Supplementary Figure 1: Vav2, but not Vav3, is preferentially expressed in the VTA of adult mice and is required for the maintenance of dopamine homeostasis.

a, Immunoblotting reveals the Vav2 expression levels in the ventral tagmental area (VTA) is significantly higher than in the substantia nigra (SN) of adult brain. b, Quantitative data of a. (n = 4); t₍₆₎= 6.317, *P = 0.0007. c. Western analysis of brain tissue lysates from adult mouse brain shows that Vav3 is ubiquitously expressed in major brain regions. The experiments were repeated at least twice. d-f, Measurement of DA, 5-HT and their metabolites levels were performed in the olfactory bulb (OB) of Vav2 mutants (d), nucleus accumbens (NAc) (e) and dorsal striatum (dSTR) (f) of Vav3 mutant and wild-type mice. d, n = 3 per group; unpaired two-tailed t-test, t₍₄₎ = 0.8303, P = 0.4530 for DA. e, f, n = 10 for WT, n = 8 for Vav3 mutants. unpaired two-tailed t-test. e, t₍₁₆₎ = 1.196, P = 0.2490 for DA. f, t₍₁₆₎ = 0.02254, P = 0.9823 for DA. g-h, Tyrosine hydroxylase immunosignal density was measured in the NAc (g) and dSTR (h) of Vav2 mutant and WT mice. (g, h, n = 4). unpaired two-tailed t-test. g, t₍₄₆₎ = 1.913, P = 0.0620. h, t(60) = 1.108, P = 0.2721. i, Western blotting shows that AAV-mediated overexpression of Vav2 following injection in the VTA or SN, validating the transfection efficiency in vivo. j, [³H]DA uptake assays were performed in the NAc or dSTR synapotosome preparation from Vav3-null and wild-type. (n = 3 mice per group). unpaired two-tailed t-test. t₍₄₎ = 0.8844, P = 0.4264 for NAc, t₍₄₎ = 0.2033, P = 0.8488 for dSTR. k, Immunoblotting reveals that the levels of Vav2 expression were significantly down-regulated 48, 72 h following transfection with shVav2 #2 in N2a cells. Error bars represent s.e.m. i, k are cropped blot images from their corresponding full-length blots presented in Supplementary Figure 12.