| Construct . | Source . |
|---|---|
| pEGFP-LC3B | Gift from T. Yoshimori, Osaka University, Osaka, Japan (rat) |
| mScarlet-LC3B | Subcloned from above with mScarlet (Addgene 85054) |
| mCh-EGFP-LC3 | Gift from T. Johansen, University of Tromsø, Tromsø, Norway |
| BFP-JIP1 | Human myc-JIP1 (gift from D. Holtzmann, Washington University in St. Louis, St. Louis, MO) subcloned into pBI-BFP plasmid |
| Halo-JIP1 | Subcloned from above with HaloTag on the N terminus of JIP1 in pEGFP vector backbone |
| Halo-HAP1 | HAP1 isoform 2 (gift from X. J. Li, Emory University, Atlanta, GA) subcloned into pHTN HaloTag vector (Promega, G7721) was lengthened into isoform 1 using transOMIC. The initial HAP1 isoform 2 differed from the National Center for Biotechnology Information database sequence by two codons and one amino acid; those differences were propagated to the final construct but had no effect on protein function or expression. HAP1 truncated and mutant constructs were all generated from this final construct. |
| HAP1-mCh-DHFR | mCherry, HAP1, and DHFR domains fused in pHTN vector backbone |
| Halo-JIP3 | HaloTag fused to the N terminus of hJIP3 from cDNA Clone (GE: MGC9053013) fused in pEGFP vector backbone |
| mCh-Htt | pARIS, Q23 (Pardo et al., 2010); gift from F. Saudou, Institut Curie, Paris, France |
| EGFP | pEGFP-N1 vector (Clontech, 6085-1) alone (for GFP fill) |
| HaloTag | pHTN vector with STOP codon following the HaloTag in order to express tag (for negative control) |
| PEX3-GFP-Halo | N-terminal 42 aa of human PEX3 for peroxisome targeting (Kapitein et al., 2010) generated in laboratory via PCR for a previous paper (Olenick et al., 2016) |
| BICD2N-Halo | Full-length mouse BICD2 in the pEGFP vector (gift from A. Akhmanova, Utrecht University, Utrecht, the Netherlands) was used to generate a truncated construct spanning residues 1–572 fused to the HaloTag and cloned into pcDNA3.1 |
| BICD2N-mCh-DHFR | Gift from E. Ballister, University of Pennsylvania, Philadelphia, PA |
| KHC-Halo | Full-length mouse kinesin-1 heavy chain (KIF5B) in pRK5 myc plasmid (gift from J. Kittler, University of Surrey, Guildford, UK) with HaloTag fused to the C terminus |
| K560-Halo | First 560 aa of human KIF5B from pET17: K560 GFP ST (gift from R. Vale, University of California, San Francisco, San Francisco, CA) subcloned into pHTC-HaloTag CMVneo vector (Promega) |
| EGFP-p25 | Gift from T. Schroer, Johns Hopkins University, Baltimore, MD |
| FLAG-p150 | Gift from T. Schwarz, Boston Children’s Hospital, Boston, MA |
| CBD-HAP1CC1 | Chitin binding domain (CBD) and HAP1 CC1 (residues 168-261) cloned in-frame into pRSF-Duet1 (EMD Millipore) |
| MBP-LIC1 (WT and FFAA) | Created in Lee et al. (2018) |
| LAMP1-EGFP | Subcloned from Addgene 1817, replacing RFP with EGFP |
| EGFP-Rab7 | Addgene 12605 |
| SNAP-Stx17 (FL and TM) | Addgene 45909-45910 subcloned into pSNAPf vector (New England Biolabs) |
| RILP-Halo | RILP (gift from J. Neefjes, Leiden University, Leiden, the Netherlands) subcloned into pHTC vector |
| Halo-Hook1 | WT and mutant constructs generated from human Hook1 sequence with HaloTag from the pHTN Halo tag CMV-neo vector (Promega; Olenick et al., 2016) |
