PMID:32487759 Functional Coupling between the Unfolded Protein Response and Endoplasmic Reticulum/Golgi Ca 2+-ATPases Promotes Stress Tolerance, Cell Wall Biosynthesis, and Virulence of Aspergillus fumigatus

[CuzickA]

This is a publication that @JonMWilkes has selected to curate.

Curation link
https://canto.phi-base.org/curs/fa48bc08d05f08e3

[JonMWilkes]

There is an issue with the hacA transcription factor. As far as I can determine there is an induced and uninduced (differentially spliced) form of this transcription factor which promoted the UPR. Thus uniprot records a hacAi and a hacA form (B9UNL5 and Q4WEY8 respectively 342 and 433 amino acids). I have included both entries but need to determine which is appropriate for the annotation -

[ValWood]

That is strange. I think UniProt might need to merge those into a single entry and represent the isoforms within that entry, with the active form as the active isoform as the canonical sequence. I am tagging @Antonialock who looks at some of the fungal pathogens in UniProt.
I don't know much about HAC1 (which is present in budding yeast but absent in fission yeast), the regulation of UPR in fission yeast is slightly different.

[ValWood]

I don't think this is spliced and unspliced from.
The first one B9UNL5 is derived from https://www.ebi.ac.uk/ena/browser/api/embl/ACJ61678.1?lineLimit=1000
listed as /isolate="H237"
The second is from
Neosartorya fumigata (strain ATCC MYA-4609 / Af293 / CBS 101355 / FGSC A1100)
DOe that help?

Both are currently in the "unreviewed" portion of Unitrot (Trembl), rather than the reviewed (Swiss-prot).

[JonMWilkes]

Thanks for this. Firstly I'm not a mycologist so get a bit confused re. the different binomial classifications - from NCBI taxonomy browser these are the same 'species' ? I used the primer sequences listed in TableS2 for the hacA induced and uninduced to blast against A. fumigatus on NCBI blast. It is this approach which has led to the flagging up of Q4WEY8 as hacA and B9NUL5 as hacAi. This began as an exercise in trying to identify the correct Uniprot code for this gene, but as can be seen the issue has got a bit out of hand. I'm minded to list the code as Q4WEY8

[JonMWilkes]

I've returned to this paper concentrating on the pathogen phenotype entries today (as I said this is something of a 'mega' paper). I have identifies a few terms in the PHIPO entries that might be needed:
Evidence code "Inferred by sequence orthology"
Thermal stress phenotypes (increased, decreased and unchanged)
Chemical sensitivity to BAPTA, calcimycin, amloderone
Disruption of homeostasis (particularly Ca2+ in the case of this paper)

[CuzickA]

Hi @JonMWilkes, thanks for noting that we may need to create the above terms.

There is an option to suggest a PHIPO term when making an annotation. It looks like you have made some good notes in the comments column. If these are to do with new PHIPO term suggestions they can be entered like this

1. either using the edit annotation option
   In the Term selection section, select 'suggest'
   
   The term suggestion and a draft definition (optional) can be added in this pop up option.
   

2. or if you are making new annotation, there is an option at the bottom of the PHIPO term selection page for 'suggest a new child term'.
   

By adding in the suggested PHIPO terms to the annotations it makes it easier for the reviewing of the curation session and future decisions about whether new terms needed to be created or edited. If new PHIPO terms/ edits are required these with then be transferred to tickets on the PHIPO GitHub tracker.

Please let me know if you have any questions.

[JonMWilkes]

Thanks for that, I'll refine these following the metagenotype entries.

[CuzickA]

Hi @JonMWilkes,

I just looked at this session and noticed that the Pathogen-Host Interaction Phenotypes have been annotated with the terms that should be used in the Annotation Extension 'infective ability'. We had some discussion about this in #107.

I have reactivated this session. Please could you go back to these annotations and enter the appropriate PHIPO term for the observed phenotype and move the terms like 'unaffected pathogenicity' over to the AE infective ability. Many thanks.

[JonMWilkes]

Hi @CuzickA ,
as requested I have edited the pathogen-host phenotypes to place the virulence phenotypes in the annotation extension fields.

[CuzickA]

From the paper 'The A. fumigatus gene srcA (Afu6g06740)', gene name in UniProt is AFUA_6G06740.

To edit the genotype in this session

1. go to pathogen genotype management
2. 

Select option to 'edit details' from right arrow

3. 
   Select 'edit'

4. 
   edit the text in the allele name
   
   and select 'finish'
   Genotype edits

I will also edit the multi-allele genotypes after the other single component genotypes have been edited.

Hi @JonMWilkes, this is a useful trick to capture the gene name in the genotype whilst we wait for UniProt to update the gene names. Did you send email requests to UniProt request gene name updates for this publication?

