csy1 / Cas8f (Q02ML9) — Pseudomonas aeruginosa UCBPP-PA14 — curation notes

Deep research was unavailable in this environment (no provider API keys), so this file
is my own literature journal. Every assertion carries an inline PMID plus a verbatim
quote from the cached record in publications/.

What this protein is

PA14_33330, 434 aa, "CRISPR-associated protein Csy1" = Cas8f, the large subunit of
the type I-F surveillance complex. It occupies the PAM-proximal tail:
PMID:32170016, in a complex of defined composition
PMID:32170016. The original biochemistry on the PA14 proteins:
PMID:21536913. GOA's two
GO:0005515 rows are the IntAct Csy1–Csy2 pair (Q02ML9–Q02MM0, NbExp=6) from that work
and from PMID:26416740.

The structures behind the claims below are of the P. aeruginosa complex:
PMID:28985564.

What Cas8f actually does

This is the structural-subunit problem, but Cas8f is not a passive scaffold — it has
three assignable jobs, all resolved in cryo-EM of the PA14 complex.

1. Binds the PAM-containing target DNA duplex.
PMID:28985564, with the pocket dominated by Cas8f side chains:
PMID:28985564.

2. Splits the duplex and reads the PAM. A Cas8f loop wedges into the fork:
PMID:28985564 and
PMID:28985564. Independently
confirmed as functionally required:
PMID:32170016.

3. Anchors the crRNA 5' handle, sequence-specifically.
PMID:32170016, and
PMID:32170016.

4. Recruits Cas3.
PMID:32170016. This is the function AcrIF3 neutralises from the
Cas3 side (PMID:27455460, read with PMID:26416740).

In vivo

Csy1 is required for the phage-dependent phenotype but, unlike Csy2 and Csy4, it is not
strictly required for crRNA accumulation:
PMID:21398535. UniProt records a reduction rather than a loss
("Decreased production of crRNA"). This asymmetry is why I propose GO:0043571 for
csy2 but not for csy1.

The complete Csy complex is required for adaptation as well as interference:
PMID:26586803 — but the gene-level alleles tested in
that study were csy3 and csy4, not csy1, so I do not transfer an IMP to Csy1.

Curation decisions

• Both GO:0005515 protein binding rows (IPI, partner Csy2) → MODIFY to
  GO:0005198 structural molecule activity. protein binding says nothing about
  function; the Csy1–Csy2 IPI samples a contact internal to the complex, where Cas8f
  and Cas5f jointly clamp the crRNA 5' handle and their hook and thumb form two of the
  three walls of the target-duplex pocket. The contribution is structural, and mutual.

Aspect correction (follow-up). My first pass proposed
part_of GO:0032991 protein-containing complex as the replacement, reasoning from the
E. coli comparator where all five Cascade subunits carry exactly that. That was wrong
in form: these GOA rows carry qualifier: enables, so they are molecular-function
rows, and a gene product cannot enable a cellular component. However
well-supported complex membership is, it cannot be expressed by swapping a CC term
into an enables MF row. The E. coli reviews were not making this mistake — their
GOA already contained separate part_of GO:0032991 rows, which they MODIFY to
GO:1990904; their enables GO:0005515 rows went to GO:0005198 instead.
Reconciled to the same convention:

• MODIFY the enables rows to GO:0005198 structural molecule activity ("the
  action of a molecule that contributes to the structural integrity of a complex").
  Aspect-correct, and none of its 22 children fits a non-ribosomal ribonucleoprotein,
  so the parent is the right level. Each row is backed by the verbatim structural
  quote naming the contact this subunit actually makes.
• Carry complex membership on its own NEW part_of GO:1990904 ribonucleoprotein complex row. PA14 has no part_of complex row for any Csy subunit, unlike all
  five E. coli Cascade subunits, so this is a real missing annotation rather than a
  reformulation. GO:1990904 rather than the bare GO:0032991 root because the
  60-nt crRNA is an integral component, not a ligand.
• core_functions[].in_complex → GO:1990904 for the same reason.
• Added a proposed_new_terms entry for a "CRISPR RNA-guided surveillance
  complex" CC term under GO:1990904, identical in substance to the one the five
  E. coli Cascade reviews carry, so the two systems request one term rather than two.
  GO's entire CRISPR vocabulary is five BP terms (GO:0099048, GO:0043571,
  GO:0098672 and the obsolete GO:0110132/GO:0110133); ComplexPortal models the
  type I-E complex as CPX-1005, GO does not.

Gaps identified (missing annotations)

• GO:0003677 DNA binding (NEW, IDA). Comparator: casA (Cas8e, Q46901) carries
  GO:0003677 by IDA from PMID:25123481. Cas8f's hook and K-wedge are the main
  protein contacts to the target duplex (quotes above).
• GO:0003723 RNA binding (NEW, IDA). Comparator: casA carries GO:0003723 by IDA
  twice. Here it is the sequence-specific 5'-handle recognition.
• GO:0099048 CRISPR-cas system (NEW, IDA). Comparator: all five E. coli Cascade
  subunits carry it by IDA (PMID:18703739); no PA14 Csy subunit does. Cas8f performs
  part of the "target interference" stage named in the term's definition — PAM reading
  and duplex splitting are steps of the process, not merely a requirement for it.
  Checked via QuickGO that GO:0099048 is not an ancestor or descendant of
  GO:0043571 or GO:0051607.

Not proposed, and why

• No molecular-function term for crRNA-guided, PAM-licensed target recognition
  exists. That is the activity the assembled Csy complex has, and Cas8f is the subunit
  that licenses it. modules/crispr_cas_adaptive_immunity.yaml records this as an open
  ONTOLOGY gap and falls back to GO:0003690 for the complex. I have not substituted
  any existing id for the missing concept; I annotate the component activities that are
  real (GO:0003677, GO:0003723) and raise the gap in suggested_questions.
• GO:0043571 maintenance of CRISPR repeat elements: declined, per the PMID:21398535
  asymmetry above.
• GO:0051607 defense response to virus: declined. No csy1 allele has been assayed
  for phage resistance, and the PA14 locus-level evidence is contested (see
  cas3-notes.md).
• A Cas3-recruitment molecular function: "proposed to interact with" is a structural
  inference in PMID:32170016, and the direct structure is of the Cas3–AcrF3 pair, not
  Cas8f–Cas3. Raised as a question rather than annotated.