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G2P.pm
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G2P.pm
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=head1 LICENSE
Copyright [1999-2015] Wellcome Trust Sanger Institute and the EMBL-European Bioinformatics Institute
Copyright [2016-2024] EMBL-European Bioinformatics Institute
Licensed under the Apache License, Version 2.0 (the "License");
you may not use this file except in compliance with the License.
You may obtain a copy of the License at
http://www.apache.org/licenses/LICENSE-2.0
Unless required by applicable law or agreed to in writing, software
distributed under the License is distributed on an "AS IS" BASIS,
WITHOUT WARRANTIES OR CONDITIONS OF ANY KIND, either express or implied.
See the License for the specific language governing permissions and
limitations under the License.
=head1 CONTACT
Ensembl <https://www.ensembl.org/info/about/contact/index.html>
=cut
=head1 NAME
G2P
=head1 SYNOPSIS
mv G2P.pm ~/.vep/Plugins
./vep -i variations.vcf --plugin G2P,file=/path/to/G2P.csv
=head1 DESCRIPTION
A VEP plugin that uses G2P allelic requirements to assess variants in genes
for potential phenotype involvement.
The plugin has multiple configuration options, though minimally requires only
the CSV file of G2P data.
For further information see:
Thormann A, Halachev M, McLaren W, et al. Flexible and scalable diagnostic filtering of genomic variants using G2P with Ensembl VEP.
Nature Communications. 2019 May;10(1):2373. doi:10.1038/s41467-019-10016-3. PMID: 31147538; PMCID: PMC6542828.
Options are passed to the plugin as key=value pairs, (defaults in parentheses):
file : Path to G2P data file. The file needs to be uncompressed.
- Download from https://www.ebi.ac.uk/gene2phenotype/downloads
- Download from PanelApp
variant_include_list : A list of variants to include even if variants do not pass allele
frequency filtering. The include list needs to be a sorted, bgzipped and
tabixed VCF file.
af_monoallelic : maximum allele frequency for inclusion for monoallelic genes (0.0001)
af_biallelic : maximum allele frequency for inclusion for biallelic genes (0.005)
confidence_levels : Confidence levels include: definitive, strong, moderate, limited
Former confidence terms are still supported: confirmed, probable, possible, both RD and IF.
Separate multiple values with '&'.
https://www.ebi.ac.uk/gene2phenotype/terminology
Default levels are confirmed and probable.
all_confidence_levels : Set to 1 to include all confidence levels
Setting the value to 1 will overwrite any confidence levels provided with the
confidence_levels option.
af_from_vcf : set value to 1 to include allele frequencies from VCF file.
Specifiy the list of reference populations to include with '--af_from_vcf_keys'
af_from_vcf_keys : VCF collections used for annotating variant alleles with observed
allele frequencies. Allele frequencies are retrieved from VCF files. If
af_from_vcf is set to 1 but no VCF collections are specified with '--af_from_vcf_keys'
all available VCF collections are included.
Available VCF collections: 'topmed', 'uk10k', 'gnomADe', 'gnomADe_r2.1.1', 'gnomADg', 'gnomADg_v3.1.2'.
Separate multiple values with '&'.
VCF collections contain the following populations:
* 'topmed' - TOPMed (available for GRCh37 and GRCh38).
* 'uk10k' - ALSPAC, TWINSUK (available for GRCh37 and GRCh38).
* 'gnomADe' & 'gnomADe_r2.1.1' - gnomADe:AFR, gnomADe:ALL, gnomADe:AMR, gnomADe:ASJ, gnomADe:EAS, gnomADe:FIN, gnomADe:NFE, gnomADe:OTH, gnomADe:SAS (for GRCh37 and GRCh38 respectively).
* 'gnomADg' & 'gnomADg_v3.1.2' - gnomADg:AFR, gnomADg:ALL, gnomADg:AMR, gnomADg:ASJ, gnomADg:EAS, gnomADg:FIN, gnomADg:NFE, gnomADg:OTH (for GRCh37 and GRCh38 respectively).
Need to use 'af_from_vcf' parameter to use this option.
default_af : default frequency of the input variant if no frequency data is
found (0). This determines whether such variants are included;
the value of 0 forces variants with no frequency data to be
included as this is considered equivalent to having a frequency
of 0. Set to 1 (or any value higher than 'af') to exclude them.
types : SO consequence types to include. Separate multiple values with '&'
(splice_donor_variant, splice_acceptor_variant, stop_gained,
frameshift_variant, stop_lost, initiator_codon_variant,
inframe_insertion, inframe_deletion,missense_variant,
coding_sequence_variant, start_lost,transcript_ablation,
transcript_amplification, protein_altering_variant)
log_dir : write stats to log files in log_dir
txt_report : write all G2P complete genes and attributes to txt file
html_report : write all G2P complete genes and attributes to html file
filter_by_gene_symbol : set to 1 if filter by gene symbol.
Do not set if filtering by HGNC_id.
This option is set to 1 when using PanelApp files.
only_mane : set to 1 to ignore transcripts that are not MANE
N/B - Information may be lost if this option is used.
