KEY RESOURCES TABLE
| REAGENT or RESOURCE | SOURCE | IDENTIFIER |
|---|---|---|
| Antibodies | ||
| Goat anti-Choline Acetyltransferase | Millipore/Sigma | (Millipore Cat# AB144P, RRID:AB_2079751) |
| Rabbit monoclonal anti-HA-tag (C29F4) - For Western Blot | Cell Signaling Technology | (Cell Signaling Technology Cat# 3724, RRID:AB_1549585) |
| Purified anti-HA.11 Epitope Tag Antibody (for RiboTag IP) |
Biolegend | (BioLegend Cat# 901503, RRID:AB_2565005) |
| mouse monoclonal anti-NeuN (A60) | Millipore | (Millipore Cat# MAB377, RRID:AB_2298772) |
| Rabbit anti-phospho-c-Jun (Ser 73) monoclonal (D47G9) | Cell Signaling Technology | (Cell Signaling Technology Cat# 3270, RRID:AB_2129575) |
| Rabbit polyclonal anti-STMN2/SCG10 | Novus | (Novus Cat# NBP1-49461, RRID:AB_10011569) |
| mouse anti-beta-Tubulin III monoclonal (clone 5H16) | Sigma-Aldrich | (Sigma-Aldrich Cat# T8578, RRID:AB_1841228) |
| Rabbit anti-ATF3 (polyclonal) | Novus | (Novus Cat# NBP1-85816, RRID:AB_11014863) |
| Rabbit anti-Iba1(polyclonal) | Wako | (FUJIFILM Wako Shibayagi Cat# 019–19741, RRID:AB_839504) |
| Rat anti-mouse CD68 monoclonal (clone FA-11) | Bio-Rad | (Bio-Rad Cat# MCA1957, RRID:AB_322219) |
| Rabbit anti-mouse c1q monoclonal (clone 4.8) | Abcam | (Abcam Cat# ab182451), RRID:AB_2732849) |
| Rabbit Monoclonal anti-C3 [EPR19394] | Abcam | (Abcam Cat# ab200999) RRID:AB_2924273 |
| Gt anti Rat Complement C3 | MP Biomedicals | MP Biomedicals (SKU:0855730) |
| Rat monoclonal anti-C3 [11H9] | Abcam | (Abcam Cat #ab11862) RRID:AB_10288263 |
| Rabbit polyclonal anti-C3 | Abcam | (Abcam Cat# ab11887) RRID:AB_298669 |
| Guinea pig anti-Synaptohysin-1 (polyclonal) | Synaptic Systems | (Synaptic Systems Cat# 101 004, RRID:AB_1210382) |
| mouse anti-Bassoon monoclonal (clone SAP7F407) | Abcam | (Abcam Cat# ab82958, RRID:AB_1860018) |
| Guinea pig anti-vesicular glutamate transporter 1 (VGlut1, polyclonal) | Millipore | (Millipore Cat# AB5905, RRID:AB_2301751) |
| Guinea pig anti-VAChT (polyclonal) | Synaptic Systems | (Synaptic Systems Cat# 139 105, RRID:AB_10893979) |
| Alpha-Bungarotoxin - 543 | Biotium | 00026 |
| Goat anti-Rabbit - 568 | ThermoFisher | (Thermo Fisher Scientific Cat# A-11011, RRID:AB_143157) |
| Goat anti -Rabbit - 488 | Jackson Immuno | (Jackson ImmunoResearch Labs Cat# 111-545-003, RRID:AB_2338046) |
| Goat anti-Mouse - 647 | Fisher Scientific | (Thermo Fisher Scientific Cat# A-21235, RRID:AB_2535804) |
| Goat anti-mouse - IgG1 - 568 | Fisher Scientific | (Thermo Fisher Scientific Cat# A-21124, RRID:AB_2535766) |
| Goat anti-mouse - IgG2a - 568 | Fisher Scientific | (Thermo Fisher Scientific Cat# A-21134, RRID:AB_2535773) |
| Goat anti - Rat - 647 | Fisher Scientific | (Thermo Fisher Scientific Cat# A-21247, RRID:AB_141778) |
| Goat anti-Mouse - 488 | Fisher Scientific | (Thermo Fisher Scientific Cat# A32723, RRID:AB_2633275) |
| Goat anti- Mouse IgG1 - 488 | Fisher Scientific | (Thermo Fisher Scientific Cat# A-21121, RRID:AB_2535764) |
| Goat anti- Mouse IgG2a - 488 | Fisher Scientific | (Thermo Fisher Scientific Cat# A-21131, RRID:AB_2535771) |
| Goat anti- Mouse IgG2a - 647 | Fisher Scientific | (Thermo Fisher Scientific Cat# A-21241, RRID:AB_2535810) |
| Goat anti- Mouse IgG1 - 647 | Fisher Scientific | A21246 |
| Goat anti Rabbit - 405 | Abcam | (Abcam Cat# ab175652, RRID:AB_2687498) |
| Donkey anti Rabbit - 405 | Abcam | (Abcam Cat# ab175649, RRID:AB_2715515) |
| Donkey anti Mouse - 543 | Biotium | (Biotium Cat# 20305, RRID:AB_2923245) |
| Donkey anti Rabbit - 543 | Biotium | 20308 |
| Donkey anti Mouse - 488 | Biotium | (Biotium Cat# 20014, RRID:AB_10561327) |
| Alexa Fluor® 488 AffiniPure Donkey Anti-Guinea Pig IgG (H + L) | Jackson Immuno | (Jackson ImmunoResearch Labs Cat# 706-545-148, RRID:AB_2340472) |
| Donkey Anti-Rabbit IgG −488 | Biotium | (Biotium Cat# 20015-1, RRID:AB_10854232) |
| Chemicals, peptides, and recombinant proteins | ||
