#Parse Command Line

parser = optparse.OptionParser()

#I/O

parser.add_option( '-x', '--reference_xml', dest='reference_xml', help='Reference protein UniProt-XML file. Sequence variant peptide entries are appended to this database to generate the ouptut UniProt-XML protein database.' )

parser.add_option( '-p', '--protein_fasta', dest='protein_fasta', help='Reference protein FASTA file. Used to generate SAV peptide entries. If no UniProt-XML is specified, SAV and NSJ entries will be appended to this database to generate an output database. By default, this output will be a UniProt-XML protein database without PTM annotations. If --output-fasta is selected, the output will be a protein FASTA.')

parser.add_option( '-g', '--gene_model', dest='gene_model', default=None, help='GTF gene model file. Used to annotate NSJ peptide entries.')

parser.add_option( '-v', '--snpeff_vcf', dest='snpeff_vcf', help='SnpEff VCF file with HGVS annotations (else read from stdin).' )

parser.add_option( '-c', '--cufflinks_bed', dest='cufflinks_bed', help='Reconstructed transcripts in bed format.' )

parser.add_option( '-b', '--splice_bed', dest='splice_bed', help='BED file (tophat junctions.bed) with sequence column added.' )

parser.add_option( '-o', '--output', dest='output', help='Output file path. Outputs UniProt-XML and FASTA format databases.' )

# parser.add_option( '-z', '--output_fasta', dest='output_fasta', action='store_true', default=False, help='Output a FASTA-format database. Place path for output file after the --output flag.')

# parser.add_option( '-m', '--threads', dest='threads', type='int', default=)

#Peptide sequence construction

parser.add_option( '-l', '--leading_aa_num', dest='leading_aa_num', type='int', default=33, help='Leading number of AAs to output for SAV peptides. Default: 33.' )

parser.add_option( '-t', '--trailing_aa_num', dest='trailing_aa_num', type='int', default=33, help='Trailing number of AAs to output for SAV peptides. Default: 33.' )

#Filtering parameters

parser.add_option( '-D', '--nsj_depth_cutoff', dest='nsj_depth_cutoff', type='int', default=0, help='Keep only NSJs found with above this depth (BED score field). Default: 0.' )

parser.add_option( '-E', '--snv_depth_cutoff', dest='snv_depth_cutoff', type='int', default=0, help='Keep only SNVs found with above this depth (DP=# field). Default: 0.' )

parser.add_option( '-M', '--minimum_length', dest='minimum_length', type='int', default=0, help='Keep only sequence variant peptides with greater than or equal to this length. Default: 0.' )

#Simple entry

parser.add_option( '-Q', '--bed_score_name', dest='bed_score_name', default="depth", help='Include in the NSJ ID line score_name:score. Default: "depth."' )

parser.add_option( '-R', '--reference', dest='reference', default="None", help='Genome Reference Name for NSJ ID location. Automatically pulled from genome_build header in GTF if present.' )

(options, args) = parser.parse_args()

##INPUTS##

#Protein FASTA

try:

protein_fasta = os.path.abspath(options.protein_fasta)

protein_fasta = open(protein_fasta, 'r')

protein_fasta = refparse.read_protein_fasta(protein_fasta)

except Exception, e:

print >> sys.stderr, "failed: %s" % e

exit(2)

#Reference XML/FASTA

ensembl, uniprot = None, None

if options.reference_xml != None:

try:

refXml = os.path.abspath(options.reference_xml)

refXml = open(refXml, 'r')

p = et.XMLParser(remove_blank_text=True) #required for pretty additions

uniprot = et.parse(refXml, p)

uniprot_root = uniprot.getroot()

for entry in uniprot_root:

refparse.condense_xml_entry(entry)

except Exception, e:

print >> sys.stderr, "Parsing and/or condensing reference xml failed: %s" % e

exit(2)

else:

print >> sys.stderr, "failed: no UniProt reference protein database specified"

#Read the Ensembl fasta into an XML structure

if options.protein_fasta != None:

try:

refFasta = os.path.abspath(options.protein_fasta)

refFasta = open(refFasta, 'r')

ensembl_root = et.Element(UP+'uniprot', nsmap=NAMESPACE_MAP)

ensembl = et.ElementTree(ensembl_root)

refparse.read_fasta_to_xml(ensembl_root, refFasta) #TODO: singluarize the entry here

except Exception, e:

print >> sys.stderr, "Parsing reference fasta failed: %s" %e

exit(2)

else:

print >> sys.stderr, "failed: no Ensembl reference protein database specified"

