Sp7 (Q9V3Z2, CG3066; synonyms MP2, PAE1) notes

2026-10-21 — Full re-review, independent of the ProtNLM2 predictions

Context: Sp7 was requested as the closest experimentally characterised fly relative
of the ProtNLM2 benchmark gene CG4793 (Q8IP30). This review treats Sp7 in its own
right. No ProtNLM2 output was consulted.

Deep research

just deep-research-falcon DROME Sp7 --fallback perplexity-lite failed: the Edison
(falcon) API returned "Failed to authenticate", and the perplexity-lite fallback timed
out after 600 s. No deep-research file was produced. The review is built from the
UniProt Swiss-Prot record and the cached primary literature listed below.

Evolutionary placement

• PANTHER 19.0 classification (.cache/panther/PTHR19.0_fruit_fly): Sp7 is in
  PTHR24256:SF537 "CLIP DOMAIN-CONTAINING SERINE PROTEASE-RELATED", whose only fly
  members are Sp7, c-SP10 (Q9VA87) and c-SP11 (Q9VA44). Easter (P13582) is in a
  different subfamily, PTHR24256:SF527. CG4793 (Q8IP30) is in PTHR24256:SF462.
  MP1 (A0A126GUP6) is in a different family altogether (PTHR24260:SF135).
• PAINT (interpro/panther/PTHR24256/PTHR24256-paint.tsv): node PTN001207268 carries
  IBD assertions for GO:0004252 serine-type endopeptidase activity, GO:0006508
  proteolysis and GO:0005576 extracellular region. Sp7 (FB:FBgn0037515) is itself a
  seed for the extracellular region IBD; easter (FBgn0000533) seeds all three.
  PAINT did not propagate melanization or any immune process from this node, so the
  immune role of Sp7 is a lineage-level/gene-level finding, not a family property.
• The catalytic triad is intact in the Sp7 sequence (checked directly from the UniProt
  sequence): His182, Asp244 and Ser341 (in the GDSGGP motif "SCDGDSGGPLM"). The chain
  is a clip-domain zymogen: signal peptide 1-27, clip domain 30-84, activation
  propeptide 28-136, and catalytic chain from Ile137 (IVYNGN...). This contrasts
  with CG4793, a serine protease homolog whose catalytic Ser is replaced by Gly
  (family review PTHR24256-review.yaml).

Consequence for transfer: Sp7's experimentally established functions (PPO1
cleavage, melanization) belong to a catalytically active clip-domain SP in a
different subfamily from CG4793. Only the family-level IBD properties (secreted,
S1A protease fold) are shared at the PTN001207268 level, and even the catalytic
activity does not transfer to SPHs that lack the triad.

Experimental findings in D. melanogaster

• Castillejo-Lopez & Hacker 2005 (PMID:16256951, abstract only): Sp7 RNAi impairs
  melanization upon injury; Sp7 is required for PO activation; expressed in blood
  cells (crystal cells); transcription up-regulated by injury and by activated Toll,
  but Toll/IMD mutants do not abolish expression.
  PMID:16256951
• Leclerc et al. 2006 (PMID:16322759, full text): PAE1 (=Sp7) mutants fail to
  activate PO in the haemolymph after microbial challenge, yet survive infections
  as well as wild type. PMID:16322759
• Tang et al. 2006 (PMID:16861233, abstract only): MP1 and MP2 act in a Spn27A-regulated
  melanization cascade; MP2 mainly in fungal infection; genetic data placed MP2
  upstream of MP1.
• Ayres & Schneider 2008 (PMID:19071960, full text): CG3066 mutants alter both
  resistance and tolerance in a microbe-dependent way; reduced survival after
  S. aureus, L. monocytogenes, S. typhimurium; longer survival with S. pneumoniae.
• An et al. 2013 (PMID:24260243, full text): recombinant Factor Xa-activatable
  proMP2 has amidase activity on IEAR-pNA when activated, cleaves recombinant and
  native PPO1, and in vivo overexpression/RNAi increases/decreases PPO1 cleavage;
  Spn27A forms SDS-stable complexes with MP2 and inhibits it. This is the direct
  biochemical evidence for serine-type endopeptidase activity.
• Dudzic et al. 2019 (PMID:31018123, abstract only): with compound mutants, Hayan is
  required for wound blackening, whereas Sp7 controls an alternate melanization that
  clears S. aureus; Sp7 is regulated by the Toll-activating SPs ModSP and Grass.
• Bidla et al. 2007 (PMID:17356067, abstract only): crystal cell rupture and PPO
  release; the abstract does not mention Sp7, the IMP annotation must come from
  the full text (deferred to the FlyBase curator).
• Cao & Jiang 2018 (PMID:30367934, full text) note the conflict over whether MP2 is
  the terminal PAP (in vitro) or acts upstream of MP1 (genetics).

Decisions summary

• Serine-type endopeptidase activity: ACCEPT (all rows), supported by An 2013
  biochemistry plus intact triad.
• Proteolysis: ACCEPT (true, supported by direct PPO1 cleavage); broad but consistent.
• Extracellular region: ACCEPT (signal peptide, hemolymph protease).
• Melanization defense response: ACCEPT (core process).
• Positive regulation of melanization defense response: ACCEPT. Sp7 activates PPO1 (or
  proMP1) and so positively regulates the melanization effector; FlyBase curators used
  this term from two independent papers.
• Defense response to Gram-positive bacterium: KEEP_AS_NON_CORE. Sp7-dependent
  melanization clears S. aureus, so it acts in the defense, but the term is a broad
  organismal context of the core melanization role.
• No NEW terms. GO:0031638 zymogen activation would describe PPO1 cleavage
  (participation test passes: Sp7 does the cleaving) but the comparator check was
  not run, so it is raised as a question instead.