For more information - https://www.ebi.ac.uk/gene2phenotype/g2p_vep_plugin
Example:
--plugin G2P,file=G2P.csv,af_monoallelic=0.05,types=stop_gained&frameshift_variant
--plugin G2P,file=G2P.csv,af_monoallelic=0.05,af_from_vcf=1
--plugin G2P,file=G2P.csv,af_from_vcf=1,af_from_vcf_keys='topmed&gnomADe_r2.1.1'
--plugin G2P,file=G2P.csv,af_from_vcf=1,af_from_vcf_keys='topmed&gnomADe_r2.1.1',confidence_levels='confirmed&probable&both RD and IF'
--plugin G2P,file=G2P.csv
=cut
package G2P;
use strict;
use warnings;
use Cwd;
use Scalar::Util qw(looks_like_number);
use FileHandle;
use Text::CSV;
use Bio::EnsEMBL::Utils::Sequence qw(reverse_comp);
use Bio::EnsEMBL::Variation::Utils::Sequence qw(get_matched_variant_alleles);
use Bio::EnsEMBL::Variation::Utils::VEP qw(parse_line);
use Bio::EnsEMBL::Variation::DBSQL::VCFCollectionAdaptor;
use Bio::EnsEMBL::Variation::Utils::BaseVepPlugin;
use base qw(Bio::EnsEMBL::Variation::Utils::BaseVepTabixPlugin);
use List::Util qw(any);
our $CAN_USE_HTS_PM;
BEGIN {
if (eval { require Bio::DB::HTS::Tabix; 1 }) {
$CAN_USE_HTS_PM = 1;
}
}
my %DEFAULTS = (
# vars must have a frequency <= to this to pass
af_monoallelic => 0.0001,
af_biallelic => 0.005,
af_keys => [qw(AA AFR AMR EA EAS EUR SAS gnomAD gnomAD_AFR gnomAD_AMR gnomAD_ASJ gnomAD_EAS gnomAD_FIN gnomAD_NFE gnomAD_OTH gnomAD_SAS
gnomADg gnomADg_AFR gnomADg_AMR gnomADg_ASJ gnomADg_EAS gnomADg_FIN gnomADg_NFE gnomADg_OTH gnomADg_SAS
gnomADe gnomADe_AFR gnomADe_AMR gnomADe_ASJ gnomADe_EAS gnomADe_FIN gnomADe_NFE gnomADe_OTH gnomADe_SAS)],
af_from_vcf_keys => [qw(uk10k topmed gnomADe gnomADe_r2.1.1 gnomADg gnomADg_v3.1.2)],
# if no MAF data is found, default to 0
# this means absence of MAF data is considered equivalent to MAF=0
# set to 1 to do the "opposite", i.e. exclude variants with no MAF data
default_af => 0,
# adding new confidence levels based on the new terminology
confidence_levels => [qw(confirmed probable definitive strong moderate)],
# only include variants with these consequence types
# currently not ontology-resolved, exact term matches only
types => {map {$_ => 1} qw(splice_donor_variant splice_acceptor_variant stop_gained frameshift_variant stop_lost initiator_codon_variant inframe_insertion inframe_deletion missense_variant coding_sequence_variant start_lost transcript_ablation transcript_amplification protein_altering_variant)},
);
my $af_key_2_population_name = {
minor_allele_freq => 'global allele frequency (AF) from 1000 Genomes Phase 3 data',
AFR => '1000GENOMES:phase_3:AFR',
AMR => '1000GENOMES:phase_3:AMR',
EAS => '1000GENOMES:phase_3:EAS',
EUR => '1000GENOMES:phase_3:EUR',
SAS => '1000GENOMES:phase_3:SAS',
AA => 'Exome Sequencing Project 6500:African_American',
EA => 'Exome Sequencing Project 6500:European_American',
gnomAD => 'Genome Aggregation Database:Total',
gnomAD_AFR => 'Genome Aggregation Database exomes v2.1:African/African American',
gnomAD_AMR => 'Genome Aggregation Database exomes v2.1:Latino/Admixed American',
gnomAD_ASJ => 'Genome Aggregation Database exomes v2.1:Ashkenazi Jewish',
gnomAD_EAS => 'Genome Aggregation Database exomes v2.1:East Asian',
gnomAD_FIN => 'Genome Aggregation Database exomes v2.1:Finnish',
gnomAD_NFE => 'Genome Aggregation Database exomes v2.1:Non-Finnish European',
gnomAD_OTH => 'Genome Aggregation Database exomes v2.1:Other (population not assigned)',
gnomAD_SAS => 'Genome Aggregation Database exomes v2.1:South Asian',
};
my $allelic_requirements = {
'biallelic' => { af => 0.005, rules => {HET => 2, HOM => 1} },
'biallelic_autosomal' => { af => 0.005, rules => {HET => 2, HOM => 1} },
'biallelic_PAR' => { af => 0.005, rules => {HET => 2, HOM => 1} },
'monoallelic' => { af => 0.0001, rules => {HET => 1, HOM => 1} },
'monoallelic_autosomal' => => { af => 0.0001, rules => {HET => 1, HOM => 1} },
'hemizygous' => { af => 0.0001, rules => {HET => 1, HOM => 1} },
'monoallelic_X_hem' => { af => 0.0001, rules => {HET => 1, HOM => 1} },
'monoallelic_Y_hem' => { af => 0.0001, rules => {HET => 1, HOM => 1} },
'x-linked dominant' => { af => 0.0001, rules => {HET => 1, HOM => 1} },
