) Average dCt values from qPCR analysis of sax-3 pre-mRNA levels in WT and unc-89(ptc1), unc-89(ptc2), unc-89(ptc3) mutants. WT expression levels are set at 1. Data are mean ± S.E.M.. Two-tailed Student’s t-test was used to calculate P values.

Figure 3—figure supplement 3
mRNA levels in WT and mutant alleles.

qPCR analysis of unc-89 (A) and sax-3 (B) mRNA levels in WT and act-5(ptc) mutants as well as of act-5 (C) and act-3 (D) in WT and unc-89(ptc1), unc-89(ptc2) and unc-89(ptc3) mutants. WT expression levels are set at 1. Data are mean ± S.E.M.; Two-tailed Student’s t-test was used to calculate P values.

Figure 4 with 3 supplements
Factors regulating transcriptional adaptation identified in RNAi-mediated knockdown screen.
Figure 4—source data 1

List of genes and RNAi clones tested in the screen; average dCt values of qPCR analyses of act-5 and act-3 mRNA levels in WT and act-5 mutants as well as of unc-89 and sax-3 mRNA levels in WT and unc-89 mutants.

https://cdn.elifesciences.org/articles/50014/elife-50014-fig4-data1-v1.xlsx
Figure 4—figure supplement 1
qPCR analysis of act-5 (A) and act-3 (B) mRNA levels in WT and act-5(ptc) mutants as well as of unc-89 (C) and sax-3 (D) mRNA levels in WT and unc-89(ptc) mutants upon drsh-1 RNAi-mediated knockdown by two independent clones.

WT expression levels are set at 1. Data are mean ± S.E.M.; average dCt values are shown in Figure 4—source data 1. Two-tailed Student’s t-test was used to calculate P values.

Figure 4—figure supplement 2
qPCR analysis of act-5 (A) and act-3 (B) mRNA levels in WT and act-5(ptc) mutants as well as of unc-89 (C) and sax-3 (D) mRNA levels in WT and unc-89(ptc) mutants upon spk-1 RNAi-mediated knockdown by two independent clones.

WT expression levels are set at 1. Data are mean ± S.E.M.; average dCt values are shown in Figure 4—source data 1. Two-tailed Student’s t-test was used to calculate P values.

Figure 4—figure supplement 3
qPCR analysis of act-5 (A) and act-3 (B) mRNA levels in WT and act-5(ptc) mutants as well as of unc-89 (C) and sax-3 (D) mRNA levels in WT and unc-89(ptc) mutants upon nrde-3 RNAi-mediated knockdown by two independent clones.

WT expression levels are set at 1. Data are mean ± S.E.M.; average dCt values are shown in Figure 4—source data 1. Two-tailed Student’s t-test was used to calculate P values.

Figure 5 with 1 supplement
Factors regulating transcriptional adaptation analyzed in double mutants.
Figure 5—source data 1

List of genes and alleles for each gene tested in the double mutant analysis; average dCt values from qPCR analyses of act-5 and act-3 mRNA levels in WT and act-5 mutants as well as of unc-89 and sax-3 mRNA levels in WT and unc-89 mutants.

https://cdn.elifesciences.org/articles/50014/elife-50014-fig5-data1-v1.xlsx
Figure 5—figure supplement 1
Partial data from double mutant analysis.

qPCR analysis of act-5 (A) and act-3 (B) mRNA levels in nrde-3(gg66) mutants and act-5(ptc); nrde-3(gg66) double mutants as well as of unc-89 (C) and sax-3 (D) mRNA levels in nrde-3(gg66) and unc-89(ptc); nrde-3(gg66) double mutants. Single mutant nrde-3(gg66) expression levels are set at 1. Data are mean ± S.E.M.; average dCt values are shown in Figure 5—source data 1. Two-tailed Student’s t-test was used to calculate P values.

