Effect of Lenacapavir and PF74 on HIV-1 capsid morphology: EM images

Lenacapavir (GS-6207; LEN) is a potent HIV-1 capsid inhibitor approved for treating multidrug-resistant infection. LEN binds to a hydrophobic pocket between neighboring capsid (CA) proteins in hexamers and stabilizes the capsid lattice, but its effect on HIV-1 capsids is not fully understood. Here, we labeled HIV-1 capsids with green fluorescent protein fused to CA (GFP-CA) or a fluid-phase GFP content marker (cmGFP) to assess LEN's impact on HIV-1 capsids. HIV-1 cores labeled with GFP-CA, but not cmGFP, could be immunostained with an anti-GFP antibody and were less sensitive to the capsid-binding host restriction factor MX2, demonstrating that GFP-CA is incorporated into the capsid lattice and is a marker for capsid lattice stability whereas cmGFP is an indicator of core integrity. LEN treatment of isolated HIV-1 cores resulted in a dose-dependent loss of cmGFP signal while preserving the GFP-CA signal, indicating that LEN disrupts core integrity but stabilizes the capsid lattice. In c..., Negative stain electron microscopy. Mature cores (5 µL of fraction 19) were incubated with 5 µL HS+IP6 buffer containing LEN (1, 10, or 100 nM final concentration) or PF74 (10 µM final concentration) in a 0.2 mL PCR tube for 20 min at 37ºC. HIV core specimen (5 µL) was applied to carbon film coated 200 or 400 mesh copper grids (Electron Microscopy Sciences). Grids were rendered hydrophilic by glow discharge prior to usage. After a 1 min incubation, the unbound specimen was wicked off with Whatman filter paper and stained with 2 droplets of 2% uranyl acetate for 1 min. After wicking off excess stain, grids were air dried and examined using a Hitachi 7800 transmission electron microscope at 80kV and imaged using a 5056X2960 pixel CCD camera (NanoSprint 15B, Advanced Microscopy Techniques). The observed capsid morphologies were categorized into four types: apparently intact, broken at the narrow end, broken (other), and aberrant. Samples were blinded and the number and percentage of capsid..., , # Effect of Lenacapavir and PF74 on HIV-1 capsid morphology: EM images

[https://doi.org/10.5061/dryad.70rxwdc7z](https://doi.org/10.5061/dryad.70rxwdc7z)

## Description of the data and file structure

Images of HIV-1 capsids were captured using electron microscopy. The raw images, with identifying information removed, were organized into TIFF stacks. The file \"EM_Raw_Images.xlsx\" includes details such as the sample name, image stack name, image number, and capsid morphology for each detected capsid. In total, 1,819 capsids were analyzed from two experiments.

### Files and variables

#### File: EM\_Raw\_Images.xlsx

**Description:** The TIFF stack name and image number for each analyzed HIV-1 capsid are provided here.

#### File: Exp#\_Stack#.tif

**Description:** raw image stacks. See Excel file for metadata

## Code/software

ImageJ can be used to view the images. ImageJ can be downloaded at: [https://imagej.net/ij/](https://imagej.net/ij/)
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