'monoallelic_X_het' => { af => 0.0001, rules => {HET => 1, HOM => 1} },
'x-linked over-dominance' => { af => 0.0001, rules => {HET => 1, HOM => 1} },
'mitochondrial' => { af => 0.0001, rules => {HET => 1, HOM => 1} },
'monoallelic_PAR' => { af => 0.0001, rules => {HET => 1, HOM => 1} },
};
my $supported_confidence_levels = {
'confirmed' => 1,
'definitive' => 1,
'probable' => 1,
'strong' => 1,
'moderate' => 1,
'possible' => 1,
'limited' => 1,
'both RD and IF' => 1,
};
my @allelic_requirement_terms = keys %$allelic_requirements;
# keys containing the assembly and the key to do a quick key and assembly lookup
my $afvcf_keys = {
"uk10k_GRCh37" => 1,
"uk10k_GRCh38" => 1,
"topmed_GRCh37" => 1,
"topmed_GRCh38" => 1,
"gnomADe_GRCh37" => 1,
"gnomADe_r2.1.1_GRCh38" => 1,
"gnomADg_GRCh37" => 1,
"gnomADg_v3.1.2_GRCh38" => 1
};
my $a_keys = {
"uk10k" => 1,
"topmed" => 1,
"gnomADe" => 1,
"gnomADe_r2.1.1" => 1,
"gnomADg" => 1,
"gnomADg_v3.1.2" => 1
};
sub new {
my $class = shift;
my $self = $class->SUPER::new(@_);
# suppress warnings that the FeatureAdpators spit if using no_slice_cache
Bio::EnsEMBL::Utils::Exception::verbose(1999);
my $params = $self->params_to_hash();
my $file = '';
# user only supplied file as first param?
if (!keys %$params) {
$file = $self->params->[0];
$params->{file} = $file;
}
else {
$file = $params->{file};
open IN, "<", $file;
# supporting panelapp by always filtering by gene symbol if PanelApp file
while (<IN>){
$params->{filter_by_gene_symbol} = 1 if (/Model_Of_Inheritance/);
last;
}
close $file;
# process types
if ($params->{types}) {
$params->{types} = {map {$_ => 1} split(/[\;\&\|]/, $params->{types})};
}
# check af
foreach my $af (qw/af_monoallelic af_biallelic/) {
if($params->{$af}) {
die("ERROR: Invalid value for af: ".$params->{$af} . "\n") unless
looks_like_number($params->{$af}) && ($params->{$af} >= 0 && $params->{$af} <= 1)
}
my $ar = $af;
$ar =~ s/af_//;
$allelic_requirements->{$ar}->{af} = $params->{$af} if (defined $params->{$af});
}
$params->{af_keys} = \@{$DEFAULTS{af_keys}};
}
my ($sec, $min, $hour, $mday, $mon, $year, $wday, $yday, $isdst) = localtime(time);
$year += 1900;
$mon++;
my $stamp = join('_', ($year, $mon, $mday, $hour, $min, $sec));
my $cwd_dir = getcwd;
my $new_log_dir = "$cwd_dir/g2p_log_dir\_$stamp";
my $log_dir = $params->{log_dir} || $new_log_dir;
if (!-d $log_dir) {
my $return = mkdir $log_dir, 0755;
die("ERROR: Couldn't create log_dir $log_dir $!\n") if (!$return);
$params->{log_dir} = $log_dir;
}
else{
opendir my $dh, $log_dir or die("ERROR: There was a problem opening the log_dir: $!\n");
my @check = grep {$_ ne '.' and $_ ne '..'} readdir $dh;
die("ERROR: The log directory ($log_dir) is not empty. You need to empty directory before using the plugin \n") if (scalar @check != 0);
closedir $dh;
$params->{log_dir} = $log_dir;
}
foreach my $report_type (qw/txt_report html_report/) {
if (!$params->{$report_type}) {
my $file_type = ($report_type eq 'txt_report') ? 'txt' : 'html';
$params->{$report_type} = $cwd_dir . "/$report_type\_$stamp.$file_type";
}
}
if ($params->{all_confidence_levels}) {
if ($params->{confidence_levels}) {
warn("Option all_confidence_levels set to 1 overwrites confidence levels provided with confidence_levels option.");
}
$params->{confidence_levels} = ['possible', @{$DEFAULTS{confidence_levels}}];
}
elsif ($params->{confidence_levels}) {
my @confidence_levels = ();
foreach my $confidence_level (split(/[\;\&\|]/, $params->{confidence_levels})) {
if (!$supported_confidence_levels->{$confidence_level}) {
die "$confidence_level is not a supported value for supported confidence levels. Supported values are: ", join(', ', keys %$supported_confidence_levels);
} else {
push @confidence_levels, $confidence_level;
push @confidence_levels, 'both DD and IF' if ($confidence_level eq 'both RD and IF'); # legacy support for using both DD and IF
}
}
if (scalar @confidence_levels > 0) {
$params->{confidence_levels} = \@confidence_levels;
}
}
if ($params->{af_from_vcf}) {
if ($CAN_USE_HTS_PM) {
my @vcf_collection_ids = ();
# adding a die if assembly is not used and af_from_vcf keys option is used
my $assembly = $self->{config}->{assembly};
