Key resources table.
| Reagent type (species) or resource |
Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| Gene (Drosophila melanogaster) | not | NA | FLYB: FBgn0013717 | |
| Gene (D. melanogaster) | Atxn7 | NA | FLYB: FBgn0031420 | |
| Gene (D. melanogaster) | SCAR | NA | FLYB: FBgn0041781 | |
| Gene (D. melanogaster) | Gcn5 | NA | FLYB: FBgn0020388 | |
| Gene (D. melanogaster) | Ada2b | NA | FLYB: FBgn0037555 | |
| Strain, strain background (D. melanogaster) | Elav-Gal4 | Bloomington Drosophila Stock Center | BDSC:458 RRID:BDSC_458 | Genotype:P(w[+mW.hs]=GawB)elav[C155] |
| Strain, strain background (D. melanogaster) | Uas-Gcn5 RNAi | Vienna Drosophila Resource Center | VDRC:21786 RRID:FlyBase_FBst0454233 | Construct ID:11218 |
| Strain, strain background (D. melanogaster) | Uas-SCAR RNAi | Bloomington Drosophila Stock Center | BDSC:36121 RRID:BDSC_36121 |
Genotype: y[1] sc[*] v[1]; P(y[+t7.7] v[+t1.8]=TRiP.HMS01536)attP40 |
| Strain, strain background (D. melanogaster) | Uas-Not RNAi | Bloomington Drosophila Stock Center | BDSC:28725 rebalanced with Tm6b, Tb RRID:BDSC_28725 |
Genotype: y[1] v[1]; P(y[+t7.7] v[+t1.8]=TRiP.JF03152)attP2/TM6B, Tb |
| Strain, strain background (D. melanogaster) | Uas-Atxn7 RNAi | Vienna Drosophila Resource Center | VDRC:102078 RRID:FlyBase_FBst0473949 | Construct ID:110634 |
| Strain, strain background (D. melanogaster) | OreR | DGGR | Catalog number: 109612 RRID:DGGR_109612 |
|
| Strain, strain background (D. melanogaster) | Non-stop[02069] | Bloomington Drosophila Stock Center | BDSC:11553 Rebalanced with Tm3 GFP RRID:BDSC_11553 | Genotype: P(ry[+t7.2]=PZ)not[02069] ry[506]/TM3, P(w[+mC]=GAL4 twi.G)2.3, P(UAS-2xEGFP)AH2.3, Sb[1] Ser[1] |
| Strain, strain background (D. melanogaster) | SCAR [Delta37] | Bloomington Drosophila Stock Center | BDSC:8754 Rebalanced with CyoGFP RRID:BDSC_8754 |
Genotype: w[*]; SCAR[Delta37] P(ry[+t7.2]=neoFRT)40A/CyO,, P(w[+mC]=GAL4 twi.G)2.2, P(UAS-2xEGFP)AH2.2. Cross BDSC:6662 and BDSC:8754 |
| Strain, strain background (D. melanogaster) | Atxn7[KG02020] | Bloomington Drosophila Stock Center | BDSC: 14255 Rebalanced with CyoGFP RRID:BDSC_14255 |
Genotype: y[1] w[67c23]; P(y[+mDint2] w[BR.E.BR]=SUPor P)CG9866[KG02020]/CyO, P(w[+mC]=GAL4 twi.G)2.2, P(UAS-2xEGFP)AH2.2. |
| Strain, strain background (D. melanogaster) | Uas-Atxn7 RNAi, SCARΔ37 | This Paper | Materials and methods Subsection Fly Strains Genotype: w[*]; SCAR[Delta37] P(ry[+t7.2]=neoFRT)40A/CyO, P(w[+mC]=GAL4 twi.G)2.2, P(UAS-2xEGFP)AH2.2.; P{KK110634} VIE-260B/TM6C, cu[1] Sb[1] Tb[1] |
|
| Cell line (D. melanogaster) | ML-DmBG3-c2 | Drosophila Genomics Resource Center #68 | FLYB: FBtc0000068; RRID:CVCL_Z728 | FlyBase symbol: ML-DmBG3-c2 |
| Cell line (D. melanogaster) | S2-DRSC | Drosophila Genomics Resource Center #181 | FLYB: FBtc0000181; RRID:CVCL_Z992 | FlyBase symbol: S2-DRSC |
| Antibody | Guinea Pig anti Non-stop | (Mohan et al., 2014a) | Western Blot Dilution (1:1000) IF Dilution (1:150) | |
