KEY RESOURCES TABLE
| REAGENT or RESOURCE | SOURCE | IDENTIFIER |
|---|---|---|
| Antibodies | ||
|
| ||
| V5 Tag Monoclonal Antibody | Invitrogen | Cat# R96025 |
| Rabbit Anti-V5 Antibody | Novus | Cat# NB600-381 |
| β-Tubulin (9F3) Rabbit mAb | Cell Signaling Technologies | Cat# 2128 |
| Anti-c-Myc Antibody (Clone #9E10) | Sigma-Aldrich | Cat# M4439 |
| Rabbit Anti-HA Antibody | Sigma-Aldrich | Cat# H6908 |
| Anti-Tubulin Beta 3 (TUBB3) Antibody-Cy5 conjugated | Biolegend | Cat# 6S7406; RRID: AB_2563610 |
| Anti-Isl ½ Antibody | Developmental Studies Hybridoma Bank | Cat# 39.4DS; RRID: AB_2314683 |
| Anti AcetylCholine Transferase Antibody | Sigma-Aldrich | Cat# AB144P |
| MAP2 Polyclonal Antibody | Proteintech | Cat# 17490-1-AP |
| Human/Mouse/Rat Sox2 Antibody (Clone #245610) | R & D Systems | Cat# MAB2018 |
| Human Oct4 Antibody (Clone #653108) | R & D Systems | Cat# MAB17591 |
| Anti-Kif5A Antibody | Genetex | Cat# GTX113761; RRID: AB_2037309 |
| RBM24 Polyclonal Antibody | Invitrogen | Cat# PAS-66881 |
| Rabbit Anti-RAN Antibody | Bethyl Laboratories | Cat# A304-297A |
| Anti-mutKif5A Antibody | This paper, Genewiz | N/A |
| Anti-GAPDH Antibody | Novus Biologicals | Cat# NB300-327 |
| Rabbit Anti-HA Tag Antibody | Sigma-Aldrich | Cat# H6908 |
| Goat Anti-Horseradish Peroxidase Antibody | Jackson Immunoresearch | Cat# 123-165-021; RRID: AB_2338959 |
| Donkey Anti-Mouse 488 | Jackson Immunoresearch | RRID: AB_2340846 |
| Donkey Anti-Rabbit IgG 647 | Jackson Immunoresearch | RRID: AB_2492288 |
| Donkey Anti-Rabbit IgG 594 | Jackson Immunoresearch | RRID: AB_2340621 |
| Donkey anti-Mouse 546 | Invitrogen | Cat# A10036 |
| Mouse anti-Hemagglutinin (HA) antibody (12CA5) | Invitrogen | Cat# MA1-12429; RRID: AB_1074049 |
| Goat anti-Mouse FITC | Jackson Immunoresearch | Cat# 115-095-003; RRID: AB_2338589 |
| Goat anti-Mouse TRITC | Jackson Immunoresearch | Cat#115-025-003; RRID: AB_2338478 |
| Rabbit anti-GFP | Proteintech | Cat# 50430-2-AP |
| Goat anti-Mouse HRP | Jackson Immunoresearch | Cat# 115-035-003; RRID: AB_10015289 |
| Goat anti-Rabbit HRP | Jackson Immunoresearch | Cat# 111-004-003; RRID: AB_2337913 |
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| Biological samples | ||
|
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| Patient Kif5A iPSC | Applied Stem Cell; This paper | N/A |
| Isogenic Control iPSC | Applied Stem Cell | ASE-9109 |
|
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| Chemicals, peptides, and recombinant proteins | ||
|
| ||
| FuGENE6 transfection reagent | Promega | Cat# E269A |
| Tetramethylrhodamine (TMR) Halo ligand | Promega | Cat# G8251 |
| Guanosine-5’-[(α,β)-methyleno] triphosphate, sodium salt (GMPCPP) | Jena Bioscience | Cat# NU-405 |
| Porcine brain tubulin | Cytoskeleton | Cat# T240 |
| HiLyte488-labeled tubulin | Cytoskeleton | Cat# TL488M |
| Purified Rigor Kinesin | Masucci et al. (2021) | N/A |
| PlusOne Repel-Silane ES | Cytiva | Cat# 17133201 |
| Pluronic F-127 | Sigma-Aldrich | Cat# P2443 |