die "Assembly needs to be defined to use af_from_vcf option" if (!defined ($assembly));
if ($params->{af_from_vcf_keys}) {
foreach my $key (split(/[\;\&\|]/, $params->{af_from_vcf_keys})) {
my $key_assembly = $key."_".$assembly;
if (!$afvcf_keys->{$key_assembly}){
# to die if key is not supported, checking with the key and the assembly
die "$key is not a supported key. Supported keys and assembly are: ", join(',', keys %$a_keys), ".\n
gnomADe and gnomADg is supported for assembly GRCh37 \n
gnomADe_r2.1.1 and gnomADg_v3.1.2 is supported for assembly GRCh38 \n" ;
}
else {
push @vcf_collection_ids, $key;
push @vcf_collection_ids, $key_assembly;
}
}
} else {
foreach my $key (@{$DEFAULTS{af_from_vcf_keys}}) {
push @vcf_collection_ids, $key;
push @vcf_collection_ids, "$key\_$assembly";
}
}
my $species = $self->{config}->{species};
my $reg = $self->{config}->{reg};
my $vca;
if (defined $self->{config}->{offline}) {
$vca = Bio::EnsEMBL::Variation::DBSQL::VCFCollectionAdaptor->new();
} else {
my $vdba = $reg->get_DBAdaptor($species, 'variation');
$vdba->dbc->reconnect_when_lost(1);
$vca = $vdba->get_VCFCollectionAdaptor;
$vca->db->use_vcf(2);
}
my $vcf_collections = $vca->fetch_all;
my @collections = ();
foreach my $vcf_collection (@$vcf_collections) {
$vcf_collection->use_db(0) if (defined $self->{config}->{offline});
my $vcf_collection_id = $vcf_collection->id;
if ($vcf_collection->assembly eq $assembly && grep {$_ =~ /$vcf_collection_id/i} @vcf_collection_ids) {
delete $vcf_collection->adaptor->{collections};
delete $vcf_collection->adaptor->{config};
my $description = $vcf_collection->description || $vcf_collection_id;
foreach my $population (@{$vcf_collection->get_all_Populations}) {
my $population_name = $population->name;
my $population_description = $population->description;
$af_key_2_population_name->{$population_name} = "$description $population_name $population_description";
}
push @collections, $vcf_collection;
}
}
warn "Couldn't find VCF collection ids for assembly " . $assembly if (!@collections);
$self->{config}->{vcf_collections} = \@collections;
$self->{config}->{use_vcf} = 1;
} else {
warn "Cannot get data from VCF without Bio::DB::HTS::Tabix";
}
}
if ($params->{variant_include_list}) {
if (! -f $params->{variant_include_list}) {
die "Variant include list (" . $params->{variant_include_list} . ") does not exist.";
}
$self->{_files} = [$params->{variant_include_list}];
}
if (defined($params->{filter_by_gene_symbol}) and $params->{filter_by_gene_symbol} != 1) {
$params->{filter_by_gene_symbol} = undef;
die "The option --filter_by_gene_symbol needs to be set to 1 \n";
}
if (defined($params->{only_mane}) and $params->{only_mane} != 1) {
$params->{only_mane} = undef;
die "The option only_mane needs to be set to 1 \n";
}
if (defined($params->{only_mane}) and $self->{config}->{assembly} ne "GRCh38") {
die "The option only_mane only works with GRCh38 assembly \n";
}
# copy in default params
$params->{$_} //= $DEFAULTS{$_} for keys %DEFAULTS;
$self->{user_params} = $params;
$self->{config}->{frequency_threshold} = _get_highest_frequency_threshold();
# read data from file
$self->{gene_data} = $self->read_gene_data_from_file($file);
$self->synonym_mappings();
$self->hgnc_mappings();
# force some config params
$self->{config}->{individual} //= ['all'];
$self->{config}->{symbol} = 1;
$self->{config}->{check_existing} = 1;
$self->{config}->{failed} = 1;
$self->{config}->{af} = 1;
$self->{config}->{af_1kg} = 1;
$self->{config}->{af_gnomad} = 1;
$self->{config}->{sift} = 'b';
$self->{config}->{polyphen} = 'b';
# tell VEP we have a cache so stuff gets shared/merged between forks
$self->{has_cache} = 1;
$self->{cache}->{g2p_in_vcf} = {};
return $self;
}
=head2 _get_highest_frequency_threshold
Description: Retrieve the highest allele frequency threshold across all defined allelic requirements.
This will speed up the filtering by allele frequency. As soon as we found a frequency
higher than this highest frequency the filtering fails and we don't need to consider
the variant further.
