NOTES.md

In rformassspectrometry/Features: Quantitative features for mass spectrometry data

One or two levels in QFeatures (closed)

One or two assay levels could be considered in QFeatures:

• one level: each SE contains only a single assay, and when an SE is processed (log-transformed, normalised, ...) in a way that its dimensions stay the same, a new SE is created and added to the QFeatures object.

• two level: SEs can contain multiple assays, and when an SE is processed (log-transformed, normalised, ...) in a way that its dimensions stay the same, a new assay is added to that SE.

This question on the bioc-devel list ask for advice on SE processing, and whether a new SE or new assay in the original SE should be preferred. While the letter is arguably more elegant, and is also used in SingleAssayExperiment pipelines, it doesn't seem to be the case when using SummarizedExperiments.

As for features (or MultiAssayExperiments in general), the two-level approach isn't readily available out-of-the-box, and would require additional developments:

• Every function that operates on an SE of a QFeatures object would need to allow the user to specify which assay to use (and/or by default use the latest one).

• The show,QFeatures method would need to display the number/names of the assays in each SE to make these two levels explicit.

Despite the elegant of the two-level option, it seems that the additional development isn't warranted at this time.

The updateAssay function was originally intended for the two-level approach, i.e. to add an assay to an SE. This is not considered anymore (for now, at least).

There is one exception though. When aggregating features with aggregateFeatures(), a second assay is added, named aggcounts that counts the number of features that were aggregate for each sample and each low-level features.

How to add new assays (closed)

1. Through aggregation with aggregateFeatures.

2. Processing an SE.

This can/could be done explicitly with addAssay

addAssay(cptac, logTransform(cptac[["peptides"]]), name = "peptides_log")
addAssay(cptac, logTransform(cptac[[1]]), name = "peptides_log")

or implicitly

logTransform(cptac, "peptides", name = "peptides_log")
logTransform(cptac, 1, name = "peptides_log")

1. Joining SEs (for example multiple TMT batches) (TODO)

joinAssays(QFeatures, c("pep_batch1", "pep_batch2", "pep_batch3"), name = "peptides")
joinAssays(QFeatures, c(1, 2, 3), name = "peptides")

See below.

QFeatures API

Processing functions

• A processing function that acts on a Feature's assay (typically a SummarizedExperiment or a SingleCellExperiment) such as process(object), returns a new object of the same type.

• A processing function such process(object, i), that acts on a Feautre object takes a second argument i, that can be a vector of indices or names, returns a new object of class QFeatures with its assay(s) i modified according to process(object[[i]]).

• The argument i mustn't be missing, i.e. one shouldn't (at least in general) permit to (blindly) apply some processing on all assays.

Assays

• Assays should have unique rownames (even though this isn't required for SEs). If they aren't, only the first occurence of the name is kept:

hlpsms <- hlpsms[1:5000, ] ## faster

ft1 <- readQFeatures(hlpsms, ecol = 1:10, name = "psms", fname = "Sequence")
sum(rownames(ft1[[1]]) == "ANLPQSFQVDTSk")
ft1 <- aggregateFeatures(ft1, "psms", fcol = "Sequence",
                         name = "peptides", fun = colSums)
sapply(rownames(ft1), anyDuplicated)
ft1

## subsetting still works
ft2 <- subsetByFeature(ft1, "ANLPQSFQVDTSk")
ft2

The underlying reason why this fails is due to matrix subsetting by name when these names aren't unique.

m <- matrix(1:10, ncol = 2)
colnames(m) <- LETTERS[1:2]
rownames(m) <- c("a", letters[1:4])
m
m["a", ]

And of course, this affects SEs ...

se <- SummarizedExperiment(m)
assay(se["a", ])

... and MultiAssayExperiments.

Note that in the example above, "ANLPQSFQVDTSk" is present in both the psms and peptides assays, and the

for (k in setdiff(all_assays_names, leaf_assay_name)) { ... }

loop in .subsetByFeature isn't executed at all. This will need to be investigated. But the behaviour above can be reproduced even when that's not the case. See

hlpsms$Sequence2 <- paste0(hlpsms$Sequence, "2")
ft1 <- readQFeatures(hlpsms, ecol = 1:10, name = "psms", fname = "Sequence2")
...

This could be fixed by switching to indices:

> (i <- which(rownames(m) == "a"))
[1] 1 2
> m[i, ]
  A B
a 1 6
a 2 7
> se[i, ]
class: SummarizedExperiment
dim: 2 2
metadata(0):
assays(1): ''
rownames(2): a a
rowData names(0):
colnames(2): A B
colData names(0):

See issue #91.

Assay links

Currently, we have

• Assay links produces by aggregateFeatures and manually with addAssayLink.

• One-to-one Assay links produced by a processing function such as logTransform or with addAssayLinkOneToOne. These contain "OneToOne" in the fcol slot (issue 42).

• There will be a need for an assay link stemming from combining assays (see below and issue 52).

Joining assays (closed)

To combine assays, we also need 1. relaxed MatchedAssayExperiment constrains (see #46) 2. assay links with multiple parent assays (see #52)

combine,MSnSet,MSnSet does two things, i.e. rbind and cbind. Here, we nedd (at least in a first instance) and have cbind,SummarizedExperiment.

• do we need some constrains requiering identical rownames? cbind,SummarizedExperiment uses the mcols to check whether rows match.
• should unique rows in one assay get NAs in the other one? yes!

We need a join-type of function, that adds NAs at the assay level. To do this, we need to have a union of features before rbinding the assays.