| Methyl cellulose | Sigma-Aldrich | Cat# M0512 |
| Adenosine 5’-triphosphate magnesium salt (ATP) | Sigma-Aldrich | Cat# A9187 |
| Guanosine 5’-triphosphate sodium salt hydrate (GTP) | Sigma-Aldrich | Cat# G8877 |
| 0.05% Trypsin EDTA | Gibco | Cat# 25300054 |
| Poly-D-Lysine | Sigma-Aldrich | Cat# P7280-5X5MG |
| Natural Mouse Laminin | Corning | Cat# 354232 |
| Neurobasal Medium, Minus Phenol Red | Gibco | Cat# 12348-017 |
| B27 Supplement | Gibco | Cat# 17504-044 |
| Glutamax | Gibco | Cat# 35050061 |
| Opti-MEM I Reduced Serum Medium, no phenol red | Gibco | Cat# 11058021 |
| Lipofectamine 2000 Transfection Reagent | Invitrogen | Cat# 11668019 |
| 2-Mercaptoethanol, 99% pure | Fisher Scientific | Cat# AC125470100 |
| Dulbecco’s Modified Eagle Medium (DMEM) | Gibco | Cat# 11965118 |
| Fetal Bovine Serum, Regular | MediaTech | Cat# 35-010-CV |
| L-glutamine (200 mM) | Gibco | Cat# 25030081 |
| Penicillin/Streptomycin | Gibco | Cat# 15140122 |
| NeuroMag Transfection Reagent | Oz Biosciences | Cat# NM50200 |
| Minimum Essential Media (MEM) | Gibco | Cat# 11095098 |
| BSA (heat shock fraction, protease free) | Sigma-Aldrich | Cat# A3294 |
| Triton X- | Sigma-Aldrich | Cat# T9284 |
| Antigen Retrieval Buffer | Abcam | Cat# 93678 |
| Prolong Gold Antifade Mountant with DAPI | Invitrogen | Cat# P-36931 |
| Hank’s Balanced Salt Solution (HBSS) | Gibco | Cat# 14175095 |
| SUPERaseIn RNase Inhibitor | Invitrogen | Cat# AM2694 |
| TRIzol Reagent | Life Technologies | Cat# 15596026 |
| Denaturing Hypotonic Lysis buffer (DHL) | Singh et al. (2014) | N/A |
| Alt-R S.p. HiFi Cas9 Nuclease v3 | Integrated DNA Technologies | Cat# 1081060 |
| Blasticidin S Hydrochloride Powder | Research Products International | Cat# B12200 |
| Ultrapure DNase/RNase-Free Distilled Water | Invitrogen | Cat# 10977015 |
| Sail Restriction Enzyme | New England Biolabs | Cat# R0138L |
| Notl Restriction Enzyme | New England Biolabs | Cat# R3198L |
| Proteinase K | Thermo Scientific | Cat# FEREO0491 |
| Laemmli SDS-Sample Buffer | Boston BioProducts | Cat# BP-111 R |
| ‘Soft Elution Buffer’ | Antrobus and Borner (2011) | N/A |
| Dulbecco’s Phosphate Buffered Saline | Corning | Cat# 20-031-CV |
| Tween 20 | Fisher BioReagents | Cat# BP337-500 |
| Dithiothrieitol Solution | Sigma-Aldrich | Cat# 43816 |
| Trypsin-EDTA | Gibco | Cat# 15400054 |
| Odyssey Blocking Buffer | Licor | Cat# 927-40010 |
| Dulbecco’s Modified Eagle Medium | Sigma-Aldrich | Cat# D7777 |
| Heat-Inactivated Fetal Bovine Serum | Corning | Cat# MT35011CV |
| All trans Retinoic Acid | Sigma-Aldrich | Cat# R2625 |
| Lipofectamine 3000 | Invitrogen | Cat# L3000015 |
| Poly-DL-Lysine | Sigma-Aldrich | Cat# P9011 |
| Formaldehyde | Fisher | Cat# BP531-500 |
| BSA (lyophilized powder) | Sigma-Aldrich | Cat# A9418 |
| Triton X-100 | Sigma-Aldrich | Cat# X-100 |
| Mowiol 4-88 Reagent | Calbiochem | Cat# 47-590-4100GM |
| Sepharose GFP-Binder Beads | Gift from Vladimir Gelfand Lab | N/A |