Tables

Key resources table
Reagent type (species) or resourceDesignationSource or referenceIdentifiersAdditional information
Gene (Caenorhabditis elegans)act-1CELE_T04C12.6WormBase ID:
WBGene00000063
Gene (Caenorhabditis elegans)act-2CELE_T04C12.5WBGene00000064
Gene (Caenorhabditis elegans)act-3CELE_T04C12.4WBGene00000065
Gene (Caenorhabditis elegans)act-4CELE_M03F4.2WBGene00000066
Gene (Caenorhabditis elegans)act-5CELE_T25C8.2WBGene00000067
Gene (Caenorhabditis elegans)unc-89CELE_C09D1.1WBGene00006820
Gene (Caenorhabditis elegans)sax-3CELE_ZK377.2WBGene00004729
Strain, strain background (C. elegans)N2CGC, Bristol strainwild type
Strain, strain background (C. elegans)IN2049MacQueen et al., 2005act-5(ptc); dtIs419[act-5+ rol-6(d)]
Strain, strain background (C. elegans)IN2051MacQueen et al., 2005act-5(Δ1); dtIs419[act-5+ rol-6(d)]
Strain, strain background (C. elegans)VC971CGC, Estes et al., 2011+/mT1; act-5(Δ2)/mT1 [dpy-10(e128)].
Strain, strain background (C. elegans)CB4043CGC, Hodgkin et al., 1989smg-2(e2008);him-5(e1490)
Strain, strain background (C. elegans)CB4355CGC, Pulak and Anderson, 1993smg-4(ma116);him-8(e1490)
Strain, strain background (C. elegans)TR1396CGC, Pulak and Anderson, 1993smg-6(r896)
Strain, strain background (C. elegans)YY168CGC, Pavelec et al., 2009ergo-1(gg100)
Strain, strain background (C. elegans)YY158CGC, Guang et al., 2008nrde-3(gg66)
Strain, strain background (C. elegans)YY13CGC, Pavelec et al., 2009rrf-3(mg373)
Strain, strain background (C. elegans)DYS0005This study, crossed IN2049 to N2act-5(ptc)
Strain, strain background (C. elegans)DYS0004This study, crossed IN2049 to N2+/act-5(Δ1)
Strain, strain background (C. elegans)DYS0012This study, injected in N2Ex[act-5p::RFP]
Strain, strain background (C. elegans)DYS0014This study, injected in N2Ex[act-3p::RFP]
Strain, strain background (C. elegans)DYS0015This study, crossed DYS0014 to DYS0004act-5(ptc);Ex[act-3p::RFP]
Strain, strain background (C. elegans)DYS0042This study, crossed DYS0012 to DYS0005act-5(ptc);Ex[act-5p::RFP]
Strain, strain background (C. elegans)VC40114CGC, Million Mutation Projectunc-89(ptc1)
Strain, strain background (C. elegans)VC40193CGC, Million Mutation Projectunc-89(ptc2)
Strain, strain background (C. elegans)VC40199CGC, Million Mutation Projectunc-89(ptc3)
Strain, strain background (C. elegans)DYS0028This study, crossed VC40114 to N2unc-89(ptc1)
Strain, strain background (C. elegans)DYS0030This study, crossed VC40193 to N2unc-89(ptc2)
Strain, strain background (C. elegans)DYS0031This study, crossed VC40199 to N2unc-89(ptc3)
Strain, strain background (C. elegans)DYS0037This study, induced by CRISPR/Cas9unc-89(Δ)
Strain, strain background (C. elegans)DYS0008This study, crossed DYS0005 to CB4043smg-2(e2008); act-5(ptc)
Strain, strain background (C. elegans)DYS0057This study, crossed DYS0005 to CB4355act-5(ptc); smg-4(ma116)
Strain, strain background (C. elegans)DYS0047This study, crossed DYS0028 to CB4355unc-89(ptc1); smg-4(ma116)
Strain, strain background (C. elegans)DYS0048This study, crossed DYS0030 to CB4355unc-89(ptc2); smg-4(ma116)
Strain, strain background (C. elegans)DYS0050This study, crossed DYS0031 to CB4355unc-89(ptc3); smg-4(ma116)
Strain, strain background (C. elegans)DYS0053This study, crossed DYS0028 to TR1396unc-89(ptc1); smg-6(r896)
Strain, strain background (C. elegans)DYS0055This study, crossed DYS0030 to TR1396unc-89(ptc2); smg-6(r896)
Strain, strain background (C. elegans)DYS0056This study, crossed DYS0031 to TR1396unc-89(ptc3); smg-6(r896)
Strain, strain background (C. elegans)DYS0010This study, crossed DYS0005 to YY168act-5(ptc); ergo-1(gg100)
Strain, strain background (C. elegans)DYS0054This study, crossed DYS0028 to YY168unc-89(ptc1); ergo-1(gg100)