Returntype : Float $highest_frequency
Exceptions : None
Caller : General
Status : Stable
=cut
sub _get_highest_frequency_threshold {
my $highest_frequency = 0.0;
foreach my $ar (keys %$allelic_requirements) {
if ($allelic_requirements->{$ar}->{af} > $highest_frequency) {
$highest_frequency = $allelic_requirements->{$ar}->{af};
}
}
return $highest_frequency;
}
sub feature_types {
return ['Transcript'];
}
sub get_header_info {
my $self = shift;
return {
G2P_flag => 'Flags zygosity of valid variants for a G2P gene',
G2P_complete => 'Indicates this variant completes the allelic requirements for a G2P gene',
G2P_gene_req => 'MONO or BI depending on the context in which this gene has been explored',
};
}
=head2 run
Arg [1] : TranscriptVariationAllele $tva
Arg [2] : Hashref $line
Description: Filter input transcript variation allele on:
- G2P gene overlap
- variant consequence
- variant include list
- allele frequency
Based on the filtering results check if the allelic requirement is fulfilled and write results to hash.
Dump annnotations to a log file for generating TXT and HTML output files after VEP has finished.
Returntype : Hashref $results
Exceptions : None
Caller : General
Status : Stable
=cut
sub run {
my ($self, $tva, $line) = @_;
# only interested if we know the zygosity
my $zyg = defined($line->{Extra}) ? $line->{Extra}->{ZYG} : $line->{ZYG};
return {} unless $zyg;
return {} if ($self->{user_params}->{only_mane} && !$tva->transcript->is_mane);
# filter by G2P gene overlap
return {} if (!$self->gene_overlap_filtering($tva));
$self->set_variant_include_list_flag($tva);
# filter by variant consequence
return {} if (!$self->consequence_filtering($tva));
# filter by allele frequency
return {} if (!$self->frequency_filtering($tva));
# dump annotations for txt and html report files
$self->dump_vf_annotations($tva);
$self->dump_individual_annotations($tva, $zyg);
# check if transcript contains enough variants to fulfill the allelic requirement of the gene
my $G2P_complete = $self->is_g2p_complete($tva, $zyg);
my $G2P_flag = $self->is_valid_g2p_variant($tva, $zyg);
my $results = {};
$results->{G2P_complete} = $G2P_complete if ($G2P_complete);
$results->{G2P_flag} = $G2P_flag if ($G2P_flag);
return $results;
}
=head2 set_variant_include_list_flag
Arg [1] : TranscriptVariationAllele $tva
Description: Check if variant is part of the variant include list.
If the variant is on the variant include list then report
it in the result set regardless if the variant passed the
filtering by variant consequence and allele frequency.
Returntype : None
Exceptions : None
Caller : General
Status : Stable
=cut
sub set_variant_include_list_flag {
my $self = shift;
my $tva = shift;
return if (!$self->{user_params}->{variant_include_list});
my $vf = $tva->variation_feature;
my $allele = $tva->base_variation_feature->alt_alleles;
foreach (@{$self->get_data($vf->{chr}, $vf->{start} - 1, $vf->{end})}) {
my @vcf_alleles = split /\//, $_->allele_string;
my $ref_allele = shift @vcf_alleles;
my $matches = get_matched_variant_alleles(
{
ref => $vf->ref_allele_string,
alts => $allele,
pos => $vf->{start},
strand => $vf->strand
},
{
ref => $ref_allele,
alts => \@vcf_alleles,
pos => $_->{start},
}
);
if (scalar @$matches) {
my $vf_cache_name = $self->get_cache_name($vf);
$self->{g2p_vf_cache}->{$vf_cache_name}->{is_on_variant_include_list} = 1;
last;
}
}
}
=head2 is_valid_g2p_variant
Arg [1] : TranscriptVariationAllele $tva
Arg [2] : String $zygosity
Example : $valid_g2p_variant = $self->is_valid_g2p_variant($tva, 'HOM')
Description: Take all allelic requirements of the gene that overlap this variant
and check if the variant passes the frequency threshold filter where the threshold is defined
by the allelic requirement of the gene.
Concatenate results for several allelic requirements by ','.
Returntype : String for example "monoallelic=HOM"
Exceptions : None
Caller : General
Status : Stable
=cut
sub is_valid_g2p_variant {
my $self = shift;
my $tva = shift;
my $zyg = shift;
my $transcript = $tva->transcript;
my $gene_stable_id = get_gene_stable_id($transcript);
my @allelic_requirements = keys %{$self->{ar}->{$gene_stable_id}};
my @results = ();
foreach my $ar (@allelic_requirements) {
my $ar_rules = $allelic_requirements->{$ar};
my $af_threshold = $ar_rules->{af};
my $variants = $self->variants_filtered_by_frequency_threshold($af_threshold, [$self->{vf_cache_name}]);
if (scalar @$variants > 0) {
push @results, "$ar=$zyg";
}
}
return join(',', @results);
}
=head2 is_g2p_complete
Arg [1] : TranscriptVariationAllele $tva
Arg [2] : String $zygosity
Description: A G2P gene is considered complete if its allelic requirement is fulfilled.
Create a summary string which connects the fulfilled allelic_requirement and all variants which pass filtering.
