DOI: 10.5061/dryad.p5hqbzkw0
title: Structure and replication cycle of a virus infecting climate-modulating alga Emiliania huxleyi

description: The globally distributed marine alga Emiliania huxleyi produces reflective
 calcite disks (coccoliths) that increase the albedo of ocean water and
 thus reduce the heat absorption in the ocean, which cools the Earth’s
 climate. The population density of E. huxleyi is restricted by
 Nucleocytoviricota viruses, including E. huxleyi virus 201 (EhV-201).
 Despite the impact of E. huxleyi viruses on the climate, there is limited
 information about their structure and replication. Here we show that the
 dsDNA genome inside the EhV-201 virion is protected by an inner membrane,
 capsid, and outer membrane decorated with numerous transmembrane proteins.
 The virions are prone to deformation, and parts of their capsids deviate
 from the icosahedral arrangement. EhV-201 virions infect E. huxleyi by
 using their fivefold vertex to bind to a host cell and fuse the virus’s
 inner membrane with the plasma membrane. Whereas the replication of
 EhV-201 probably occurs in the nucleus, virions assemble in the cytoplasm
 at the surface of endoplasmic reticulum-derived membrane segments. Genome
 packaging initiates synchronously with the capsid assembly and completes
 through an aperture in the forming capsid. Upon the completion of genome
 packaging, the capsids change conformation, which enables them to acquire
 an outer membrane by budding into intracellular vesicles. EhV-201
 infection induces a loss of surface protective layers from E. huxleyi
 cells, which allows the continuous release of virions by exocytosis. Our
 results provide insight into how EhVs bypass the surface protective layers
 of E. huxleyi and exploit the organelles of an infected cell for progeny
 assembly.
description: # Structure and replication cycle of a virus infecting climate-modulating
 alga Emiliania huxleyi 1\)
 *Emiliania huxleyi* virus 201 life cycle visualization *in situ*: * a)
 Cryo-electron tomography of lamellas of *Emiliania huxleyi* noncalcifying
 2090 alga infected by *E. huxleyi* virus 201 (EhV-201) and control
 uninfected samples. * b) Segmentation of two selected cryo-tomographic
 lamellae volumes (infected and control *E. huxleyi* cells). * c) Block
 face imaging data of high-pressure frozen and resin-embedded samples of
 *E. huxleyi* infected by EhV-201. 2\) Scripts used during the
 reconstruction of EhV-201 by subtomogram averaging: * a) TomoStarPy
 scripts available on GitHub (22 February 2024 clone):
 * b) Script used for
 alignment of EhV-201 virion pentamer vertices during generation of virion
 composite map 3\) Super-resolution fluorescence data of EhV-201 attachment
 to the *E. huxleyi* cells. 4\) Proteomic analysis of EhV-201 virion - full
 report ## Description of the data and file structure **Note:** All the
 data are compressed as fileName.tar.bzip2. To extract them, you can use
 Linux terminal (tar -xjf filename.tar.bzip2) or Windows program 7-Zip or
 WinRAR (extract gradually: fileName.tar.bzip2 &gt; extract here &gt;
 fileName.tar &gt; extract here). #### 1.a) *E. huxleyi* cryo-lamellas
 tomography data: * 1a_lamellas_ehux_control_ehux:
 xxxxxx_ehux2090_ctrl_f2medium and \_100kDa_lysate (control *E. huxleyi*
 samples treated with F/2-Si medium or by 100 kDa viral lysate flow-through
 (more details in the article)). * 1a_lamellas_ehux_infected_48h_dataset_1
 and \_2-4 (infected *E. huxleyi* samples at late stage of EhV-201
 infection at 48 hours post-infection) * 1a_lamellas_ehux_infected_10min
 and \_30min (infected *E. huxleyi* samples at early stage of EhV-201
 infection at 10 or 30 minutes post-infection) *
 1a_lamellas_ehux_infected_coccoliths (infected *E. huxleyi* samples which
 spontaneously reanimate the coccolith production at early stage of EhV-201
 infection at 30 or 60 minutes post-infection) To visualize the
 reconstructed tomographic volume use 3dmod from IMOD package. Available
 for Windows (Open file &gt; tomoX_full_rec.mrc) and Linux (3dmod -Y
 tomoX_full_rec.mrc) Intermediate reconstruction image files removed. To
 regenerate them use the eTomo GUI from IMOD package workflow (File &gt;
 Open &gt; tomoX.edf) --- #### 1.b) Segmentation of *E. huxleyi* lamellae
 volumes i) Original segmentation data:
 "origData_dragonflySession" * control_cell - noninfected *E.