##OUTPUT##

outXml, outFasta = None, None

if options.output == None: outXml = sys.stdout

else:

try:

outBasename = os.path.basename(options.output)

outXml = open(outBasename + '.xml', 'w')

outFasta = open(outBasename + '.fasta', 'w')

except Exception, e:

print >> sys.stderr, "Opening outfile failed: %s" % e

exit(3)

#Process gene model

try:

geneModelFile = os.path.abspath(options.gene_model)

linect = sum(1 for line in open(geneModelFile))

geneModelFile = open(geneModelFile, 'r')

geneModel = refparse.GeneModel()

for i, gtf_line in enumerate(geneModelFile):

if i % 20000 == 0: print "gene_model line " + str(i) + " of " + str(linect)

if gtf_line.startswith('#') and gtf_line.find('genome_build') >= 0: options.reference = gtf_line.split()[1]

elif gtf_line.startswith('#'): continue

else: geneModel.new_entry(gtf_line)

except Exception, e:

print >> sys.stderr, "Parsing gene model failed: %s" % e

exit(2)

#Process VCF

try:

snpeff_vcf = os.path.abspath(options.snpeff_vcf)

linect = sum(1 for line in open(snpeff_vcf))

snpeff_vcf = open(snpeff_vcf, 'r')

for i, vcf_line in enumerate(snpeff_vcf):

if i % 1000 == 0: print "snpeff_vcf line " + str(i) + " of " + str(linect)

variantcalls.parse_vcf_line(ensembl_root, vcf_line, protein_fasta, options.snv_depth_cutoff, options.minimum_length, options.leading_aa_num, options.trailing_aa_num)

snpeff_vcf.close()

except Exception, e:

print >> sys.stderr, "VCF processing failed: %s" % e

exit(1)

#Process Splice BED

try:

splice_bed = os.path.abspath(options.splice_bed)

linect = sum(1 for line in open(splice_bed))

splice_bed = open(splice_bed, 'r')

for i, bed_line in enumerate(splice_bed):

if i % 2000 == 0: print "splice_bed line " + str(i) + " of " + str(linect)

if bed_line.startswith('track'): continue

novelsplices.translate_bed_line(ensembl_root, geneModel, bed_line, options.nsj_depth_cutoff, options.minimum_length, options.reference, options.bed_score_name)

splice_bed.close()

except Exception, e:

print >> sys.stderr, "Splice BED processing failed: %s" % e

exit(2)

#Enter PTM information from uniprot into the EnsemblXML

uniprot_root.remove(uniprot_root.find(UP+'copyright'))

ensembl_entries_by_seq = {entry.find(UP+'sequence').text.replace('\n', '').replace('\r', ''): entry for entry in ensembl_root}

uniprot_entries_by_seq = {entry.find(UP+'sequence').text.replace('\n', '').replace('\r', ''): entry for entry in uniprot_root}

common_seqs = list(set(ensembl_entries_by_seq) & set(uniprot_entries_by_seq))

uniprot_only = list(set(uniprot_entries_by_seq) - set(common_seqs))

ensembl_only = list(set(ensembl_entries_by_seq) - set(common_seqs))

for seq in common_seqs:

featureshare.share_all_features(uniprot_entries_by_seq[seq], ensembl_entries_by_seq[seq])

aa_num_dict = featureshare.aa_num_dict()

for i, ensembl_seq in enumerate(ensembl_only):

for j, uniprot_seq in enumerate(uniprot_only):

if len(ensembl_seq) == len(uniprot_seq):

ensembl_num_seq = featureshare.seq_to_num(ensembl_seq, aa_num_dict)

uniprot_num_seq = featureshare.seq_to_num(uniprot_seq, aa_num_dict)

comparison = uniprot_num_seq - ensembl_num_seq

if np.count_nonzero(comparison) <= 3:

indel.append_seqvar(ensembl_entries_by_seq[ensembl_seq], uniprot_only[j], comparison) # j is a holder for now; planning to remove this...

elif np.abs(len(ensembl_seq) - len(uniprot_seq)) <= 3:

result = indel.get_indel_locations(ensembl_seq, uniprot_seq)

indel.append_indel(ensembl_entries_by_seq[ensembl_seq], uniprot_only[j], result)

#Write the sample-specific xml to outfile

ensembl.write(outXml, pretty_print=True)

#Write the sample-specific fasta to outfile

entryct = len(ensembl_root)

if outFasta != None:

for i, entry in enumerate(ensembl_root):

if i % 1000 == 0: print "writing entry " + str(i) + " of " + str(entryct)

entry = refparse.xml_to_fasta(entry)

if entry == None: continue

else: outFasta.write(entry[0] + '\n' + entry[1] + '\n')

outXml.close()