My changes now look this

[CuzickA]

@JonMWilkes FYI for Fig.2A

edited to

In the conditions I changed the 'IMA plates' to 'rich medium' and the detail can be left in the comments.
I have used the '3 dpi' term. There is a ticket on the PHI-ECO tracker where I plan to change the def on these terms to include inoculation onto media or host so that i don't have to duplicate all of these timing terms.

For Fig2D
Changed the PHIPO term here as the expt is looking at hyphae morphology

[CuzickA]

Fig 3

1. We could use the FYPO term here (and add it to PHIPO)
   sensitive to heat shock (FYPO_0001408)
   Def: A cell phenotype in which cells show increased sensitivity to heat shock. Heat shock is a form of heat stress in which cells are briefly exposed to a very high temperature.

But, in this expt it is not a brief exposure to a high temp. The cells are grown at the high temp for the duration of the expt.

2. Perhaps it would be better to create a new PHIPO term for 'sensitive to heat stress' and then capture that the temp is increased in the conditions? If we go down this route we would need to remove the 'high temp' from all the annotations where the expt were conducted at the standard (?) 37oC and just add 'high temp' for those at 45oC.

The current 'high temp' definition is
'The experiment was performed, or cells were grown, at a high temperature (typically exceeding ~32 degrees Celsius).'

Which is why it has been added to all the standard temp 37oC expt.

@ValWood, do you think option 1 or 2 is best for this expt where the cells are grown at either 37oC or 45oC for 7 days but without a temp shift.

[CuzickA]

Fig 4

It might be better to use 'decreased cell wall integrity' as the PHIPO term and then add the cell wall perturbers into the conditions

[CuzickA]

Note to self: I have checked pathogen phenotype annotations for Fig 2,3,4,5 and S6C,D.

@JonMWilkes You are right- this is a bit of a 'mega' paper!
Please could you

1. read through this ticket
2. look through the changes I have made for the above checked pathogen phenotype annotations
3. add in the WT control metagenotype annotations as explained below (I will reactive the session for you to do this)

Where possible for metagenotype annotations we also make a wild type control metagenotype and annotate this. This can then be connected to the mutant/altered metagenotype using the Annotation extension option 'compared to control'

Here is an example

[JonMWilkes]

@CuzickA - I have looked over the ticket and the modifications to the pathogen phenotypes. I agree with them, adds a lot of detail. On the question of defining the wild type genotype, when there are several genes being manipulated in the study can we/ how do we specify a single wild type genotype or do we need to create a WT counterpoint to every mutant combination?

[CuzickA]

Hi @JonMWilkes, sounds good :-).

We need a WT counterpoint to every mutant combination (when possible).
For a single pathogen gene within metagenotype see example above
For two pathogen genes within metagneotype

Note: the compared to control AE looks incorrect here. I think it should be compared to

[JonMWilkes]

Hi @CuzickA , I have in the meantime specified a genotype expressing all genes of interest (within a particular background strain) similar to what you suggest for the double deletion control. Logically this should be usable as a wild-type control for all mutant genotypes involving any combination of the listed genes. Additionally I have attempted to code for a deletion/rescue genotype - let me know what you think of this. Is there a case for simply being able to denote the 'parental strain' as a genotype in its own right ( including background genotypes such as NHEJ deficiency)?

[JonMWilkes]

Hi @CuzickA - following our discussion re. WT genotypes I need to access the annotations in order to correct them. Could you release it to me please,
Thanks - Jon

[CuzickA]

Hi @JonMWilkes, thanks for reminding me. The session has now been reactivated.

Hi @JonMWilkes, this is a useful trick to capture the gene name in the genotype whilst we wait for UniProt to update the gene names. Did you send email requests to UniProt request gene name updates for this publication?

Also, just following up from my comment above - did you manage to send gene name update requests to UniProt for this paper?

[JonMWilkes]

Hi @CuzickA , thanks for the reminder, have now contacted Uniprot re. gene names. Does the reference to a 'useful trick' above have a link, or what does it refer to? Thanks JON

[CuzickA]

That's great @JonMWilkes.

The 'useful trick' above is where I have provided screenshots about editing the genotype name to use the updated 'gene' name within the genotype rather than the gene name given by UniProt.

The comment above with these screenshots starts with the text "From the paper 'The A. fumigatus gene srcA (Afu6g06740)', gene name in UniProt is AFUA_6G06740." in case you are trying to wade through the comments to find it.

[CuzickA]

[CuzickA]

Email dated 23 Aug 2022

Links to UniProt entries - looks like gene name updates have not been release yet.
https://www.uniprot.org/uniprotkb/Q4WHC8/entry
https://www.uniprot.org/uniprotkb/Q4WND5/entry

[Antonialock]

I see that the names have gone into UniProt and will be relased next release (I don't know the exact date but we recently did a free so should be within the next few weeks I suppose)

[CuzickA]

Thanks @Antonialock, I'll try and remember to check back in a few weeks ;-)

[Antonialock]

It was meant to say "we recently did a freeze"!

[CuzickA]

Note: This session still needs curation/re-curation/checking.