Strain, strain
background (C. elegans)
DYS0051This study, crossed DYS0030 to YY168unc-89(ptc2); ergo-1(gg100)
Strain, strain
background (C. elegans)
DYS0052This study, crossed DYS0031 to YY168unc-89(ptc3); ergo-1(gg100)
Strain, strain background (C. elegans)DYS0045This study, crossed DYS0005 to YY158act-5(ptc); nrde-3(gg66)
Strain, strain background (C. elegans)DYS0065This study, crossed DYS0028 to YY158unc-89(ptc1); nrde-3(gg66)
Strain, strain background (C. elegans)DYS0072This study, crossed DYS0030 to YY158unc-89(ptc2); nrde-3(gg66)
Strain, strain background (C. elegans)DYS0066This study, crossed DYS0031 to YY158unc-89(ptc3); nrde-3(gg66)
Strain, strain background (C. elegans)DYS0046This study, crossed DYS0005 to YY13rrf-3(mg373); act-5(ptc)
Strain, strain background (C. elegans)DYS0070This study, crossed DYS0028 to YY13unc-89(ptc1); rrf-3(mg373)
Strain, strain background (C. elegans)DYS0062This study, crossed DYS0030 to YY13unc-89(ptc2); rrf-3(mg373)
Strain, strain background (C. elegans)DYS0063This study, crossed DYS0031 to YY13unc-89(ptc3); rrf-3(mg373)
Commercial assay or kitIn-Fusion HD CloningClontechClontech:639647
Commercial assay or kitSuperscript III reverse transcriptaseTakaraCat. No: 18080–044
Commercial assay or kitSMARTer RACE cDNA Amplification KitTakaraCat. N. 634860
Commercial assay or kitAdvantage 2 PCR kitTakaraCat. N. 639207
RNAi constructmv_C18D11.4BioSciencersp-8
RNAi constructsjj2_C18D11.4BioSciencersp-8
RNAi constructsmv_C33H5.12BioSciencersp-6
RNAi constructssjj2_C33H5.12BioSciencersp-6
RNAi constructsmv_W02B12.3BioSciencersp-1
RNAi constructssjj2_W02B12.3BioSciencersp-1
RNAi constructsmv_D2089.1BioSciencersp-7
RNAi constructssjj2_D2089.1BioSciencersp-7
RNAi constructsmv_B0464.5BioSciencespk-1
RNAi constructssjj2_B0464.5BioSciencespk-1
RNAi constructsmv_R05D11.6BioSciencepaxt-1
RNAi constructssjj2_R05D11.6BioSciencepaxt-1
RNAi constructsmv_F43E2.8BioSciencehsp-4
RNAi constructssjj2_F43E2.8BioSciencehsp-4
RNAi constructssjj2_Y39G8C.1BioSciencexrn-1
RNAi constructsmv_Y48G8AL.6BioSciencesmg-2
RNAi constructssjj2_Y48G8AL.6BioSciencesmg-2
RNAi constructssjj2_F46B6.3BioSciencesmg-4
RNAi constructsmv_Y54F10AL.2BioSciencesmg-6
RNAi constructssjj2_Y54F10AL.2BioSciencesmg-6
RNAi constructsmv_F26B1.2BioSciencehrpk-1
RNAi constructssjj2_F26B1.2BioSciencehrpk-1
RNAi constructsmv_F26E4.10BioSciencedrsh-1
RNAi constructssjj2_F26E4.10BioSciencedrsh-1
RNAi constructsmv_T22A3.5BioSciencepash-1
RNAi constructssjj2_T22A3.5BioSciencepash-1
RNAi constructssjj2_F26A3.8BioSciencerrf-1
RNAi constructsmv_ R06C7.1BioSciencewago-1
RNAi constructssjj2_ R06C7.1BioSciencewago-1
RNAi constructsmv_F58G1.1BioSciencewago-4
RNAi constructssjj2_F58G1.1BioSciencewago-4
RNAi constructssjj2_F10B5.7BioSciencerrf-3
RNAi constructsmv_M88.5BioSciencezbp-1
RNAi constructssjj2_M88.5BioSciencezbp-1
RNAi constructssjj2_K12H4.8BioSciencedcr-1
RNAi constructsmv_T20G5.11BioSciencerde-4
RNAi constructssjj2_T20G5.11BioSciencerde-4
RNAi constructsmv_F36H1.2BioSciencekdin-1
RNAi constructsmv_K12B6.1BioSciencesago-1
RNAi constructssjj2_K12B6.1BioSciencesago-1
RNAi constructsmv_K08H10.7BioSciencerde-1
RNAi constructssjj2_K08H10.7BioSciencerde-1
RNAi constructssjj2_R09A1.1BioScienceergo-1
RNAi constructsmv_R04A9.2BioSciencenrde-3 
RNAi constructssjj2_R04A9.2BioSciencenrde-3

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Vahan Serobyan
  2. Zacharias Kontarakis
  3. Mohamed A El-Brolosy
  4. Jordan M Welker
  5. Oleg Tolstenkov
  6. Amr M Saadeldein
  7. Nicholas Retzer
  8. Alexander Gottschalk
  9. Ann M Wehman
  10. Didier YR Stainier
(2020)
Transcriptional adaptation in Caenorhabditis elegans
eLife 9:e50014.
https://doi.org/10.7554/eLife.50014