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. 2020 Dec 11;9:e60132. doi: 10.7554/eLife.60132

Figure 1. Cell type- and developmental stage-dependent variation in nucleocytoplasmic localization of TBPH.

(A) Subcellular localization of overexpressed TBPH-Flag proteins in sensory (C4da), motor, and dopaminergic (DA) neurons, and in glial cells [Genotype: Sensory (C4da) neurons, +/+;UAS-TBPH-Flag-HA/ppk1a-Gal4, Motor neurons, +/+;UAS-TBPH-Flag-HA/D42-Gal4,UAS-mCD8-RFP, DA neurons, +/+;UAS-TBPH-Flag-HA/TH-Gal4,UAS-CD4-tdTom, Glial cells, +/+;UAS-TBPH-Flag-HA/repo-Gal4,UAS-mCD8-RFP]. DAPI staining was used to mark the nuclei. Merged immunohistochemical images of TBPH proteins (green), plasma membranes (red), and DAPI (blue) are presented at the bottom. Outer and inner dashed lines indicate borders of cell bodies and nuclei, respectively (Scale bars, 5 μm). (B) Quantification of cytoplasmic/nuclear (Cyt/Nuc) ratio of TBPH-Flag proteins in four different cell types described in A. ****p<1.0×10−4 by one-way ANOVA with Tukey’s post-hoc correction; error bars, ± SEM; n = 12 for sensory neurons, n = 24 for motor neurons, n = 7 for DA neurons, n = 28 for glial cells. (C) Subcellular localization of overexpressed TBPH-Flag proteins in C4da neurons at five different developmental time points (120 hr AEL, 0 hr APF, 18 hr APF, 10-day adult, and 40-day adult) [Genotype: +/+;UAS-TBPH-Flag-HA/ppk1a-Gal4,UAS-mCD8-RFP]. Outer and inner dashed lines indicate the borders of cell bodies and nuclei, respectively (scale bars, 5 μm). The intensity profile of fluorescent signals representing TBPH proteins across cell bodies along yellow lines are presented at the bottom. The gray dashed lines mark borders of nuclei (bottom panels). (D) Quantification of Cyt/Nuc ratio of TBPH-Flag proteins at five different developmental time points described in C. ****p<1.0×10−4, ***p=0.0007, **p=0.0042, *p=0.0369 by two-tailed t-test; error bars, ± SEM; n = 6 for 120 hr AEL, n = 9 for 0 hr APF, n = 6 for 18 hr APF, n = 11 for 10-day adult, n = 8 for 40-day adult.

Figure 1—source data 1. Numerical data plotted in Figure 1B, D and Figure 1—figure supplement 1B.

Figure 1.

Figure 1—figure supplement 1. Developmental stage-dependent changes in nucleocytoplasmic localization of endogenous TBPH.

Figure 1—figure supplement 1.

(A) Negative control for Flag immunostaining in sensory, motor, and DA neurons and glial cells at 120 hr AEL [Genotype: Sensory (C4da) neurons, +/+;ppk1a-Gal4/+, Motor neurons, +/+;D42-Gal4,UAS-mCD8-RFP/+, DA neurons, +/+;TH-Gal4,UAS-CD4-tdTom/+, Glial cells, +/+;repo-Gal4,UAS-mCD8-RFP/+]. (B) Subcellular localization of endogenous TBPH proteins in C4da neurons at 120 hr AEL and 18 hr APF. Merged images of TBPH proteins (green) and plasma membranes (red) are presented at the bottom [Genotype: +/+;UAS-mCD8-RFP/ppk1a-Gal4]. Outer and inner dashed lines indicate borders of cell bodies and nuclei, respectively (Scale bar, 5 μm). (C) Quantification of Cyt/Nuc ratio of endogenous TBPH proteins in C4da neurons at 120 hr AEL and 18 hr APF. **p=0.0021 by two-tailed t-test; error bars, ± SEM; n = 9 for 120 hr AEL, n = 3 for 18 hr APF.