Mapple lamp1

EGFP-LC3 was generated in Dr. M. Ward’s laboratory (NIH/NINDS) by cloning human LC3A into the lentiviral vector pLEX with a PGK promotor and an EGFP tag at the N terminus of LC3. α-Synuclein-mApple was generated by Epoch Life Science Inc. by cloning human SNCA into the pLEX vector with a PGK promotor and an mApple tag at the C terminus of α-Synuclein. EGFP-LC3 was generated previously (Cheng et al., 2015 (link)). α-Synuclein-A53T-RFP was a gift from Dr. H. Cai (NIH/NIA). Stealth siRNAs are from Thermo Fisher Scientific (Arl8a/b-siRNA#1: MSS290361 and Arl8b-siRNA MSS289173; Arl8a/b-siRNA#2: Arl8a-siRNA MSS229276 and Arl8b-siRNA MSS289171). mApple-Lamp1 was from Addgene (Cat# 54627), also named mApple-Lysosomes-20. Arl8b-mCherry is a gift from Dr. J. Bonifacino (NICHD/NIH). An siRNA-resistant Arl8b mutant was created by substituting nine nucleotides in the Arl8b-siRNA#1-targeting region (G459A, T462C, A463C, A465T, A468G, T471C, C474T, C477T and T480C).

Lentiviral constructs of caPAK5 and kdPAK5, which contain only the coding sequence (CDS) of the mRNAs, were generated by PCR and cloned into the entry vector pDONR221, followed by transferring into destination lentiviral vectors pHAGE-CMV-n-HA-FLAG as previously reported ^{9 (link)}. DsRed-Mito and GFP-Mito were gifts from R. Youle (NINDS, NIH). GO-ATeam2 and GO-ATeam3 were gifts from H. Imamura (Kyoto University, Japan). PAK5-shRNAs and Miro-1-shRNA were purchased from Dharmacon. Scramble shRNA, mApple-LAMP-1, caPAK1, caPAK5, kdPAK5, and caAKT1 constructs were purchased from Addgene. SNPH truncated mutants were generated in a previous study.^{19 (link)}