Returntype : String $g2p_complete, for example "monoallelic=HET:7_941481_C/T&HET:7_929274_A/G,HOM:7_931481_C/T|biallelic=HOM:7_931481_C/T"
Exceptions : None
Caller : General
Status : Stable
=cut
sub is_g2p_complete {
my $self = shift;
my $tva = shift;
my $zyg = shift;
my $vf = $tva->base_variation_feature;
my $individual = $vf->{individual};
my $transcript = $tva->transcript;
my $gene_stable_id = get_gene_stable_id($transcript);
my $transcript_stable_id = $transcript->stable_id;
$self->{per_individual}->{$individual}->{$transcript_stable_id}->{$zyg}->{$self->{vf_cache_name}} = 1;
my @allelic_requirements = keys %{$self->{ar}->{$gene_stable_id}};
my @g2p_complete = ();
foreach my $ar (@allelic_requirements) {
my $zyg2var = $self->{per_individual}->{$individual}->{$transcript_stable_id};
my $filtered_zyg2var = $self->zyg2var_filtered_by_allelic_requirement_rule($ar, $zyg2var);
if (defined $filtered_zyg2var) {
my @filtered_variants = ();
foreach my $zyg (keys %$filtered_zyg2var) {
push @filtered_variants, join('&', map {"$zyg:$_"} @{$filtered_zyg2var->{$zyg}});
}
push @g2p_complete, "$ar=" . join(',', @filtered_variants);
}
}
return join('\|', @g2p_complete);
}
=head2 zyg2var_filtered_by_allelic_requirement_rule
Arg [1] : String $ar allelic requirement
Arg [2] : Hashref $zyg2var
Example : $zyg2var_filtered = $self->zyg2var_filtered_by_allelic_requirement_rule('biallelic', {
'HET' => {
'4_32941481_C/T' => 1,
'4_32929274_A/G' => 1
}
});
Description: Check if the variants fulfil the given allelic requirement. Variants are grouped by their
zygosity. The subroutine checks for each variant if the internally stored frequency for a variant
is lower than the allele frequency threshold defined by the allelic requirement. Then consider
the variants which pass frequency filtering and check if the number is sufficient to fulfil the
allelic requirement.
Returntype : Hashref $zyg2var_filtered: with zygosity as key and filtered variants in arrayref as value
For example: {
'HET' => [
'4_32941481_C/T',
'4_32929274_A/G'
]
};
Undef, if none of the variants pass the filtering
Exceptions : None
Caller : General
Status : Stable
=cut
sub zyg2var_filtered_by_allelic_requirement_rule {
my $self = shift;
my $ar = shift;
my $zyg2variants = shift;
my $ar_rules = $allelic_requirements->{$ar};
# allele frequency threshold for given allelic requirement
my $af_threshold = $ar_rules->{af};
# number of variants with a particular zygosity that need to pass filtering
# to fulfil allelic requirement
my $zyg2counts = $ar_rules->{rules};
my $results = {};
foreach my $zyg (keys %$zyg2counts) {
my $count = $zyg2counts->{$zyg};
my @all_variants = keys %{$zyg2variants->{$zyg}};
my $variants = $self->variants_filtered_by_frequency_threshold($af_threshold, \@all_variants);
if (scalar @$variants >= $count) {
$results->{$zyg} = $variants;
}
}
if (scalar keys %$results > 0) {
return $results;
} else {
return undef;
}
}
=head2 variants_filtered_by_frequency_threshold
Arg [1] : Float $af_threshold allele frequency threshold
Arg [2] : Arrayref $variants
Example : $variants_filtered = $self->variants_filtered_by_frequency_threshold(0.001, [
'4_32941481_C/T' => 1,
'4_32929274_A/G' => 1
]
);
Description: Check for each variant if the highest frequency that has been observed in any
population (stored internally under the highest_frequencies hash key for a variant) is lower
than the given allele frequency threshold. If yes, add the variant to the result set.
Also add the variant to the result set if the variant hasn't any observed allele frequencies
or if the variant is in the variant include list.
Returntype : Arrayref $variants_filtered
For example: [
'4_32941481_C/T' => 1,
'4_32929274_A/G' => 1
];
Exceptions : None
Caller : General
Status : Stable
=cut
sub variants_filtered_by_frequency_threshold {
my $self = shift;
my $af_threshold = shift;
my $variants = shift;
my @pass_variants = ();
foreach my $variant (@$variants) {
# get the highest frequency that has been observed for the variant
my $highest_frequency = $self->highest_frequency($variant);
if (! defined $highest_frequency || (defined $af_threshold && $highest_frequency <= $af_threshold) ||
$self->{g2p_vf_cache}->{$variant}->{is_on_variant_include_list}
) {
push @pass_variants, $variant;
}
}
return \@pass_variants;
}
=head2 gene_overlap_filtering
Arg [1] : TranscriptVariationAllele $tva
Example :
Description: returns 1 or 0 depending on if the gene is part of the input panel. If the gene is part of the panel we store
the allelic requirement in the internal cash under the name 'ar'. We also write G2P_gene_data and G2P_in_vcf
information to the log file. We call _dump_transcript_annotations and write transcript information to the log file.
