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El PMN: Tu amigo en el metabolismo de plantas. Kate Dreher curator PMN/AraCyc/TAIR. PMN = The P lant M etabolic N etwork Created in 2008 Funded by the National Science Foundation www. plantcyc.org How do I find information?

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el pmn tu amigo en el metabolismo de plantas

El PMN:Tu amigo en el metabolismo de plantas

Kate Dreher

curator

PMN/AraCyc/TAIR

putting the pmn to use
PMN = The Plant Metabolic Network

Created in 2008

Funded by the National Science Foundation

www. plantcyc.org

How do I find information?

How do I build a metabolic pathway database for my favorite plant?

Sue Rhee

(PI)

Peifen Zhang

(Director)

Putting the PMN to use
basic searching practice problems
1. What are some of the pathways that talk about “salt stress” in the PMN?Basic searching: practice problems
  • 2. How many “amides” are in PlantCyc?
  • 3. Are there more PlantCyc or AraCyc reactions listed under the E.C. category 3 (hydrolases)?
basic searching practice problems1
1. What are some of the pathways that talk about “salt stress” in the PMN?Basic searching: practice problems

Arabidopsis thaliana proline biosynthesis I

Measurement of free proline content and gene expression and enzyme activity levels from salt-stress treated plants [ Roosens98 ] showed that in young ...Arabidopsis thaliana choline biosynthesis I

Summers93: Summers PS, Weretilnyk EA (1993). "Choline Synthesis in Spinach in Relation to Salt Stress." Plant Physiol 103(4);1269-1276. PMID: 12232019 ...Arabidopsis thaliana arginine biosynthesis III

"Isolation of the ornithine-delta-aminotransferase cDNA and effect of salt stress on its expression in Arabidopsis thaliana." Plant Physiol 117(1);263-71. ...PlantCyc phosphatidylcholine biosynthesis I

"Regulation of phosphatidylcholine biosynthesis under salt stress involves choline kinases in Arabidopsis thaliana." FEBS Lett 566(1-3);115-20. ...PlantCyc putrescine biosynthesis via N-carbamoylputrescine

... and biosynthetic gene expression in Arabidopsis thaliana under salt stress. ... knockout mutation of ADC2 gene reveals inducibility by osmotic stress. ...

  • 2. How many “amides” are in PlantCyc?
    • 9
  • 3. Are there more PlantCyc or AraCyc reactions listed under the E.C. category 3 (hydrolases)?
    • More AraCyc reactions
basic searching practice problems2
Find a pathway in PlantCyc whose end product is tryptophan.

Is it part of any superpathways?

How many anthinilate synthases from Medicago truncatula are associated with this pathway?

Is there experimental or computational support for their assignment to this reaction?

Which enzyme is subject to feedback regulation? Please give gene code, too.

What type of experimental support exists for this enzyme?

What is its Km for chorismate?

Find the protein sequence for the closest ortholog in papaya

What is the length of the full-length coding sequence for its closest homolog in soybean (in soybeans)?

How many species are associated with reaction 5.3.1.24?

Find a paper that talks about an enzyme with similar activity in Zea mays?

What is the chemical formula of anthranilate?

What are two synonyms for this compound?

What other pathway is it found in?

Basic searching: practice problems
basic information practice problems
Find a pathway in PlantCyc whose end product is tryptophan.

TRPSYN-PWY – tryptophan biosynthesis

Is it part of any superpathways?

YES, superpathway of phenylalanine, tyrosine and tryptophan biosynthesis

How many anthinilate synthases from Medicago truncatula are associated with this pathway? 4

Is there experimental or computational support for their assignment to this reaction? Computational

Which enzyme is subject to feedback regulation?anthranilate synthase , At5g05730

What type of experimental support exists for this enzyme? mutant phenotype; genetic interaction

What is its Km for chorismate? 180 uM

Find the protein sequence for the closest ortholog in papaya

MQTLGFSYRLVPSGRRFSPVPANGISGARSSSLVNVRTFKCMSLSSPSLVCDVKKFAEASKHGNVVPLYH SIFSDQLTPVLAYRCLVKEDDREAPSFLFESVEPGFQASSVGRYSVVGAQPTIEIVAKENKVTIMDHEGG TLSEEYVQDPMMIPRRISEGWKPQLIDELPDTFCGGWVGYFSYDTIRYVEKKKLPFSMAPEDDRNLADIH LGLYDDVIVFDHVEKKAHVIHWVRLDQYSSAEKAYNDGLKRLEKLVAKVQDIDPPRLSPGSVDLQTRQFG PSLRKSTMTSEEYKMAVLEAKEHILAGDIFQIVLSQRFERRTFADPFEVYRALRVVNPSPYMTYLQARGC ILVASSPEILTRVEKKKIVNRPLAGTVRRGKTTAEDEMLEKQLLNDAKQCAEHIMLVDLGRNDVGKVTGE LHDHLTCWDVLRAALPVGTVSGAPKVKAMELIDQLEVTRRGPYSGGFGGISFTGNMDVALALRTIVFPTG THYNTMYSYKDVENRRDWIAHLQAGAGIVADSNPDDENQECHNKVAGLARAIDLAESAFVNK*

What is the length of the full-length coding sequence for its closest homolog in soybean? 1593 bp

How many species are associated with reaction 5.3.1.24? three

Find a paper that talks about an enzyme with similar activity in Zea mays?