| Antibody | Mouse anti Chaoptin (Monoclonal) | Developmental Studies Hybridoma Bank | 24B10 RRID:AB_528161 |
IF Dilution (1:250) |
| Antibody | Goat anti Rat IGG −568 (Polyclonal) | Invitrogen | A-11077 RRID:AB_141874 |
IF Dilution (1:1000) |
| Antibody | Goat anti Mouse IGG-488 (Polyclonal) | Invitrogen | A-11001 RRID:AB_2534069 |
IF Dilution (1:1000) |
| Antibody | Goat anti Guinea pig IGG- 488 (Polyclonal) | Invitrogen | A11073 RRID:AB_142018 |
IF Dilution (1:1000) |
| Other | Phalloidin-488 | Invitrogen | A12379 | IF Dilution (1:20) |
| Other | Phalloidin-568 | Invitrogen | A12380 RRID:AB_2810839 |
IF Dilution (1:20) |
| Other | Vecta Shield | Vector Labs | H-1200 | |
| Antibody | Rat anti HA-HRP (Monoclonal) | Roche | 12013819001 RRID:AB_390917 |
Western Blot Dilution (1:500) |
| Antibody | Mouse anti SCAR (Monoclonal) | Developmental Studies Hybridoma Bank | P1C1-SCAR RRID:AB_2618386 |
Western Blot Dilution (1:250) IF Dilution (1:100) |
| Antibody | Rabbit anti Atxn7 | (Mohan et al., 2014a) | Western Blot Dilution (1:2000) | |
| Antibody | Guinea Pig anti Ada2b | Gift from Jerry L Workman (Kusch et al., 2003) | Western Blot Dilution (1:1000) | |
| Antibody | Goat anti Guinea Pig HRP (Polyclonal) | Jackson ImmunoResearch INC | 106-035-003, RRID:AB_2337402 | Western Blot Dilution (1:10000) |
| Antibody | Goat anti mouse HRP (Polyclonal) | Jackson ImmunoResearch INC | 115-035-003, RRID:AB_10015289 |
Western Blot Dilution (1:5000) |
| Antibody | Goat anti Rabbit HRP (Polyclonal) | Jackson ImmunoResearch INC | 111-035-003 RRID:AB_2313567 |
Western Blot Dilution (1:10000) |
| Antibody | Rat anti HA (Monoclonal) | Roche | 11867423001 RRID:AB_390918 |
IF Dilution (1:250) |
| Recombinant DNA reagent | pMT-HA-Ub | gift from Jianhang Jia | gift from Jianhang Jia | |
| Recombinant DNA reagent | Scar-FH | Berkley Expression Clone Collection | FMO14142 | |
| Recombinant DNA reagent | Scar-Flag | This paper | Materials and methods Subsection Plasmids Quick change on FMO14142 F Primer:GATGACGACAAGGTCAAACTTGCTGCTTAGACTAGTTCTAGT R Primer: ACTAGAACTAGTCTAAGCAGCAAGTTTGACCTTGTCGTCATC |
|
| Recombinant DNA reagent | Prmha3-Non-stop-2XFlag-2XHA | This Paper | Sequence ID: AAD53181.1 | Materials and methods Subsection Plasmids |
| Recombinant DNA reagent | Prmha3-Non-stop-2XFlag-2XHA-0FA | This Paper | Materials and methods Subsection Plasmids Quick change mutagenesis on Prmha3-Non-stop-2XFlag-2XHA WIRS0 phenylalanine to alanine F: GCAGTGGCCGAAGCGCCGGCAGGGGAACGGAACGGTGGGC WIRS0 phenylalanine to alanine R: CCGTTCCCCTGCCGGCGCTTCGGCCACTGCTGCTGCTGC |
|
| Recombinant DNA reagent | Prmha3-Non-stop-2XFlag-2XHA-1FA | This Paper | Materials and methods Subsection Plasmids Quick change mutagenesis on Prmha3-Non-stop-2XFlag-2XHA WIRS1 phenylalanine to alanine F: CAGCTACGATACAGCCCGGGTCATCGACGCCTACTTCGCTGCTTGCG WIRS1 phenylalanine to alanine R: GGCGTCGATGACCCGGGCTGTATCGTAGCTGTGCTCCTTCACATAGC |
|
