PXD072625-1

PXD072625 is an original dataset announced via ProteomeXchange.

Title	Single-molecule imaging reveals RNA Polymerase II dynamics and TAF1-dependent promoter-proximal pause release
Description	Transcription initiation is a highly regulated process that determines gene expression outcomes1,2, yet the dynamics and efficiency of individual initiation events in living cells remain poorly understood. Here we combine endogenous tagging of human RNA polymerase II (Pol II) and the TFIID subunit TAF1 with dual-color live-cell single-molecule imaging to quantitatively map Pol II behavior during transcription initiation and early elongation. Using GRID (Genuine Rate Identification) analysis, we resolved four kinetic populations of chromatin-bound Pol II and find that more than 94% of promoter-bound molecules dissociate within tens of seconds, indicating globally inefficient initiation. Kinetic partitioning of Pol II dwell times into promoter-proximal pausing and gene body elongation reveals a global pausing index that is selectively increased by CDK9 inhibition. Single-cell analysis uncovers substantial heterogeneity in initiation efficiency and pausing that is not explained by cell-cycle state. Spatial heatmap analysis identifies recurrent Pol II ‘hotspots’, nuclear regions enriched for repeated long-lived Pol II binding events, whose abundance and architecture are selectively perturbed by inhibitors that trap Pol II at promoters or disrupt TFIID. Finally, acute depletion of TAF1 leaves initiation efficiency unchanged but significantly increases the pausing index and reduces elongation efficiency, consistent with impaired pause release. Together, these findings establish a general framework to quantify transcription efficiency in vivo and support a model in which TAF1, and by extension TFIID, promotes pause release and productive elongation.
HostingRepository	PRIDE
AnnounceDate	2026-06-03
AnnouncementXML	Submission_2026-06-03_09:33:47.785.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Simone Sidoli
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: NEWT:9606;
ModificationList	No PTMs are included in the dataset
Instrument	timsTOF HT

Revision	Datetime	Status	ChangeLog Entry
0	2026-01-04 16:19:12	ID requested
⏵ 1	2026-06-03 09:33:48	announced

Dataset with its publication pending

submitter keyword: single particle tracking, TFIID, transcription efficiency,RNA Polymerase II, proximal pausing

Simone Sidoli
contact affiliation	Department of Biochemistry, Albert Einstein College of Medicine, Bronx, NY 10461, United States
contact email	simone.sidoli@einsteinmed.edu
lab head
Simone Sidoli
contact affiliation	Albert Einstein College of Medicine
contact email	simone.sidoli@gmail.com
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD072625

PRIDE project URI

• PRIDE
  1. PXD072625
     1. Label: PRIDE project
     2. Name: Single-molecule imaging reveals RNA Polymerase II dynamics and TAF1-dependent promoter-proximal pause release