Family Curator Notes

Annotation inferences using phylogenetic treesThe goal of the GO Reference Genome Project, described in PMID 19578431, is to provide accurate, complete and consistent GO annotations for all genes in twelve model organism genomes.To this end, GO curators are annotating evolutionary trees from the PANTHER database with GO terms describing molecular function, biological process and cellular component. GO terms based on experimental data from the scientific literature are used to annotate ancestral genes in the phylogenetic tree by sequence similarity (ISS), and unannotated descendants of these ancestral genes are inferred to have inherited these same GO annotations by descent. The annotations are done using a tool called PAINT (Phylogenetic Annotation and INference Tool).

Description of phylogeny
The family consists of 3 paralogous vertebrate clades -- p53, p63, and p73 -- with (an urchin and) an insect outgroup. In P-POD OrthoMCL, the p63 and p73 clades form a single family, and the fly p53 is in the same family as the vertebrate p53's. There are divergent rat (Q6QI11) and platypus p53's.

-p53 forms a homotetramer, but the only annotation that reflects this is "p53 binding" on mouse p73. Propagate to AN0.
-DNA binding terms: Propagate "damaged," "double-stranded," and "sequence-specific" DNA binding to AN0. Also "chromatin binding."
-Transcription: Propagate "transcription factor activity," "transcription repressor activity," and "transcription activator activity" to AN0.
-Do not propagate rat p53 "ubiquitin protein ligase binding" because it is based on ubiquitination _of_ p53 shown in PMID 16330492.
-Mouse p53 "histone deacetylase regulator activity" IDA from PMID 15657445 is based on an in vitro assembly of chromatin; the authors suggest that p53 recruits HDAC and do not suggest any sort of enzymatic activation of HDAC. Do not propagate. A better term is the BP "positive regulation of histone deacetylation" IDA from the same paper.

-p53 is a transcription factor, so propagate "nucleus," "transcription factor complex," and "chromatin" to AN0; limit "replication fork" to p53 clade. Note that there are nucleus-specific children of "chromatin" and "replication fork," but they haven't been used for annotation.
-Propagate "dendrite" to p63 clade.
-"Cytosol" is only seen in p53 clade, but I can't find any membrane-spanning regions, so propagate to AN0.

-The first priority should be to make sure that annotations to BP terms specifically pertaining to the p53 pathway are used. There are seven:
GO:0030330 : DNA damage response, signal transduction by p53 class mediator
3 is_a children (high priority):
GO:0006977 : DNA damage response, signal transduction by p53 class mediator resulting in cell cycle arrest
GO:0006978 : DNA damage response, signal transduction by p53 class mediator resulting in transcription of p21 class mediator
GO:0042771 : DNA damage response, signal transduction by p53 class mediator resulting in induction of apoptosis
3 "regulates" children (lower priority):
GO:0043516 : regulation of DNA damage response, signal transduction by p53 class mediator
GO:0043517 : positive regulation of DNA damage response, signal transduction by p53 class mediator
GO:0043518 : negative regulation of DNA damage response, signal transduction by p53 class mediator

(Also, GO:0006917 "induction of apoptosis" has the narrow synonym "induction of apoptosis by p53," but this is a parent of 42771, so annotations to 42771 will cover this term.)

