Figure 4. SST⁺ neuron-specific ER stress induced by preproSST.

a. AAV-vector design: A human synapsin promoter (hSyn) drives the transgene expression when the host-derived Cre recombinase excises the stop cassette (lox_SV40-polyA_lox). Each transgene is fused in frame with eGFP via T2A self-cleavage signal. Transgene-expressing cells are labeled by eGFP, which is released from the expressed transgene product. Full-length preproSST and two truncation mutants (processing incompetent and mature SST) used are shown.

b. Fractionation of cytosolic and membrane fractions for proteins expressed from AAV vectors in HEK293T heterologous expression system. Proteins were detected by Western blot or dot blot using the indicated antibodies.

c. GFP-tagged full-length preproSST transgene or GFP alone were introduced via AAV into SST⁺ neurons in PFC of male mice (N=4/group, 3–4 months old). The p-eIF2α (Ser-51) levels (normalized by total eIF2α levels) in GFP⁺ cells (pointed by arrow) were evaluated in IF assays. Scale bar: 50 μm.

d. AAV encoding each transgene was delivered into a specific cell type in PFC (“cell type” > “transgene”) and the p-eIF2α levels in each infected cell were plotted (mean±SEM in red). Kruskal-Wallis test with Dunn’s multiple comparisons; *P<0.05.

e. Overexpressing preproSST in SST⁺ neurons is sufficient to induce elevated behavioral emotionality. Z-scoring of emotionality assays in mice (Sst^IRES-Cre/+) injected into PFC with the indicated AAV (N=7/group/sex). Results of independent tests are depicted in Fig. S4. Kruskal-Wallis test with Dunn’s multiple comparisons; *P<0.05, #P=0.086.