| Construct . | Source . |
|---|---|
| pEGFP-LC3B | Gift from T. Yoshimori, Osaka University, Osaka, Japan (rat) |
| mScarlet-LC3B | Subcloned from above with mScarlet (Addgene 85054) |
| mCh-EGFP-LC3 | Gift from T. Johansen, University of Tromsø, Tromsø, Norway |
| BFP-JIP1 | Human myc-JIP1 (gift from D. Holtzmann, Washington University in St. Louis, St. Louis, MO) subcloned into pBI-BFP plasmid |
| Halo-JIP1 | Subcloned from above with HaloTag on the N terminus of JIP1 in pEGFP vector backbone |
| Halo-HAP1 | HAP1 isoform 2 (gift from X. J. Li, Emory University, Atlanta, GA) subcloned into pHTN HaloTag vector (Promega, G7721) was lengthened into isoform 1 using transOMIC. The initial HAP1 isoform 2 differed from the National Center for Biotechnology Information database sequence by two codons and one amino acid; those differences were propagated to the final construct but had no effect on protein function or expression. HAP1 truncated and mutant constructs were all generated from this final construct. |
| HAP1-mCh-DHFR | mCherry, HAP1, and DHFR domains fused in pHTN vector backbone |
| Halo-JIP3 | HaloTag fused to the N terminus of hJIP3 from cDNA Clone (GE: MGC9053013) fused in pEGFP vector backbone |
| mCh-Htt | pARIS, Q23 (Pardo et al., 2010); gift from F. Saudou, Institut Curie, Paris, France |
| EGFP | pEGFP-N1 vector (Clontech, 6085-1) alone (for GFP fill) |
| HaloTag | pHTN vector with STOP codon following the HaloTag in order to express tag (for negative control) |
| PEX3-GFP-Halo | N-terminal 42 aa of human PEX3 for peroxisome targeting (Kapitein et al., 2010) generated in laboratory via PCR for a previous paper (Olenick et al., 2016) |
| BICD2N-Halo | Full-length mouse BICD2 in the pEGFP vector (gift from A. Akhmanova, Utrecht University, Utrecht, the Netherlands) was used to generate a truncated construct spanning residues 1–572 fused to the HaloTag and cloned into pcDNA3.1 |
| BICD2N-mCh-DHFR | Gift from E. Ballister, University of Pennsylvania, Philadelphia, PA |
| KHC-Halo | Full-length mouse kinesin-1 heavy chain (KIF5B) in pRK5 myc plasmid (gift from J. Kittler, University of Surrey, Guildford, UK) with HaloTag fused to the C terminus |
| K560-Halo | First 560 aa of human KIF5B from pET17: K560 GFP ST (gift from R. Vale, University of California, San Francisco, San Francisco, CA) subcloned into pHTC-HaloTag CMVneo vector (Promega) |
| EGFP-p25 | Gift from T. Schroer, Johns Hopkins University, Baltimore, MD |
| FLAG-p150 | Gift from T. Schwarz, Boston Children’s Hospital, Boston, MA |
| CBD-HAP1CC1 | Chitin binding domain (CBD) and HAP1 CC1 (residues 168-261) cloned in-frame into pRSF-Duet1 (EMD Millipore) |
| MBP-LIC1 (WT and FFAA) | Created in Lee et al. (2018) |
| LAMP1-EGFP | Subcloned from Addgene 1817, replacing RFP with EGFP |
| EGFP-Rab7 | Addgene 12605 |
| SNAP-Stx17 (FL and TM) | Addgene 45909-45910 subcloned into pSNAPf vector (New England Biolabs) |
| RILP-Halo | RILP (gift from J. Neefjes, Leiden University, Leiden, the Netherlands) subcloned into pHTC vector |
| Halo-Hook1 | WT and mutant constructs generated from human Hook1 sequence with HaloTag from the pHTN Halo tag CMV-neo vector (Promega; Olenick et al., 2016) |