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. 2022 Sep 6;119(37):e2206817119. doi: 10.1073/pnas.2206817119

Fig. 2.

Fig. 2.

Molecular IL2 subtype diversification is controlled by unc-39. (A) Expression pattern of the unc-39(syb4537[unc-39::gfp]) reporter allele across embryonic and larval stages. (B) Lateral IL2 marker expression is lost in unc-39R203Q mutant animals (either canonical e257 allele or CRISPR/Cas9 genome engineered ot1172 or ot1173 alleles with identical nucleotide change). Markers are nlp-69(syb4512[nlp-69::SL2::gfp::H2B]), ins-1(syb5452[ins-1::SL2::gfp::H2B]), flp-14(syb3323[flp-14::SL2::gfp::H2B]), degl-1 (otIs825), and egas-4 (otIs833). NeuroPAL images for cell identification and quantification are in SI Appendix, Fig. S4 B and C. (C) Dorsoventral IL2 markers dmsr-2(syb4514[dmsr-2::SL2::gfp::H2B]), flp-32(syb4374[flp-32::SL2::gfp::H2B]), degl-2(syb5229[degl-2::SL2::gfp::H2B]), and egas-1(otIs846) are gained in all IL2 neurons of unc-39R203Q mutant animals (either canonical e257 allele or CRISPR/Cas9 genome engineered ot1173 allele with identical nucleotide change). We counted gain of expression as an all or none phenotype, but the converted lateral IL2 sometimes can be dimmer (in the Bottom Right subpanel, egas-1, the asterisk marks such a case). NeuroPAL images for cell ID and quantification are in SI Appendix, Fig. S4 B and C. Note that the IL2 are often displaced (Fig. 3A) in unc-39R203Q mutant animals. In the unc-39(ok2137) null mutant animals, the front part of the anterior ganglion moves past the anterior bulb of the pharynx (SI Appendix, Fig. S4D). (D) Ectopic expression of unc-39 in the IL2, URA, and URB neurons in an unc-39R203Q mutant background (e257, ot1172, or ot1173 allele) results in rescue of mutant phenotype in the lateral IL2 neurons and conversion of dorsal/lateral IL2 neurons. In each subpanel, the unc-39R203Q control is shown on the Left and two independent extrachromosomal array lines on the Right. Two lateral IL2 markers [nlp-69(syb4512) rescue lines otEx7870 and otEx7871, egas-4/otIs833 otEx7874, and otEx7875] that are lost in the hypomorph are now partially rescued and even expanded in more than just the two lateral IL2s or even the six IL2 (nlp-69, 10 cells in the Middle panel vs. 6 on the Right). (E) Three dorsoventral IL2 markers [degl-2(syb5229) rescue lines otEx7868 otEx7869, egas-1/otIs846 rescue lines otEx7872 and otEx7873, and flp-32(syb4374) rescue lines otEx7917 and otEx7918] that are found in all the IL2 in the hypomorphic mutant animals are repressed in all subtypes upon unc-39 misexpression. unc-39 is able to suppress an unc-86–dependent marker like flp-32(syb4374) in all IL2s but also the relatively similar URAs, which all share unc-86 as identity regulators. In the flp-32(ot1073) image a ventral view is provided. Quantification is in SI Appendix, Fig. S4 E and F.