| SMART-Seq v4 Ultra Low Input RNA Kit | Takara Bio | 634889 |
| TSMARTer® DNA Unique Dual Index Kit for library Prep |
Takara Bio | R400665–R400668 |
| RNasin Plus RNase Inhibitor | Promega | N2615 |
| Cycloheximide | Sigma | C7698-5G |
| Protease Inhibitor Cocktail | Sigma | P8340-5mL |
| Heparin sodium salt from porcine intestinal mucosa | Sigma | H3393-50KU |
| Superscript III First-Strand Synthesis SuperMix |
Invitrogen | 11752–250 |
| FastStart Universal SYBR Green Master Mix (ROX) |
Fisher Scientific | 4913850001 |
| M.O.M blocking reagent | Vector Laboratories | 27090 |
| Molecular Biology Grade Ethanol | Fisher Scientific | BP2818100 |
| Paraformaldehyde | EMS | 15710 |
| Normal Goat serum | Fisher Scientific | 16210064 |
| Normal Donkey Serum | Fisher Scientific | NC9624464 |
| Beta Mercaptoethanol | Sigma | M6250 |
| RNeasy RLT buffer | Qiagen | 79216 |
| Tissue Plus OCT compound Clear | Fisher Scientific | 4585 |
| Deposited data | ||
| Raw RNA sequencing Files | GEO: GSE213987 | https://www.ncbi.nlm.nih.gov/geo |
| Experimental models: Organisms/strains | ||
| Dlk fx/fx | (Miller et al. 2009)28 | |
| RiboTag: B6N.129-Rpl122tm1.1Psam/J | Jackson laboratories | 011029 |
|
ChAT-IRES-Cre: B6; 129S6-ChATtm(Cre)/Lowl/J |
Jackson laboratories | 006410 |
| Rosa-tdTomato line (Ai14) | Jackson laboratories | 007914 |
| Tmem119-2A-EGFP reporter mouse line | Jackson laboratories | 031823 |
| C3 Knockout: B6.129S4-C3tm1Crr/J | Jackson laboratories | 029661 |
| Oligonucleotides | ||
| See Table S1 | ||
| Software and algorithms | ||
| R | R Foundation for Statistical Computing, Vienna, Austria |
R Foundation for Statistical Computing, Vienna, Austria |
| R.Studio | posit | https://posit.co/download/rstudio-desktop/ |
| GraphPad Prism 9 | GraphPad Software, La Jolla, CA | https://www.graphpad.com/scientific-software/prism/ |
| FastQC | Babraham Institute Bioinformatics | https://github.com/s-andrews/FastQC |
| BBDuK from BBtools | BBMap – Bushnell B. | sourceforge.net/projects/bbmap/ |
| STAR/2.5.2a aligner | (Dobin et al. 2013)72 | https://github.com/alexdobin/STAR |
| Samtools | 73 | https://samtools.sourceforge.net/ |
| QoRTs | (Hartley and Mullikin 2015)74 | https://github.com/hartleys/QoRTs |
| DESeq2 | (Love, Anders, and Huber 2014)75 | https://github.com/mikelove/DESeq2 |
| ggplot2 | (Wickham 2011)76 | https://github.com/tidyverse/ggplot2 |
| EnhancedVolcano | (Blighe, Rana, and Lewis 2019)77 | https://github.com/kevinblighe/EnhancedVolcano |
| DAVID (The Database for Annotation, Visualization and Integrated Discovery) | (“DAVID Functional Annotation Bioinformatics Microarray Analysis”) |
https://david.ncifcrf.gov/ |
| GeneSCF | (Subhash and Kanduri 2016)78 | https://github.com/genescf/GeneSCF |
| GoPlot | (Walter, Sánchez-Cabo, and Ricote 2015)79 | https://wencke.github.io/ |
| Volocity Image analysis software (6.3.1) | Perkin Elmer/Quarum Technologies | Volocity 3D Image Analysis Software (RRID:SCR_002668) |
| Other | ||
| Mm- Chat - C2 (in situ probe) | ACD | 408731-C2 |
| Mm- Atf3 - C3 (in situ probe) | ACD | 426891-C3 |
| Mm- C1qb - C1 (in situ probe) | ACD | 438101 |
| TSA® Plus Fluorescein | Akoya Biosciences | NEL741001KT |
| TSA® Plus Cyanine 5 (Cy5) | Akoya Biosciences | NEL745001KT |
| TSA® Plus Cyanine (Cy3) | Akoya Biosciences | NEL744001KT |
| RNAscope® Multiplex Fluorescent Reagent Kit V2 |
ACD | 323100 |
| Protein G Magnetic Beads | NEB | S1430S |
| RNeasy Plus Micro Kit | Qiagen | 74034 |
| DAPI Fluoromount-G | Southern biotech | 0100–20 |
| Fluoromount-G anti-fade | Southern biotech | 0100–35 |
| DynaMag™−2 Magnet | ThermoFisher | 12321D |
| ImmEdge hydrophobic pen | Vector Biosciences | H-4000 |
| Csf1r inhibitor PLX5622 | Plexxikon | |
| Chow made with PLX5622 or Control Diet (AIN-76A formula) | Research Diets | |
| Fine forceps for SNC | Fine Science Tools | 11399–80 |