 huxleyi* cell with a segmented thin slice of the volume. * infected_cell -
 infected *E. huxleyi* cell by EhV-201 at a late stage of infection. Due to
 the PC memory issues individual materials are in separate sessions
 (_cellEnvelope; \_innerVesicle; \_membranes; \_viruses). Data contains
 Tomograms (tomoX_rec_noBytes_noFilter.mrc); and Segmentations
 (fileName.ORSSession). To open the segmentation session files use
 Dragonfly ORS software (Windows, Linux). ii) Segmentation movies
 generation: "movie_generation" To visualize the segmented
 volumes use the ChimeraX software (Windows, Linux) command line:
 "open movie_segmentation_xxx_preparation.cxc" To generate the
 movie, run in the same session: "open
 movie_segmentation_xxx_movie.cxc" --- #### 1.c) Block face imaging
 data To open the stitched volumes use ImageJ (Fiji) (Windows, Linux). ---
 #### 2.a) TomoStarPy scripts master branch clone: Find the installation
 instructions and individual scripts description in: README.md inside the
 "tomostarpy" folder. Alternatively you can invoke the help for
 each individual script in terminal by entering (python3 scriptName.py).
 --- #### 2.b) Script for virion pentamer orientation: The script requires
 Cyton, Cuda-python, Numpy. Building: build_libs.bsh; Running: set
 parameters in run.py (help in the source code) and execute it. \ This is
 an single-use script without any possibility of changing the parameters
 too much. --- #### 3) *E. huxleyi* fluorescence data from super-resolution
 Elyra 7 microscope. * Medium F/2-Si control (220708_F2CTRL_xxx.czi) *
 Virus in F/2-Si medium control (220708_virusCTRL_xxx.czi) * Uninfected *E.
 huxleyi* alga control (220708_algaCTRL_xxx.czi) * Infected *E. huxleyi* by
 EhV-201 at 30 min post infection using various multiplicity of infection
 (220708_MOI10 and \_MOI100 and \_MOI1000_xxx.czi) * Fluorescent beads used
 for channel alignment (220708_beads_xxx.czi). Original 9-phase data
 Z-stack in a leap mode (no identifier) and SIM^2 reconstructed Z-stack
 data (_SIM). To visualize the reconstructed Z-stack data (_SIM), please
 use ZEN (Blue or Black versions) software (Windows), or ImageJ (Fiji)
 (both available for Windows, Linux). --- #### 4) Proteomic data analysis
 Open the file in MS Excel. Individual sheets in the Excel file are named:
 \ i) Gel image (image of the gel with depicted protein bands used for MS
 analysis).\ ii) 1 - 27 (summary of the detected proteins identified in the
 corresponding protein bands. Individual columns correspond to: Protein
 accession number; Protein description; Molecular weight (MW); Calculated
 isoelectric point (calc. pI); Spectral abundance factor (SAF), Normalized
 abundance factor (NSAF), Sequence coverage; Sum of proteins; Sum of
 peptides; and Sum of unique peptides. More information can be found in the
 notes of the column's headings, depicted by a red triangle in the
 cell upper right corner). \ iii) 1 - 27_peptides (contains information
 about all the peptides detected for each protein. Lists of peptides are in
 the hidden lines depicted by a plus sign on the left from the line number.
 Click the plus sign to expand them). ## Sharing/Access information Links
 to other publicly accessible locations of the data. Code availability: *