Some physical characteristics of the enzymes of L-tryptophan biosynthesis in higher plants

What is the chemical formula of anthranilate? C7H7NO2

What are two synonyms for this compound? anthranilic acid, 2-aminobenzoic acid

What other pathway is it found in? acridone alkaloid biosynthesis

Basic information: practice problems
advanced searching
Advanced Query Form

Downloading the whole database

Advanced searching
advanced searching practice problems
Find a list of all of the enzymes with a pI between 5 and 6.5 that are glucosyltransferases.

What species are they found in?

What is the highest pI and the lowest pI in the group?

Get a list of all of the reactions that are children of E.C. 2.4.1. and identify the compounds in the right and the left

How many reactions are in your original list?

If you eliminate all of the reactions that have rhamnose in their name, how many do you have left?

Look for all the pathways that have more than one hypothetical reaction in them.

Get all of the citations associated with them

How many of the pathways have more than 4 hypothetical reactions associated with them?

Advanced searching: practice problems
advanced searching practice problems1
Find a list of all of the enzymes in PlantCyc with a pI between 5 and 6.5 that are glucosyltransferases.

How many are there?

What species are they found in?

What is the highest pI and the lowest pI in the group?

Get a list of all of the reactions that are children of E.C. 2.4.1. in PlantCyc and identify the compounds in the right and the left in your output list

How many reactions are in your original list?

If you eliminate all of the reactions that have rhamnose in their name, how many do you have left?

Look for all the pathways in PlantCyc that have more than one hypothetical reaction in them.

Get all of the citations associated with them

How many of the pathways have more than 4 hypothetical reactions associated with them?

Advanced searching: practice problems
advanced searching practice problems2
Find a list of all of the enzymes in PlantCyc with a pI between 5 and 6.5 that are glucosyltransferases.

How many are there? 13

What species are they found in?

Zea mays, Dorotheanthus bellidiformis, Cicer arietinum, Brassica napus

Pinus strobus

What is the highest pI and the lowest pI in the group? 5, 6.36

Get a list of all of the reactions that are children of E.C. 2.4.1. in PlantCyc and identify the compounds in the right and the left in your output list

How many reactions are in your original list? 189

If you eliminate all of the reactions that have rhamnose in their name, how many do you have left? 174

Look for all the pathways in PlantCyc that have more than one hypothetical reaction in them. 17

Get all of the citations associated with them

How many of the pathways have more than 4 hypothetical reactions associated with them? 6

Advanced searching: practice problems
omics viewer
Do your experiment (or get your data)

Create your input file

Upload and analyze your data

Absolute values

Time course

Relative values

OMICs viewer
omics viewer practice problems
Use “PMN_taller_OMICS_practice_problem”

The columns are:

0 – gene identifiers

1 – time point 1

2 – time point 2

3 – time point 3

4 – rapid change (ratio of time point 1 to 2)

Find the pathways that have gene expression values of over 600 at time point 2. Which one has the highest number of steps affected in the pathway?

Create an animation of all three time points

Look for pathways that had genes whose expression changed more than 4-fold between the first and last time points. How many appear?

Use the data you have already calculated and look for pathways that have gene expression changes of more than 6-fold in the first time interval. How many appear?

OMICs viewer: practice problems
omics viewer practice problems1
Use “PMN_taller_OMICS_practice_problem”

The columns are:

0 – gene identifiers

1 – time point 1

2 – time point 2

3 – time point 3

4 – rapid change (ratio of time point 1 to 2)

Find the pathways that have gene expression values of over 600 at time point 2. Which one has the highest number of steps affected in the pathway? starch degradation

Create an animation of all three time points

Look for pathways that had genes whose expression changed more than 4-fold between the first and last time points. How many appear? 13

Use the data you have already calculated and look for pathways that have gene expression changes of more than 6-fold in the first time interval. How many appear? 6

OMICs viewer: practice problems
building your own metabolic database
Step 1: Genes / Nucleotide Sequences

Genome sequencing

Unigene builds

ESTs

Step 2: Annotate Predicted Proteins

Annotation source:

PMN pipeline

JGI or other sequencing consortium

Small group of dedicated scientists, etc.