| Recombinant DNA reagent | Prmha3-Non-stop-2XFlag-2XHA-2FA | This Paper | Materials and methods Subsection Plasmids Quick change mutagenesis on Prmha3-Non-stop-2XFlag-2XHA WIRS2 phenylalanine to alanine F: CCAGCGTGGTGTCGGCCCATTTGAAACGCTTCGAGCACTCAGCTCTG WIRS2 phenylalanine to alanine R: CGAAGCGTTTCAAATGGGCCGACACCACGCTGGGCAGAGTGCGCAG |
|
| Recombinant DNA reagent | Prmha3-Non-stop-2XFlag-2XHA-3FA | This Paper | Materials and methods Subsection Plasmids Quick change mutagenesis on Prmha3-Non-stop-2XFlag-2XHA WIRS3 phenylalanine to alanine F: CGCAAGATCTCCTCGGCCATTCAATTCCCCGTGGAGTTCGACATG WIRS3 phenylalanine to alanine R: CCACGGGGAATTGAATGGCCGAGGAGATCTTGCGATCGATCAGAGC |
|
| Recombinant DNA reagent | Prmha3-Non-stop-2XFlag-2XHA-WIRSFA | This Paper | Materials and methods Subsection Plasmids Quick change mutagenesis on Prmha3-Non-stop-2XFlag-2XHA all of the above primers sequentially |
|
| Recombinant DNA reagent | Prmha3-Non-stop-2XFlag-2XHA-C406A | This Paper | Materials and methods Subsection Plasmids Quick change mutagenesis on Prmha3-Non-stop-2XFlag-2XHA Non-stop C406A F: CTTAATCTGGGCGCCACTGCCTTCATGAACTGCATCGTC Non-stop C406A R: GACGATGCAGTTCATGAAGGCAGTGGCGCCCAGATTAAG |
|
| Recombinant DNA reagent | Prmha3-Non-stop-2XFlag-2XHA-C406S | This Paper | Materials and methods Subsection Plasmids Quick change mutagenesis on Prmha3-Non-stop-2XFlag-2XHA Non-stop C406S F: CTTAATCTGGGCGCCACTAGCTTCATGAACTGCATCGTC Non-stop C406S R: GACGATGCAGTTCATGAAGCTAGTGGCGCCCAGATTAAG |
|
| Sequence-based reagent | dsRNA Lacz | dsRNA-LacZ-R: GCTAATACGACTCACTATAGGCCAAACATGACCARGATTACGCCAAGCT dsRNA-LacZ-F: GCTAATACGACTCACTATAGGCCAAACGTCCCATTCGCCATTCAGGC | ||
| Sequence-based reagent | dsRNA not | http://www.flyrnai.org | DRSC11378 | Primer F: TAATACGACTCACTATAGGCGCAGGCTGAACTGTTTG Primer R: TAATACGACTCACTATAGGTCTATTCCGGCTCCCGTT |
| Sequence-based reagent | dsRNA not | http://www.flyrnai.org | BKN21994 | Primer F: TAATACGACTCACTATAGGACTTGACCCACGTGTCCTTC Primer R:TAATACGACTCACTATAGGATTGACCAGATCTTCACGGG |
| Sequence-based reagent | dsRNA Atxn7 | http://www.flyrnai.org | DRSC35628 | Primer F:TAATACGACTCACTATAGGCGACATGGAAAAGGTCATCA Primer R:TAATACGACTCACTATAGGGGAAACCTGCCTTCGTGTAA |
| Sequence-based reagent | dsRNA Atxn7 | http://www.flyrnai.org | DRSC23138 | Primer F: TAATACGACTCACTATAGGCTGTTAAGCTGGAGGCCAAGPrimer R: TAATACGACTCACTATAGGGCCCTCTTATTGCACCTCAG |
| Sequence-based reagent | Taqman Assay: not | Thermofisher | TaqManID: Dm01823071_g1 | |
| Sequence-based reagent | Taqman Assay: Atxn7 | Thermofisher | TaqManID: Dm01800874_g1 | |
| Sequence-based reagent | Taqman Assay: SCAR | Thermofisher | TaqManID: Dm01810606_g1 | |
| Sequence-based reagent | Taqman Assay: RPL32 | Thermofisher | TaqMan ID: Dm02151827_g1 | |
| Commercial assay or kit | high capacity cDNA reverse transcription kit | ThermoFisher | 4374966 | |
| Commercial assay or kit | TaqMan Universal PCR Master Mix | ThermoFisher | 4364340 | |
| Chemical compound, drug | MG132 | Sigma-Aldrich | C2211 | |
| Software, algorithm | ImageJ | https://imagej.nih.gov/ij/ |