Returntype : Boolean
Exceptions : None
Caller : run
Status : Stable
=cut
sub gene_overlap_filtering {
my $self = shift;
my $tva = shift;
my $transcript = $tva->transcript;
my $gene_stable_id = get_gene_stable_id($transcript);
my $pass_gene_overlap_filter = $self->{g2p_gene_cache}->{$gene_stable_id};
my @gene_xrefs = ();
if (! defined $pass_gene_overlap_filter && ! defined $self->{user_params}->{filter_by_gene_symbol}) {
my @gene_hgnc = split /:/, $transcript->{_gene_hgnc_id} if (defined $transcript->{_gene_hgnc_id});
my $hgnc_id = $gene_hgnc[1] if (@gene_hgnc);
if (defined $hgnc_id) {
foreach my $id (keys %{ $self->{hgnc_mapping} }) {
if ( $hgnc_id == $id) {
my $gene_symbol = $self->{hgnc_mapping}{$id};
my $gene_data = $self->gene_data($gene_symbol) if defined ($gene_symbol);
if (defined $gene_data) {
if (defined $gene_data->{'allelic requirement'} && scalar @{$gene_data->{'allelic requirement'}}) {
foreach my $ar (@{$gene_data->{'allelic requirement'}}) {
$self->{ar}->{$gene_stable_id}->{$ar} = 1;
}
$self->write_report('G2P_gene_data', $gene_stable_id, $gene_data, $gene_data->{'gene_xrefs'}, $gene_data->{'HGNC'}, $gene_data->{'confidence_category'}, $gene_data->{'confidence_value'} );
}
$self->write_report('G2P_in_vcf', $gene_stable_id);
$pass_gene_overlap_filter = 1;
last;
}
}
}
}
$self->{g2p_gene_cache}->{$gene_stable_id} = $pass_gene_overlap_filter;
}
if (! defined $pass_gene_overlap_filter && defined $self->{user_params}->{filter_by_gene_symbol} == 1) {
my $gene_symbol = $transcript->{_gene_symbol} || $transcript->{_gene_hgnc};
$pass_gene_overlap_filter = 0;
foreach my $gene_id ($gene_symbol, $gene_stable_id) {
my $gene_data = $self->gene_data($gene_id) if (defined $gene_id);
if (defined $gene_data) {
if (defined $gene_data->{'allelic requirement'} && scalar @{$gene_data->{'allelic requirement'}}) {
foreach my $ar (@{$gene_data->{'allelic requirement'}}) {
$self->{ar}->{$gene_stable_id}->{$ar} = 1;
}
$self->write_report('G2P_gene_data', $gene_stable_id, $gene_data, $gene_data->{'gene_xrefs'}, undef, $gene_data->{'confidence_category'}, $gene_data->{'confidence_value'});
}
$self->write_report('G2P_in_vcf', $gene_stable_id);
$pass_gene_overlap_filter = 1;
last;
}
}
$self->{g2p_gene_cache}->{$gene_stable_id} = $pass_gene_overlap_filter;
}
$self->_dump_transcript_annotations($transcript) if ($pass_gene_overlap_filter);
return $self->{g2p_gene_cache}->{$gene_stable_id};
}
=head2 consequence_filtering
Arg [1] : TranscriptVariationAllele $tva
Description: returns 1 or 0 depending on if the variant passes consequence filtering.
If the variant is on the variant include list we return 1, regardless
if the variant consequence is defined in types.
Returntype : Boolean
Exceptions : None
Caller : General
Status : Stable
=cut
sub consequence_filtering {
my $self = shift;
my $tva = shift;
my $vf = $tva->base_variation_feature;
my $vf_cache_name = $self->get_cache_name($vf);
return ((grep { $self->{user_params}->{types}->{$_->SO_term} } @{$tva->get_all_OverlapConsequences}) ||
$self->{g2p_vf_cache}->{$vf_cache_name}->{is_on_variant_include_list});
}
=head2 _dump_transcript_annotations
Arg [1] : Transcript $transcript
Description: Write G2P_transcript_data to log file.
Returntype : None
Exceptions : None
Caller : General
Status : Stable
=cut
sub _dump_transcript_annotations {
my $self = shift;
my $transcript = shift;
my $transcript_stable_id = $transcript->stable_id;
if (!defined $self->{g2p_transcript_cache}->{$transcript_stable_id}) {
my $gene_stable_id = get_gene_stable_id($transcript);
if ($transcript->is_canonical) {
$self->write_report('G2P_transcript_data', "$gene_stable_id\t$transcript_stable_id\tis_canonical");
}
$self->{g2p_transcript_cache}->{$transcript_stable_id} = 1;
}
}
=head2 get_cache_name
Arg [1] : VariationFeature $vf
Description: Create a variant identifier for internal use which is stored in the internal cache
Returntype : String $vf_cache_name for example 19_8412862_G/A
Exceptions : None
Caller : General
Status : Stable
=cut
sub get_cache_name {
my $self = shift;
my $vf = shift;
my $cache_name = ($vf->{original_chr} || $vf->{chr}) . '_' . $vf->{start} . '_' . ($vf->{allele_string} || $vf->{class_SO_term});
return $cache_name;
}
=head2 get_gene_stable_id
Arg [1] : Transcript $transcript
Description: Retrives the Ensembl or RefSeq gene stable id from the transcript object
Returntype : String $gene_stable_id
Exceptions : None
Caller : General
Status : Stable
=cut
sub get_gene_stable_id {
my $transcript = shift;
return $transcript->{_gene}->stable_id;
}
=head2 frequency_filtering
Arg [1] : TranscriptVariationAllele $tva
Description: returns 1 or 0 depending on if the variant passes frequency filtering.
