paint conf call 062414
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PAINT conference call GO:0000725 recombinational repair PTHR16771 and PTHR23074 Monica Munoz-Torres, PhD | @monimunozto Berkeley Bioinformatics Open-Source Projects (BBOP)
Genomics Division, Lawrence Berkeley National Laboratory 24 June, 2014
UNIVERSITY OF CALIFORNIA
Outline 1. Thought process 2. GO Term: recombinational repair 3. PTHR16771
• Proteasome Complex Subunit DSS.
4. PTHR23074 • AAA ATPase.
PAINT Conf Call GO:0000725 recombinational repair
Outline 2
Thought Process
• Find gene with annotation to GO term of interest
• Inspect details at UniProt (FASTA, GO terms, etc.)
• Link to PubMed, extract information
• Inspect CDD (e.g. BLAST vs nr)
• Inspect all annotations associated with gene in question reviewing both tree and MSA
• Propagate as appropriate
Thought Process 3
GO Term
Accession: GO:0000725
Name: recombinational repair
Ontology: biological_process
Definition: A DNA repair process that involves the exchange, reciprocal or nonreciprocal, of genetic material between the broken DNA molecule and a homologous region of DNA. Source: GOC:elh
GO Term 4
PTHR16771
PTHR16771: Proteasome Complex Subunit DSS1
• BRCA2 is a breast cancer susceptibility gene implicated in the repair of double-strand breaks by homologous recombination with RAD51.
• DSS1 interacts with BRCA2 in a domain-specific way
• DSS1 is required for the BRCA2-RAD51 complex to become associated with sites of DNA damage and for the maintenance of genomic stability, a function conserved from lower eukaryotes to humans.
• DSS1 is involved in DNA damage repair via homologous recombination.
PTHR16771 6
DSS1_SEM1 [cd13768, pfam05160] proteasome complex subunit DSS1/SEM1 (s:CDD)
• The evolutionarily conserved deleted in split hand/split foot protein 1 (DSS1)/Sem1 is a subunit of the regulatory particle (RP) of the proteasome.
PTHR16771 7
Pruned
• Inspecting MSA and comparing with the Conserved domains of DSSm1/SEM1 family, I pruned the following species:
20140505: Pruned TETTS_TTHERM_00227230
20140505: Pruned ORYSJ_P0693B08.32
20140505: Pruned PUCGT_PGTG_08936
PTHR16771 8
Reasons for pruning
• ORYSJ_P0693B08.32 and PUCGT_PGTG_08936 have long (20 - 40 aa) strings of residues interrupting the conserved domain of DSS1 (increasing the length of the peptide), which has been very well characterized to 71aa in length.
• TETTS_TTHERM_00227230 has an extended string of ~40 aa at the beginning of the protein, and the second portion of its conserved domain is severely corrupted (degenerate) in comparison to all other proteins in the family
PTHR16771 9
Reasons for pruning
PTHR16771 10
Decision # cellular_component 20140513: Has function proteasome complex (GO:0000502) in
Eukaryota_PTN000423400 20140513: Colocalizes with nucleus (GO:0005634) in
Eukaryota_PTN000423400 # biological_process 20140513: Has function proteolysis (GO:0006508) in
Eukaryota_PTN000423400 20140509: Has function double-strand break repair via homologous
recombination (GO:0000724) in Eukaryota_PTN000423400 # Pruned from tree 20140513: Pruned ORYSJ_P0693B08.32 20140513: Pruned TETTS_TTHERM_00227230 20140513: Pruned PUCGT_PGTG_08936
PTHR16771 11
Questions
• Should I annotate other terms? or only concentrate on the one I am reviewing?
• I find myself hesitating about propagating due to degeneracy of conserved domains in MSA (thus, I pruned three species).
How much weight should MSA have on decisions? Does everyone else rely heavily on them?
PTHR16771 12
MF: Peptidase activity?
• Although Human_SHFM1 has an annotation to peptidase activity GO:0008233, the paper reporting the IDA does not make any claims of peptidase activity, or any of its synonyms (i.e. hydrolase, acting on peptide bonds; peptide hydrolase activity; protease activity; proteinase activity).
• “Yeast and mammalian two-hybrid assays showed that DSS1 can associate with BRCA2 in the region of amino acids 2472 to 2957 in the C terminus of the protein. Using coimmunoprecipitation of epitope-tagged BRCA2 and DSS1 cDNA constructs transiently expressed in COS cells, authors demonstrated an association. Furthermore, endogenous BRCA2 could be coimmunoprecipitated with endogenous DSS1 in MCF7 cells, demonstrating an in vivo association.”