| Accutase Cell Detachment Solution | Corning | Cat# MT25058CI |
| Matrigel hESC-Qualified Matrix | Corning | Cat# 08-774-552 |
| ROCK Inhibitor | Selleck Chemicals | Cat# S1049 |
| Lipofectamine Stem Transfection Reagent | Invitrogen | Cat# STEM00001 |
| Chelex 100 Sodium form | Sigma-Aldrich | Cat# C7901 |
| Stemflex Medium | Gibco | Cat# A3349401 |
| B27 Plus Neuronal Culture System | Gibco | Cat# A3653401 |
| Poly D-L-Ornithine | Sigma | Cat# P3655 (3000-7000 kDa) |
| Laminin | Invitrogen | Cat# 23017-015 |
| Neurobasal medium | Invitrogen | Cat# 21103-049 |
| Glutamax 200mM | Invitrogen | Cat# 35050-061 |
| B27 | Invitrogen | Cat# 17504-044 |
| hsBDNF | PeproTech | Cat# 450-02 |
| rnCNTF | PeproTech | Cat# 450-50 |
| mmGDNF | PeproTech | Cat# 450-44 |
| Horse Serum | Sigma | Cat# H1138-100ML |
| BSA | Gold Biotechnology | Cat# A-421-50 |
| 4-20% Mini-PROTEAN® TGX™ Gel, 15 well, 15 μL | Bio-Rad | Cat# 456-1096 |
| Nitrocellulose membrane/filter paper pack, pkg of 50 | Bo-Rad | Cat# 1620213 |
| Protease inhibitor tables | Roche | Cat# 11873580001 |
| NeuroMag paramagnetic beads | Oz Biosciences | Cat# NM50200 |
| Magnetic plate | Oz Biosciences | Cat# MF10000 |
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| Critical commercial assays | ||
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| Pierce BCA Protein Assay Kit | ThermoFisher Scientific | Cat# 23227 |
| AmpliTaq Gold 360 Master Mix | Applied Biosystems | Cat# 4398876 |
| High Capacity cDNA Reverse Transcription Kit | Applied Biosystems | Cat# 436814 |
| TaqMan Fast Advanced Master Mix | Applied Biosystems | Cat# 4444556 |
| Taqman Gene Expression Assay, SLITRK2 | ThermoFisher Scientific | Hs01028461_s1 |
| Taqman Gene Expression Assay, TCEAL2 | ThermoFisher Scientific | Hs04194669_sH |
| Taqman Gene Expression Assay, PCDHA13 | ThermoFisher Scientific | Hs00259032_s1 |
| Taqman Gene Expression Assay, HNRNPM | ThermoFisher Scientific | Hs01115690_m1 |
| Taqman Gene Expression Assay, DNAJA4 | ThermoFisher Scientific | Hs00388055_m1 |
| Taqman Gene Expression Assay, ELAVL4 | ThermoFisher Scientific | Hs00956610_mH |
| Taqman Gene Expression Assay, GAPDH | ThermoFisher Scientific | Hs02786624_g1 |
| NEBuilder HiFi DNA Assembly Cloning Kit | New England Biolabs | Cat# E5520S |
| HiSpeed Plasmid Midi Kit | Qiagen | Cat# 12662 |
| PureLink HiPure Plasmid Midiprep Kit | Invitrogen | Cat# K210005 |
|
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| Deposited data | ||
|
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| Sequence data, analyses, and resources related to the RIPseq of V5-KIF5AΔ and V5-KIF5AΔExon27 immunoprecipitation eluates | This paper | Gene Expression Omnibus GSE196539 |
| Sequence data, analyses, and resources related to the RNA sequencing of KIF5AR1007K and isogenic control iMNs | This paper | Gene Expression Omnibus GSE196539 |
| Sequence data, analyses and resources related to the mass spectrometry analysis of V5-KIF5AwT and V5-KIF5AΔExon27 immunoprecipitation eluates | This paper | ProteomeXchange; PXD031012 |
