Journal: bioRxiv

Article Title: The Antiviral and Cancer Genomic DNA Deaminase APOBEC3H Is Regulated by a RNA-Mediated Dimerization Mechanism

doi: 10.1101/203000

Figure Lengend Snippet: ( A and B ) Comparison of the DNA deaminase activities of untagged wild-type (WT) human A3H or the indicated amino acid substitution mutants in 293T extracts in the absence and presence of RNase (S, substrate; P, product). Corresponding immunoblots show levels of A3H in cell lysates using a murine mAb (P1D8) or a rabbit pAb (Novus). β-actin is a loading control.

Article Snippet: A fraction of each extract was separated by 12% SDS-PAGE and transferred to low-fluorescence PVDF overnight at 20 V. Membranes were blocked with 4% milk in PBST then incubated with mouse anti-A3H mAb P1D8 ( ) or rabbit anti-β-actin pAb (Cell Signaling: 13E5), at dilutions of 1:1,000 or 1:10,000 respectively.

Techniques: Western Blot

Journal: bioRxiv

Article Title: The Antiviral and Cancer Genomic DNA Deaminase APOBEC3H Is Regulated by a RNA-Mediated Dimerization Mechanism

doi: 10.1101/203000

Figure Lengend Snippet: ( A ) Representative images of A3H, A3H-R175E/R176E, A3B, and A3G in 293T cells. The no APOBEC control shows low background fluorescence. DAPI staining defines the nuclear compartment (10 μm scale bar). ( B ) Quantification of the subcellular localization of the indicated constructs (n =25 cells with average +/− SD; N, nuclear; WC, whole cell; C, cytoplasmic). ( C ) Single-cycle infectivity data for Vif-null HIV-1 LAI produced in the presence of 200 ng of the indicated untagged, human A3H expression constructs (WT, wild-type; average +/− SD of 3 technical replicates). Corresponding immunoblots are shown below for proteins in virus-like particles (VLPs) and virus-producing 293T cells (A3H detection with mouse mAb P1D8 and rabbit pAb Novus; anti-p24 and anti-tubulin are loading controls for VLPs and cells, respectively).

Article Snippet: A fraction of each extract was separated by 12% SDS-PAGE and transferred to low-fluorescence PVDF overnight at 20 V. Membranes were blocked with 4% milk in PBST then incubated with mouse anti-A3H mAb P1D8 ( ) or rabbit anti-β-actin pAb (Cell Signaling: 13E5), at dilutions of 1:1,000 or 1:10,000 respectively.

Techniques: Fluorescence, Staining, Construct, Infection, Produced, Expressing, Western Blot