As for rowData, we want to

• keep the mcols that match exactly between assays (ex: PeptideSequence, ProteinAccession, ...)
• remove mcols that differ between assays (ex: PEP, qvalues, charge, rtime, ...)

The row data will be accessible through links between assays anyway.

Naming:

joinAssays(QFeatures, c("pep_batch1", "pep_batch2", "pep_batch3"), name = "peptides")
joinAssays(QFeatures, c(1, 2, 3), name = "peptides")

Algorithm: 1. Find which mcols to keep 2. Extend with rownames and NAs (depending on type of join) 3. Order assays 4. cbind assays (see cbind,SummarizedExperiment)

Do we want a public join for SummarizedExperiments? Discuss with SE maintainers.

Note: if we were to have assay from multiple fractions to be rbinded, we could consider a rbindAssays, mergeFractions, bindFractions, ...

Replacing vs adding assays

Issues https://github.com/rformassspectrometry/QFeatures/issues/193 and https://github.com/rformassspectrometry/QFeatures/issues/186.

Currently, assays are replaced with - filterNA() - filterFeatures() (and possibly others)

Sometimes, we want to add, rather than replace, for example if we want to test/assess the effect of different filters. This could be defined by the names argument. If missing (default), the assays are replaced. If present and of same length than i, new assays are added.

• When it comes to data processing, we could also have a subset argument, that would implicitly only process a subset of rows so as to avoid to explicitly store the subset/intermediate assay.

• A more radical change would be for filterFeatures() to add a rowData logical that defines the rows to be filtered.

There are multiple ideas/discussion replated to QFeatures becoming very large (and slow). Rather than adding more assays, we could: - use logical for subsetting; - use multiple assays within a SingleCellExperiment (or SE), when the dimensions remain identical (for exmple logTransform()); - have a unique database to handle and manage all data (assays and rowData).

But we agree that the interface, for the user, should remain simple, i.e. different assays. For now, keep the same philosophy and create new assays for all operations, and start a reflexion for more in-depth refactoring.

See also HDF5 backend issue.

Devel roadmap

scp

• scp data import documentation. Current approach doesn't work well for LFQ. Add documentation, and align scpGUIImport.
• Compute mertics per cell is clumsy/ad hoc at the moment. We need a general function to iterates over cells/features and store automatically in col/rowData. What is computed defined by a user-defined function. For example *OverSample() and *OverFeatures() and a more general computeMetric(). These would added to QFeatures with specialised metrics implemented in scp.
• Merge scplainer into scp (issue 45): needs unit tests
• Missing data vignette/report: unit tests coming soon
• readScpFromDIANN() needs unit test and some refactoring
• divideByReference() is a batch correction method, and should be moved to QFeatures, and made accessible as part of a more general batchCorrect() interface, that can also be used for combat and limma.

scpdata

• Keep adding new datasets.

QFeatures

• All QFeatures functions should add assays - see 'Replacing vs adding assays' above.
• filterNA() on columns (see https://github.com/rformassspectrometry/QFeatures/issues/173]. Or should we use computeMetricOverCells() with metric nNA?
• Small bugs: nNA() on empty assays and longFormat(se, rowvars) (eventhough the latter should probbaly use tidy versions).
• Interest in pseudo-bulking, which should work out-of-the-box, since the colData is unique for all assaya (Chris to address during post-doc).

Tabular input (issue 199)

1. Single-set case, multiplexed: requires colAnnotation only. Also LF with a re-ordered peptide/protein-level table (runs are missing in this case).

|------+------------+-----------|
| cols | Quant 1..N | more cols |
|      |            |           |
|      |            |           |
|      |            |           |
|------+------------+-----------|

readQFeatures(hlpsms, quantCols = 1:10)
readQFeatures(hlpsms, colAnnotation = colann)

## also possible, but redundant
readQFeatures(hlpsms, colAnnotation = colann, quantCols = 1:10)

1. Multi-set case, multiplexed: requires colAnnotation and runCol.

|-----+------+------------+-----------|
| Run | cols | Quant 1..N | more cols |
|   1 |      |            |           |
|   1 |      |            |           |
|-----+------+------------+-----------|
|   2 |      |            |           |
|-----+------+------------+-----------|

readQFeatures(hlpsms, quantCols = 1:10, runCol = "file")
readQFeatures(hlpsms, colAnnotation = colann, runCol = "file")

1. Multi-set case, LF: requires colData and runCol with a optional multiplexing (for plexDIA).

|-----+------+---------+-----------+-----------|
| Run | cols | Quant 1 | more cols | multiplex |
|   1 |      |         |           |           |
|   1 |      |         |           |           |
|-----+------+---------+-----------+-----------|
|   2 |      |         |           |           |
|-----+------+---------+-----------+-----------|

1. Special case DIANN. A specialised function that parses the table to case 2.

Users can either use the arguments above or a colAnnotation data.frame (that will become the colData).

DIANN data

• DIANN data is in long format
• readQFeaturesFromDIANN() would need to transform from long to wide, then use readQFeatures()
• multiplexing: NULL or "none"

dfr |>
  diannWider() |>
  readQFeatures()

readQFeaturesFromDIANN <- funtion(dfr, multiplexing = NULL, ...) {
    if (!is.null(multiplexing))
        x <- .diannWider(multiplexing)
    readQFeatures(x, ...)
}

rformassspectrometry/Features documentation built on Sept. 25, 2024, 11:30 a.m.