PXD041843

PXD041843 is an original dataset announced via ProteomeXchange.

Title	Viral pIC-pocketing: RSV sequestration of eIF4F Initiation Complexes into bi-phasic biomolecular condensates
Description	Orthopneumoviruses characteristically form membrane-less cytoplasmic inclusion bodies (IBs) wherein RNA replication and transcription occur. Herein, we report a strategy whereby the orthopneumoviruses sequester various components of the eiF4F Initiation Complex machinery into viral IBs to facilitate translation of their own mRNAs; pIC-pocketing. Mass spectrometry analysis of sub-cellular fractions from RSV-infected cells identified significant modification of the cellular translation machinery; however; interestingly, ribopuromycylation assays showed no changes to global levels of translation. Electron micrographs of RSV-infected cells revealed bi-phasic organisation of IBs; specifically, spherical “droplets” nested within the larger inclusion. Using correlative light and electron microscopy (CLEM), combined with fluorescence in situ hybridisation (FISH), we showed that the observed bi-phasic morphology represents functional compartmentalisation of the IB and that these domains are synonymous with the previously reported inclusion body associated granules (IBAGs). Detailed analysis demonstrated that IBAGs concentrate nascent viral mRNA, the viral M2-1 protein as well as many components of the eIF4F complex, involved in translation initiation. Interestingly, although ribopuromycylation-based imaging indicates the majority of viral mRNA translation likely occurs in the cytoplasm, there was some evidence for intra-IBAG translation, consistent with the likely presence of ribosomes in a subset of IBAGs imaged by electron microscopy. The mechanistic basis for this pathway was subsequently determined; the viral M2-1 protein interacting with eukaryotic translation initiation factor 4G (eIF4G) to facilitate its transport between the cytoplasm and the separate phases of the viral IB. In summary, our data shows that IBs function to spatially regulate early steps in viral translation within a highly selective biphasic liquid organelle.
HostingRepository	PRIDE
AnnounceDate	2024-02-02
AnnouncementXML	Submission_2024-02-02_05:51:16.275.xml
DigitalObjectIdentifier	https://dx.doi.org/10.6019/PXD041843
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Supported dataset by repository
PrimarySubmitter	Stuart Armstrong
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	monohydroxylated residue; iodoacetamide derivatized residue
Instrument	Q Exactive HF

Revision	Datetime	Status	ChangeLog Entry
0	2023-04-27 00:17:46	ID requested
⏵ 1	2024-02-02 05:51:16	announced

10.6019/PXD041843;

submitter keyword: Respiratory syncytial virus,eIF4F complex, inclusion bodies

Dalan Bailey
contact affiliation	The Pirbright Institute, Woking, Surrey, GU24 0NF, UK
contact email	dalan.bailey@pirbright.ac.uk
lab head
Stuart Armstrong
contact affiliation	Infection Biology
contact email	sarmstro@liv.ac.uk
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD041843
     1. Label: PRIDE project
     2. Name: Viral pIC-pocketing: RSV sequestration of eIF4F Initiation Complexes into bi-phasic biomolecular condensates