We consider allele frequencies from the VEP cache and VCF files for filtering.
Returntype : Boolean
Exceptions : None
Caller : General
Status : Stable
=cut
sub frequency_filtering {
my $self = shift;
my $tva = shift;
my $vf = $tva->base_variation_feature;
# Set up caching to avoid looking up frequencies for each overlapping transcript
my $vf_cache_name = $self->get_cache_name($vf);
$self->{vf_cache_name} = $vf_cache_name;
$self->{g2p_vf_cache} = {} if (!defined $self->{g2p_vf_cache}->{$vf_cache_name});
# Retrieve cached result
my $pass_frequency_filter = $self->{g2p_vf_cache}->{$vf_cache_name}->{pass_frequency_filter};
return $pass_frequency_filter if (defined $pass_frequency_filter);
# Check frequencies from cache files first
$pass_frequency_filter = $self->_vep_cache_frequency_filtering($tva);
# Check frequencies from VCF files if user is providing use_vcf flag
if ($pass_frequency_filter && $self->{config}->{use_vcf}) {
$pass_frequency_filter = $self->_vcf_frequency_filtering($tva);
}
$self->{g2p_vf_cache}->{$vf_cache_name}->{pass_frequency_filter} = $pass_frequency_filter;
return $self->{g2p_vf_cache}->{$vf_cache_name}->{pass_frequency_filter};
}
=head2 _vep_cache_frequency_filtering
Arg [1] : TranscriptVariationAllele $tva
Description: Returns 1 or 0 depending on if the variant passes frequency filtering where allele frequencies come from the VEP cache.
Return 1 if no observed allele frequencies exist for the given variant which means that the filtering passes.
Returntype : Boolean
Exceptions : None
Caller : General
Status : Stable
=cut
sub _vep_cache_frequency_filtering {
my $self = shift;
my $tva = shift;
my $allele = $tva->base_variation_feature->alt_alleles;
my $vf = $tva->base_variation_feature;
my $frequency_threshold = $self->{config}->{frequency_threshold};
my $existing = $vf->{existing}; # Get existing variants from cache file which are stored on VF level
my @keys = @{$self->{user_params}->{af_keys}}; # Consider user defined list of af keys
my $dumped_annotations = 0; # Indicates if existing annotations have already been dumped for txt and html report files
my $vf_cache_name = $self->{vf_cache_name};
foreach my $existing_var (@$existing) {
my @frequencies = grep defined, @{$existing_var}{@keys};
if ($existing_var->{matched_alleles}) { # Get matched alleles from input variant and existing variant, in case input variant was normalized to match variant from cache file
$allele = $existing_var->{matched_alleles}[0]->{b_allele};
}
next if (!@frequencies);
if ($self->_exceeds_frequency_threshold(\@frequencies, $allele, $frequency_threshold) && !$self->{g2p_vf_cache}->{$vf_cache_name}->{is_on_variant_include_list}) {
return 0; # Return 0 (failed filtering) if frequencies exceed threshold and variant is not on variant_include_list
} else {
# Dump annotations for txt and html report files
$self->_dump_existing_vf_frequencies($existing_var, $allele);
$self->_dump_existing_vf_annotations($existing_var);
$dumped_annotations = 1;
}
}
# If we get to this point it means that there were no frequencies for the input variant in the cache files
# and we pass the filtering step.
# We need to dump 'empty' annotations for such variants to indicate that there are no available frequencies
$self->_dump_existing_vf_annotations() if (!$dumped_annotations);
return 1;
}
=head2 _dump_existing_vf_frequencies
Arg [1] : Hashref $existing_var
Description: The $existing_var contains everything that is stored for the variant in the VEP cache file.
Extract allele frequencies from $existing_var and write it to the log file as G2P_frequencies.
Store the highest observed frequency for the variant which is used later for filtering.
Returntype : None
Exceptions : None
Caller : General
Status : Stable
=cut
sub _dump_existing_vf_frequencies {
my $self = shift;
my $existing_var = shift;
my $allele = shift;
my @keys = @{$self->{user_params}->{af_keys}};
my @frequencies = ();
my $higest_frequency = 0;
foreach my $population_name (@keys) {
my $af = $existing_var->{$population_name};
next if (!defined $af);
foreach my $pair (split(',', $af)) {
my ($a, $f) = split(':', $pair);
if(($a || '') eq $allele && defined($f)) {
push @frequencies, "$population_name=$f";
$higest_frequency = $f if ($f > $higest_frequency);
}
}
}
$self->highest_frequency($self->{vf_cache_name}, $higest_frequency);
$self->write_report('G2P_frequencies', $self->{vf_cache_name}, \@frequencies);