Fig. 3

Knockdown of 4.1Ba causes a reduction in the number of synapses at CaPs.

(A) Labeling of 4.1Ba-GFP puncta colocalized with the presynaptic marker Synapsin 1/2 (Syn) and the postsynaptic marker panMAGUK (PM) at CaPs. Arrows indicate sites of colocalization. (B) Representative images of Syn and PM in Ctl MO, 4.1Ba MO, and 4.1Ba MO + 4.1Ba RNA (4.1Ba Res) injected embryos. (C) Knockdown of 4.1Ba reduced the number of synapses at CaPs. The phenotype was rescued with 4.1Ba and 4.1Bb mRNA (Rescue). Misexpression of 4.1Ba did not affect synapse number. (D—E) Quantification of SV2 puncta (D) and Synaptotagmin 2b puncta (E) at CaPs. Both presynaptic proteins are reduced in the absence of 4.1Ba. (F) Cell specific rescue of 4.1Ba in CaPs. Embryos were collected from a tg(mnx1:Gal4:UAS:GFP) and tg(mnx1:Gal4) cross and injected with the Ctl MO, 4.1Ba MO, or the 4.1Ba MO + UAS:4.1Ba-GFP plasmid. Embryos were analyzed based on GFP expression. Expression of 4.1Ba-GFP restored the number of synapses specifically in CaPs. N = 10 for all conditions and experiments. Significance was evaluated with a One-way ANOVA followed by Tukey’s post-hoc analysis. All experiments were repeated in triplicate. Error bars represent s.e.m. * p < 0.05; ** p< 0.01. Scale bars in A—B, D—F: 5 μm. Abbreviations: Ctl MO = control morpholino; 4.1Ba MO = 4.1Ba morpholino; 4.1Ba Rescue = 4.1Ba MO + 4.1Ba mRNA; 4.1Bb Rescue = 4.1Ba MO + 4.1Bb mRNA; 4.1Ba GFP Rescue = 4.1Ba MO + UAS:4.1Ba GFP DNA.