-42771 is found in all 3 clades plus the fly outgroup. Propagate to AN0.
-6978 is found in the p53 clade only, but there are examples in other clades curated to parent terms:
-Mouse p73 induces p21 (PMID 16044147); this is recorded as "positive regulation of transcription, DNA-dependent."
-Zebrafish (weak effect) and human (stronger effect) p73 induce p21 (PMID 15541366); recorded as "regulation of transcription, DNA-dependent" for zebrafish.
-Mouse p63 inhibits p21 transcription (PMID 16618808); recorded as "negative regulation of transcription from RNA polymerase II promoter."
Propagate 6978 to AN0.
-There are no annotations to 6977 and only one (mouse p73) to GO:0007050 "cell cycle arrest." There are multiple annotations in all clades to GO:0051726 "regulation of cell cycle" and its children, but the most specific and most accurate is GO:0031571 "G1/S DNA damage checkpoint." Propagate 31571 to AN0.
-Propagate "positive regulation of histone deacetylation" to p53 clade. See MF section for explanation.
-Propagate "cellular response to UV," "response to gamma radiation," and "response to X-ray" to AN0.
-There are multiple annotations to terms involved in transcriptional regulation, but they are all covered by GO:0000122 "negative regulation of transcription from RNA polymerase II promoter" and GO:0010552 "positive regulation of gene-specific transcription from RNA polymerase II promoter." Propagate these to AN0.
-There are many annotations to terms involved in apoptosis and regulation of apoptosis, including annotations in 3 clades to "neuron apoptosis" (or its children) and in 2 clades for "regulation of neuron apoptosis." Propagate the regulation term to AN0; first, though (to reduce redundancy), propagate "pos. reg. of neuron apoptosis" to p53 clade and "neg. reg. of neuron apoptosis" to p73 clade. Also, propagate "anti-apoptosis" to p63 clade.
-A "cell cycle arrest" IDA is found for mouse p73, but there is a "neg. reg. of cell cycle" IMP (potential parent term, see ontology question below) for zebrafish p53 which is probably due to arrest. Also, this is a known function of p53, so propagate to AN0.
-Mouse p73 has an IDA to GO:0043508 "negative regulation of JUN kinase activity." The activity was observed in the delta-N-p73 isoform, "which is the only p73 isoform expressed in sympathetic neurons" (PMID 15483136). This seems very specific, so propagate the term to the p73 clade.

Questions for MOD curators

-Worm: Is there a worm homolog that should be in this tree?

WB comment July 8, 2010 1:20:57 PM EDT:

There does appear to be a p53 superfamily member, CEP-1, in elegans.

CEP-1's role in germline apoptosis has been fairly well studied, and I actually just came across a review today about invertebrate p53-like proteins:

PANTHER reply, July 8, 2010 3:06:08 PM EDT:

This is an interesting one. Apparently CEP-1 has diverged so extensively that its homology to other members of the P53 family is no longer detectable by BLAST, or by any HMMs that were constructed using sequence information alone (from InterPro, including PANTHER, Pfam, SMART, etc). A structural analysis reveals the homology, though, and HMMs using this information recognize the homology (e.g. Superfamily and CDD). We?ll need to manually add it to the family, which will take a little time. If we can get a decent alignment using sequence information alone, it shouldn?t take too long, but if we?d need to manually adjust the alignment based on a protein structure alignment this would not be easy. I?ll keep you posted.

-Rat: Does Q6QI11 belong in this family?

-All organisms with "protein binding" annotations: Is there a more specific term you can use?

Human (EBI) replies, July 29, 2010 7:53:51 AM EDT: I have updated all the protein binding annotations that I could

-All organisms with "replication fork" annotations: Can you justify "nuclear replication fork" (GO:0043596)?
-All organisms with "chromatin" annotations: Can you justify "nuclear chromatin" (GO:0000790)?

-Mouse: Is the Ptprv-p53 IGI to GO:0006977 "DNA damage response, signal transduction by p53 class mediator resulting in cell cycle arrest" from MGI:3589512 (PMID 16107883) reciprocal? I.E., does it justify making an annotation to 6977 for p53? (SF# 3025875)

MGI, July 13, 2010 10:11:08 AM EDT: yup, Li already made these.

-Mouse p63: Please verify the annotation to "cloacal septation." Do mice have a cloaca?

MGI, July 13, 2010 10:11:08 AM EDT: Yup. In the embryonic stages they have one.

-Mouse: There are several developmental terms annotated with IDA's. Should they be IMP's instead? Examples:
GO:0048147 : negative regulation of fibroblast proliferation

MGI, July 13, 2010 10:11:08 AM EDT: Changed to IMP.

GO:0002064 : epithelial cell development
GO:0001736 : establishment of planar polarity
GO:0030216 : keratinocyte differentiation
GO:0043616 : keratinocyte proliferation

MGI, July 13, 2010 10:11:08 AM EDT:

These four are tricky ones. They used knock-out cells, but transfected an isoform of the gene product back in to test the function of the specific isoform. I think that the IDAs are more appropriate here because they positively test the activity of the isoform in a null background.

Questions for ontology curators
-Shouldn't GO:0007050 "cell cycle arrest" be a child of GO:0045786 "negative regulation of cell cycle," which is defined as "Any process that _stops_, prevents or reduces the rate or extent of progression through the cell cycle" (emphasis mine)? (SF# 3025958)
MGI, July 13, 2010 10:11:08 AM EDT: Fixed