Journal: mBio

Article Title: Identification and Characterization of a Candidate Wolbachia pipientis Type IV Effector That Interacts with the Actin Cytoskeleton

doi: 10.1128/mBio.00622-16

Figure Lengend Snippet: The α-synuclein domain-containing protein is conserved across the Wolbachia genus. (A) Alignment of the synuclein domain from WD0830 and Wolbachia homologs compared to mammalian alpha-synuclein (mouse). Conserved residues are shown at the top of the alignment. (B) Conservation (the percent protein identity) and length of the WD0830 homolog across the Wolbachia genus. Area and percent similarity are indicated by the colored line across the WD0830 open reading frame. (C) Phylogeny of these same open reading frames generated from codon alignments (ClustalW) edited by eye and then used as input to RAxML (GTR+γ) recovered major Wolbachia clades.

Article Snippet: Either WD0830 (at a 40-ng/ml final concentration), the actin binding protein alpha-actinin (Cytoskeleton, Inc.; used as a positive control for F-actin binding and sedimentation), BSA, or nothing additional (negative controls) was added to individual actin samples.

Techniques: Generated

Journal: mBio

Article Title: Identification and Characterization of a Candidate Wolbachia pipientis Type IV Effector That Interacts with the Actin Cytoskeleton

doi: 10.1128/mBio.00622-16

Figure Lengend Snippet: WD0830 cosediments with and bundles actin. (A) Purified WD0830 and alpha-actinin (positive-control actin binding protein) were incubated with polymerized rabbit skeletal actin and subjected to centrifugation at 150,000 × g , fractionated by SDS-PAGE, and silver stained to visualize proteins in the supernatant (S) and the pellet (P). (B) To identify actin bundling activity, polymerized rabbit skeletal actin was incubated with or without WD0830 as well as with or without BSA (as a negative control) and subjected to low-speed (14,000 × g) centrifugation before high-speed (150,000 × g) centrifugation. LSP, low-speed pellet; HSS, high-speed supernatant; HSP, high-speed pellet. (C) To visualize actin bundling, polymerized rabbit skeletal actin was incubated with either WD0830 or BSA and then directly stained with Acti-stain 555, mounted on a slide, and visualized by fluorescence microscopy. Bar, 100 µm. All images were taken at the same magnification.

Article Snippet: Either WD0830 (at a 40-ng/ml final concentration), the actin binding protein alpha-actinin (Cytoskeleton, Inc.; used as a positive control for F-actin binding and sedimentation), BSA, or nothing additional (negative controls) was added to individual actin samples.

Techniques: Purification, Positive Control, Binding Assay, Incubation, Centrifugation, SDS Page, Staining, Activity Assay, Negative Control, Fluorescence, Microscopy