Journal: Current biology : CB

Article Title: Animal septins contain functional transmembrane domains.

doi: 10.1016/j.cub.2025.03.004

Figure Lengend Snippet: Figure 3. Animal septins are predicted to contain transmembrane domains (TMDs) (A) Cladogram of all septin sequences deposited on UniProt (magenta, sequences containing TMDs). Rim colors: organismal phyla (dark blue, Ascomycota; brown, Basidiomycota; fuchsia, Mucoromycota; purple, Chytridiomycota; lime green, Viridiplantae; yellow, Chordata; teal, Arthropoda; light green, Nematoda; blue, Platyhelminthes; light pink, Mollusca; hot pink, Cnideria). (B) msfGFP::CsSEPT10C (green) expressed in U2OS cells and colocalized with translocase of outer mitochondrial membrane 20 (TOMM20; magenta). Images shown are a single plane, inverted contrast, and scaled to display minimum and maximum pixel intensities. Merged image is not inverted contrast. Dashed box (white) indicates region used for 23 inset (not inverted). Scale bar, 10 mm. (C) msfGFP::CsSEPT10C (green) co-expressed with HaloTag::Sec61b (ER; magenta) in U2OS cells. Images shown are a single plane, inverted contrast, and scaled to display minimum and maximum pixel intensities. Merged image is not inverted contrast. Dashed box (white) indicates region used for 23 inset (not inverted). Scale bar, 10 mm. (D) (Left) Neighbor-joining tree of evolutionary relationships. (Right) Schematic of CsSEPT10 and HsSEPT10 protein architecture. Colors as in Figure 1C. (E) Localization of msfGFP::HsSEPT10 and the chimera msfGFP::HsSEPT10DC-CsSEPT10C. Scale bar, 10 mm. (F) Colocalization of the chimera msfGFP::HsSEPT10DC-CsSEPT10C (green) and mApple::TOMM20 (magenta). Images shown are a single plane, inverted contrast, and scaled to display minimum and maximum pixel intensities. Merged image is not inverted contrast. Dashed box (white) indicates region used for 23 inset (not inverted). Scale bar, 10 mm. See also Figure S3.

Article Snippet: This work N/A C. elegans strain MDX152; genotype: knuSi934 [pNU3400 (AMAD01- unc-61p:: wrmScarlet::UNC-61(B/C):: unc-61u, unc-119(+))] II; unc-119(ed)III; unc-61a (mon26 [unc-61a L7_V29del]) V. This work N/A Oligonucleotides Primers for mammalian transgenes, See Table S1 This work N/A Primers to confirm MosSCI animals, See Table S2 This work N/A crRNA, ssDNA donor, confirmatory primers for CRISPR-edited C. elegans, See Table S3 This work N/A Recombinant DNA cDNA: HsSEPT10_i1 Gift from Andrew Wilde (University of Toronto) N/A Gene fragment: msfGFP::CsSEPT10C Eurofins Genomics N/A Gene fragment: HsSEPT10_i1DC-CsSEPT10C Eurofins Genomics N/A Plasmid: Double UP Halotag to mScarlet Gift from Erik Dent (University of Wisconsin-Madison) Addgene plasmid #125138 Plasmid: mScarlet3 Gift from Dorus Gadella (University of Amsterdam) Addgene plasmid #189753 Plasmid: UNC-61a TMD1-48:HaloTag This work pJAP9 Plasmid: msfGFP::HsSEPT10_i1 This work pM321 Plasmid: msfGFP::CsSEPT10C This work pM324 Plasmid: msfGFP::HsSEPT10_i1DC-CsSEPT10C This work pM325 Plasmid: mScarlet3-CAAX This work pM300 Plasmid: UNC-61a TMD1-48::msfGFP This work pJAP11 Plasmid: TOMM20::mApple Gift from Michael Davidson (Florida State University) Addgene plasmid #54955 Plasmid: GalT::mCherry Gift from Michael Davidson (Florida State University) Addgene plasmid #55052 Plasmid: HaloTag::Sec61b Gift from Christopher Obara (UCSD) N/A Software and algorithms UniProt Uniprot Consortium29 https://uniport.org Interactive Tree of Life Letunic and Bork57 https://itol.embl.de (Continued on next page) ll Current Biology 35, 1910–1917.e1–e5, April 21, 2025 e2 Report

Techniques: Membrane