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| Experimental models: Cell lines | ||
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| COS-7 | ATCC | CRL-1651; RRID:CVCL_0224 |
| SKNAS | ATCC | CRL-2137; RRID:CVCL_1700 |
| HEK293FT | Invitrogen | R70007 |
| Neuro2A | ATCC | CCL-131; RRID:CVCL_0470 |
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| Experimental models: Organisms/strains | ||
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| C57BL/6 female mice (primary neuron preps) | Charles River Laboratories | Cat# 027 |
|
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| Oligonucleotides | ||
|
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| sgRNA targeting CYBL locus - ATGTTGGAAGGATGAGGAAA | Fernandopulle et al. (2018) | N/A |
| Primer for HA-KIF5A Forward − ACTGACGCTAGCCACCATGTACCCATACGATGTTCCAGATTACGCTGGTGGTTCTGGTGGTGGTTCTGGTATGGCGGAGACCAACAACG | This Paper | N/A |
| Primer for HA-KIF5A WT Reverse − AGAATCGGATCCTGTGGGAGATTAGCTGGCTG | This Paper | N/A |
| Primer for HA-KIF5A Del27 Reverse − AGATACGGATCCCGAAGTTATGGTACCTTAGAAACTGA | This Paper | N/A |
| Primer for HA-KIF5A C-Term Reverse − AGAATCGGATCCTTATCCATTGTCCATGTTGGCCTT | This Paper | N/A |
| Primer for eGFP Forward − AGTCAGGCTAGCCACCATGGTGAGCAAGGGAGAGGAG | This Paper | N/A |
| Primer for eGFP Reverse − AGATACGGATCCCTTGTACAGCTCGTCCATGCC | This Paper | N/A |
| Primer for GFP-KIF5A Forward − | This Paper | N/A |
| AGTCAGGGATCCGGTGGTTCTGGTGGTGGTTCTGGTATGGCGGAGACCAACAACG | ||
| Primer for GFP-KIF5A WT Reverse − AGATACTCTAGATGTGGGAGATTAGCTGGCTG | This Paper | N/A |
|
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| Recombinant DNA | ||
|
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| DsRed-Mito | Takara Bio | Cat# 632421 |
| pRK5:Myc-KIF5A | Addgene | Clone# 127616 |
| pRK5-Myc-KIF5AWT-Halo | This paper | N/A |
| pRK5-Myc- KIF5AΔExon27-Halo | This paper | N/A |
| pRK5-Myc-KIF5AΔC-term-Halo | This paper | N/A |
| pRK5-Myc-KIF5A1-560-Halo | This paper | N/A |
| pcDNA4TO-K560-E236A-24xGCN4_v1-IRES-Puro | Tanenbaum et al. (2014) | Addgene Clone #60909 |
| pGW1-Kif5AWT | This paper | N/A |
| pGW1-Kif5AΔExon27 | This paper | N/A |
| pGW1-Kif5AΔC-term | This paper | N/A |
| pGW1-tdTomato | This paper | N/A |
| V5-Kif5AΔExon27 | This paper | Addgene Clone #15239 |
| V5-Kif5AWT | This paper | Addgene Clone #15239 |
| GFP-Kif5AWT | This paper | Addgene Clone #15239 |
| HA-Kif5AWT | This paper | Addgene Clone #15239 |
| Kif5AAExon27 | This paper | N/A |
| pCMV-beta-Rat Lic1.FL-HA | Tynan et al. (2000) | N/A |
| pMyc-MOV10 | Addgene | Cat# 10977 |
| RNT1-GFP | Addgene | Cat# 17708 |
| HA-p62 | Addgene | Cat# 28027 |
| pCDNA 3.1 (+) HA- KIF5A WT | This Paper | N/A |
| pCDNA 3.1 (+) HA- KIF5A Del27 | This Paper | N/A |
| pCDNA 3.1 (+) HA- KIF5A C-Term | This Paper | N/A |
| pCDNA 3.1 (+) GFP-KIF5A WT | This Paper | N/A |
| pCDNA 3.1 (+) GFP- KIF5A Del27 | This Paper | N/A |
