(A) LIN-29 neuronal expression is regulated by the sex-determination pathway. Mutant hermaphrodites for the downstream effector of the sex-determination pathway tra-1 showed lin-29a/b::gfp (xe61) expression in hermaphrodite neurons in L4 and adult stages. Masculinization of the hermaphrodite nervous system by a pan-neuronal promoter driving fem-3 (otEx7321), that represses tra-1 function, induced lin-29a/b::gfp (xe61) expression in hermaphrodite neurons in L4 and onwards. (B) lin-29a/b::gfp neuronal expression is lost in neuron-specific transcription factor mutants. The POU homeobox transcription factor unc-86 is required for lin-29a/b::gfp expression in AIM interneurons. The LIM-HD transcription factor ttx-3 is required for lin-29a/b::gfp expression in ASK. The COE/EBF transcription factor unc-3 is required for lin-29a/b::gfp expression in AVA, SAB, DA, DB, VA, VB, PDA, PDB and PVC neurons. Cholinergic neurons are labeled with the cho-1/CHT mCherry fosmid reporter. (C) mab-3::gfp expression in the SMD neurons in control hermaphrodites, males and mutant animals. mab-3::gfp (ot931) expression was determined during larval development. GFP was first observed in the SMD neurons at the L4 stage and onwards, only in males. mab-3::gfp expression was lost from SMD neurons in a let-7(n2853ts) mutant male. L1 let-7(n2853ts) animals were shifted to the restricted temperature (25°C) and imaged as adults, after 48hs. mab-3::gfp expression was lost from SMD neurons in an unc-42(e419) mutant male. Scale bar: 5 µm. (D) dmd-3::gfp expression in the PHC neuron in control hermaphrodites, males and mutant animals. GFP was first observed in the PHC neurons at the L4 stage and onwards, only in males. dmd-3::gfp expression was decreased from PHC neurons in a let-7(n2853) mutant male. L1 let-7(n2853) animals were shifted to the restricted temperature (25°C) and imaged as adults, after 48hs. (E) dmd-3::gfp expression in PHC was lost in ceh-14 null mutant males and in let-7(n2853ts) mutant males. L1 let-7(n2853ts) animals were shifted to the restricted temperature (25°C) and imaged as adults, after 48hs. (F) Masculinization of the hermaphrodite nervous system by a pan-neuronal promoter driving fem-3 (otEx7322 and otEx7323, respectively), that promotes TRA-1 degradation, induced mab-3::gfp (ot931) expression in hermaphrodite SMD neurons and dmd-3::gfp (ot932) expression in hermaphrodite PHC neurons in L4 and onwards.