• No further details on the nature of the bond are discussed. Should this term be removed? or changed to 'catalytic activity' instead?
PTHR16771 13
Questions
From the "Evidence" tab:
• What does "has function nucleus" mean? and "has function proteasome complex"? and how can I avoid that syntax? am I supposed to avoid the syntax? “has function” vs “contributes to”?
• Warnings: (see next)
PTHR16771 14
Warnings that disappear… #WARNINGS
WARNING: Experimental data changed!
The following annotations are no longer supported by experimental evidence
and have been removed:
Opisthokonta_PTN000423402 to double-strand break repair via homologous recombination
Bilateria_PTN000423403 to double-strand break repair via homologous recombination
PTHR16771 15
• I got a warning about experimental evidence being removed, but the experimental evidence is still being displayed. My initial thought was that the culprit was my "redundant" propagation of a process annotation to a less derived node, then a more ancestral one. Is this the cause? ---> Update: the errors disappeared. ???
Previously: Decision
#cellular component
20140513: Has function proteasome complex (GO:0000502) in Eukaryota_PTN000423400
20140513: Colocalizes with nucleus (GO:0005634) in Bilateria_PTN000423403
20140513: Colocalizes with nucleus (GO:0005634) in Eukaryota_PTN000423400
20140507: Has function nucleus (GO:0005634) in Bilateria_PTN000423403
#biological process
20140513: Has function proteolysis (GO:0006508) in Eukaryota_PTN000423400
20140509: Has function proteolysis (GO:0006508) in Opisthokonta_PTN000423402
20140509: Has function double-strand break repair via homologous recombination (GO:0000724) in Eukaryota_PTN000423400
#Pruned
20140513: Pruned ORYSJ_P0693B08.32
20140513: Pruned TETTS_TTHERM_00227230
20140513: Pruned PUCGT_PGTG_08936
PTHR16771 16
PTHR23074
PTHR23074: AAA ATPase • AAA+ Superfamily: “ATPases Associated with a wide variety of
cellular Activities”
• An ancient group of ATPases belonging to the ASCE (for additional strand, catalytic E) division of the P-loop NTPase fold.
• Members function as molecular chaperons, ATPase subunits of proteases, helicases, or nucleic-acid stimulated ATPases.
• AAA+ proteins contain several distinct features in addition to the conserved alpha-beta-alpha core domain structure and the Walker A and B motifs of the P-loop NTPases.
PTHR23074 18
FIGNL1: RAD51-binding protein fidgetin-like 1
• RAD51 recombinase plays central role in homologous recombination (HR), which is critical for repair of DNA double-strand breaks.
• FIGNL1 specifically interacts with RAD51 through its conserved RAD51 binding domain.
• Forms part of protein complex FIGNL1 + KIAA0146/SPIDR, required for DNA repair
PTHR23074 19
PTHR23074: FIGNL1
PTHR23074 20
Inspecting Conserved Domains
PTHR23074 21
FIGNL1
KATNAL2
VPS4A
PTHR23074: KATNAL2
PTHR23074 22
Observations:
• Paraphyletic
• Plants + animals
• No Experimental Annotations
• AAA ATPase conserved domain
Decision
FIGNL1's RAD51 binding domain (FRBD) is required for "regulation of double-strand break repair via homologous recombination" (GO:0010569) activity.
FIGNL1 and its homologs contain also an AAA superfamily domain, and vsp4 domain. The required binding domain is only present in this family. Members from other clades were reviewed. Neither family had the required FRBD and one did not have a vsp4 domain either.
Decision to propagate GO:0010569 only to Eukaryota_PTN000553645 as the Most Recent Common Ancestor for this biological process for the FIGNL1 gene.
PTHR23074 23
Propagated term
Footer 24
Propagated term
PTHR23074 25
Evidence (text) # molecular_function
20140603: Contributes to ATPase activity (GO:0016887) in root_PTN000553509
# cellular_component
20140603: Has function nucleus (GO:0005634) in root_PTN000553509
# biological_process
20140603: Has function regulation of double-strand break repair via homologous recombination (GO:0010569) in Eukaryota_PTN000553645
# Notes
PTHR23074 26
Software Reports • Space to enter PANTHER protein
family ID not visible
• Arrow heads do not act as expected: i.e. do not move spaces along MSA
Software Reports 27
Software Reports
• Large family eats up CPUs; PAINT freezes (tmp)
• Is right-click on experimental annotation supposed to highlight the MRCA node for them? If so, not evident.
• Permanent Tree ID (PANTREE) links broken / blank pages
• Resizing a window takes several seconds
• Challenging to propagate to “off-screen” node
Software Reports 28
UNIVERSITY OF CALIFORNIA
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