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| Software and algorithms | ||
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| ||
| FastQC | Babraham Institute | http://www.bioinformatics.babraham.ac.uk/projects/fastqc/ |
| MultiQC | Ewels et al. (2016) | https://github.com/ewels/MultiQC |
| STAR aligner (2.7.0a) | Dobin et al. (2013) | https://github.com/alexdobin/STAR |
| HTSeq | Anders et al. (2015) | https://htseq.readthedocs.io/en/master/overview.html |
| DESeq2 | Love et al. (2014) | https://bioconductor.org/packages/release/bioc/html/DESeq2.html |
| rMATS version 4.0.2 | Shenetal. (2014) | http://rnaseq-mats.sourceforge.net/ |
| Metascape | Zhou etal. (2019) | https://metascape.org/gp/index.html#/main/step1 |
| rMAPS2 | Hwang et al. (2020) | http://rmaps.cecsresearch.org/ |
| ImageJ | NIH | https://imagej.nih.gov/ij/ |
| MTrackJ | NIH | https://imagescience.org/meijering/software/mtrackj/ |
| Volocity | PerkinElmer | https://www.perkinelmer.com |
| Prism 9 | GraphPad | https://www.graphpad.com/scientific-software/prism |
| Survival Analysis YOLO | This paper | https://drive.google.com/drive/folders/1V36q_9QjX_0SHbLS_IN2PPngOSLKYS1_?usp=sharing |
| Biorender | Biorender | https://www.biorender.com |
| AutoQuant x3 | Media Cybernetics | https://www.mediacy.com/79-products/autoquant-x3 |
| Proteome Discoverer 2.1.1.21 | Thermo Scientific | https://www.thermofisher.com/us/en/home/industrial/mass-spectrometry/liquid-chromatography-mass-spectrometry-lc-ms/lc-ms-software/multi-omics-data-analysis/proteome-discoverer-software.html |
| Mascot Server 2.6.2 | Matrix Science Ltd. | http://www.matrixscience.com/ |
| Scaffold 4.10.0 | Proteome Software, Inc. | https://www.proteomesoftware.com/products/scaffold-5 |
| Peptide Prophet | Keller et al. (2002); Nesvizhskii et al. (2003) | http://peptideprophet.sourceforge.net/ |
| PyMOL | The PyMOL Molecular Graphics System, Version 2.0 Schrödinger, LLC | https://pymol.org/2/ |
| I-TASSER | Yang et al. (2015) | https://zhanggroup.org/I-TASSER/ |
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| Other | ||
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| Cytation 5 Cell Imaging Multi Mode Reader | Agilent (BioTek) | https://www.biotek.com/products/imaging-microscopy-cell-imaging-multi-mode-readers/cytation-5-cell-imaging-multi-mode-reader/ |
| BioSpa 8 Automated Incubator | Agilent (BioTek) | https://www.biotek.com/products/software-robotics-robotics/biospa-8-automated-incubator/ |
| Nikon Ti Eclipse | Nikon | Ti Eclipse |
| Leica DMI6000 Widefield Microscope | Leica Microsystems | DMI6000 |
| Anti-V5 Tagged Magnetic Beads | MBL International | Cat#M167-11 |
| CFX 384 Real-Time PCR Detection System | BioRad | https://www.bio-rad.com/en-us/product/cfx384-touch-real-time-pcr-detection-system?ID=LJB22YE8Z |
| C1000 Touch Thermal Cycler | BioRad | https://www.bio-rad.com/en-us/sku/1851138-c1000-touch-thermal-cycler-with-384-well-reaction-module?ID=1851138 |
| Trans-Blot Turbo Transfer System | BioRad | Cat# 1704150 |