Annotation tasks:

Give enzymes a name

Add GO molecular function terms

Assign to a MetaCyc or EC reaction

Use Pathologic and MetaCyc or PlantCyc to create new pathway database

Remove non-enzymatic proteins before prediction

After first round of prediction, review unassigned enzymes

Repredict

Validate new database

Building your own metabolic database
validating your own metabolic database
Validating your own metabolic database
  • PathoLogic errs on the side of over-prediction
  • Curators / Scientists validate pathways . . .
examine predicted pathways
Examine predicted pathways
  • Search for evidence in published papers, books, etc.
    • Is the pathway described in the literature for my species?
    • Are the crucial metabolites described in in the literature for my species?
    • Are there unique reactions associated with the pathway that have assigned genes?
    • Is there evidence that this is a universal plant pathway?
      • Is it on the PMN “green list?”
    • Is there evidence that this pathway is NEVER found in plants?
      • Only relevant when prediction is made using MetaCyc
      • Is it on the PMN “black list?”
make necessary changes
Make necessary changes
  • Remove pathways not found in My species
    • glycogen biosynthesis
    • C4 photosynthesis
    • caffeine biosynthesis
  • Edit pathways operating via a different route
    • Phenylalanine biosynthesis in bacteria vs. My species
make necessary changes1
Make necessary changes
  • Edit pathways operating via a different route

AraCyc Pathway: phenylalanine biosynthesis

increase the data content
Increase the data content
  • Addpathways from My species not present in the reference database
      • Secondary metabolites
  • Add additional compounds, reactions, and enzymes from My species that cannot be put in a known pathway
  • Write new summaries or revise imported summaries for pathways
provide evidence codes
Provide evidence codes
  • Evidence codes are used for:
    • Pathways
    • Enzyme activities
  • General types of support
    • EV-EXP -> experimental
    • EV-COMP -> computational
    • EV-IC -> inferred by a curator
    • EV-AS -> author statement
  • Additional information about evidence can be captured:
    • EV-EXP-IDA-PURIFIED-PROTEIN
      • Inferred from direct assay
    • EV-EXP-IMP-REACTION-BLOCKED
      • Inferred from mutant phenotype
    • EV-COMP-AINF
      • Inferred by computation automatically without human oversight
provide evidence codes1
Provide evidence codes
  • Enzyme activity assays
  • Radiotracer experiments
we d love to have you as part of our plant metabolic network of experts and friends
We’d love to have you as part of ourPlant Metabolic Network of experts and friends!
  • Add your database to the PMN
    • The PMN can host it as a separate individual species or multi-species database
    • Enzymes from your validated pathways can be incorporated into PlatCyc pathways
    • Newly curated pathways from your species can be added to PlantCyc
    • Pathways
    • Enzyme activities
  • . . . but you don’t have to wait to make a whole new database!
please help us to improve the pmn
Please help us to improve the PMN
  • Review existing pathways
    • Volunteer to go over a set of pathways in your area of expertise
    • E-mail us any time you see a problem
    • Use our feedback form

curator@plantcyc.org

please give us advice and information1
Please give us advice and information
  • Provide new data
    • Submit any information concerning:
      • Pathways
      • Enzymes
      • Reactions
      • Compounds
we are here to help www plantcyc org
We are here to help: www.plantcyc.org
  • Please use our data
  • Please use our tools
  • Please help us to improve our databases!
  • Please contact us if we can be of any help!
    • Make an appointment to meet with me during my visit
      • (Puedo tratar de hablar en español)

curator@plantcyc.org

www.plantcyc.org

pmn acknowledgements
PMN Acknowledgements

Peifen Zhang (Director)

Sue Rhee (PI)

Eva Huala (Co-PI)

Current Curators:

- A. S. Karthikeyan (curator)

Recent Past Contributors:

- Christophe Tissier (curator)

- Hartmut Foerster (curator)

Collaborators:

- Peter Karp (SRI)

- Ron Caspi (SRI)

- SRI Tech Team

- Lukas Mueller (SGN)

- Anuradha Pujar (SGN)

- Gramene and MedicCyc

Tech Team Members:

- Bob Muller (Manager)

- Larry Ploetz (Sys. Administrator)

- Raymond Chetty

- Anjo Chi

- Vanessa Kirkup

- Cynthia Lee

- Tom Meyer

- Shanker Singh

- Chris Wilks

we are here to help www plantcyc org1
We are here to help: www.plantcyc.org
  • Please use our data
  • Please use our tools
  • Please help us to improve our databases!
  • Please contact us if we can be of any help!
    • Make an appointment to meet with me during my visit
      • (Puedo tratar de hablar en español)

curator@plantcyc.org

www.plantcyc.org