PXD042090-1

PXD042090 is an original dataset announced via ProteomeXchange.

Title	Cas1-Cas2 and DnaK physically and functionally interact to modulate CRISPR Adaptation
Description	Prokaryotic Cas1-Cas2 protein complexes generate adaptive immunity to mobile genetic elements (MGEs), by capture and integration of MGE DNA in to CRISPR sites. De novo immunity relies on Cas1-Cas2 targeting MGE DNA, without the aid of pre-existing immunity complexes, through mechanisms of ‘naive adaptation’ that are not clear. Using E. coli we show that the chaperone DnaK inhibits Cas1-Cas2 from DNA binding and integration, and that DnaK expression prevents naïve adaptation from chromosomal self-targeting. We show that that inhibition of naïve adaptation is reversible  by eliminating DnaK from cells, by mutation of the DnaK substrate binding domain, and by expression of an MGE (phage )protein. We show that a fluorescently labelled Cas1 fusion can be visualised in living cells. Formation of foci depends on active DNA replication, and that the number of foci per cell is much increased in cells lacking DnaK. We discuss a model in which DnaK provides a mechanism for restraining naïve adaptation from self-targeting. This restraint is released once MGE DNA is present in the cell.
HostingRepository	PRIDE
AnnounceDate	2023-05-18
AnnouncementXML	Submission_2023-05-18_04:51:47.311.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	TomKillelea
SpeciesList	scientific name: Escherichia coli; NCBI TaxID: 562;
ModificationList	biotinylated residue
Instrument	LTQ

Revision	Datetime	Status	ChangeLog Entry
0	2023-05-09 09:43:25	ID requested
⏵ 1	2023-05-18 04:51:47	announced
2	2023-11-14 09:00:22	announced	2023-11-14: Updated project metadata.

submitter keyword: CRISPR, Cas1-Cas2, Adaptaion, DnaK

Edward L.Bolt
contact affiliation	School of Life Sciences, University of Nottingham
contact email	ed.bolt@nottingham.ac.uk
lab head
TomKillelea
contact affiliation	University of Nottingham
contact email	tom.killelea@nottingham.ac.uk
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD042090
     1. Label: PRIDE project
     2. Name: Cas1-Cas2 and DnaK physically and functionally interact to modulate CRISPR Adaptation