Pedigrees of three PCG families (PCG049, PCG050 and PCG067) having more than one affected member and segregating novel disease causing variant/s in <i>CYP1B1</i> gene.

Filled squares and circles indicate affected members of family. Cousin marriage is indicated by double line.</p

A comparison of <i>CYP1B1</i> gene conservation among different homologs for novel variants detected in this study.

a) Clustal Omega multiple sequence alignment (MSA) (Shown in red) for p.Arg153Gly detected in PCG049. b) Multiple sequence alignment (MSA) for p. Ser172Arg among homologs detected in PCG049 showing less conservation in Mus musculus. c) MSA result for p.Gly211Argfs*13 among homologs detected in PCG050. d) MSA result for variant p.Val241Glu detected in PCG052. e) MSA result for p.Leu97Ala*127 among homologs detected in PCG053. f) MSA result for p.Arg222Profs*2 detected in PCG054 showing less conservation in Mus musculus. g) MSA result for variant p.Val254Glyfs*73 detected on PCG059 showing complete conservation among homologs. h) MSA result for p. Leu247Gln among homologs detected in PCG060. i) MSA result for p.Phe421Leu detected in PCG062. j) MSA result for p.Leu264Alafs*63 detected in PCG063. k) MSA result for p.Asp242His detected in PCG067. a-k) WebLogo results of all novel CYP1B1 protein variants showing comparison of conservation among homologs are on right side of the figure. All the variants except p.Ser172Arg and p.Arg222Profs*73 show 100% conservation among different mammals (Large size of amino acid abbreviation letter show full conservation while small size show less conserved position among homologs).</p

Chromatograms of novel duplications and an insertion detected in PCG patients.

A) A homozygous variant c.629dup detected in PCG050 leading to p.Gly211Argfs*13. B) Chromatogram of homozygous variant c.287dup detected in PCG053 that resulted in protein change i.e., p.Leu97Alafs*127. C) Sequence chromatogram of homozygous variant c.662dup leading to protein change p.Arg222Profs*2 in PCG054. D) Chromatogram showing c.789dup homozygous variant in PCG063 leading to a p.Leu264Alafs*63. E) Homozygous insertion c.758_759insA detected in PCG059 resulting in p.Val254Glyfs*73. All the chromatograms on left side indicate normal sequence while right side of figure shows mutated chromatograms.</p

Fig 5 -

(a) Overview of CYP1B1 wild type protein in ribbon-presentation (b-l) In-silico protein prediction regarding variant protein structures for all detected variants by HOPE software. (b-l) i: for each variant show the position of amino acid in protein structure. (b-l) ii: for each detected variant show the zoomed in change in protein structure due to each respective variant (Wild type amino acid residue is colored as green while mutant is colored as red).</p

In-silico analysis data of disease-causing variants identified in this study.

In-silico analysis data of disease-causing variants identified in this study.</p

List of reported and novel disease-causing variants detected in this study upon sequencing of <i>CYP1B1</i> gene in PCG patients.

List of reported and novel disease-causing variants detected in this study upon sequencing of CYP1B1 gene in PCG patients.</p

Chromatograms of novel disease-causing single nucleotide substitutions detected in PCG patients upon sequencing of <i>CYP1B1</i> gene.

Right side of figure shows substituted nucleotides in chromatograms. A) A heterozygous variant c.457C>G detected in PCG049 leading to p.Arg153Gly. B) Second heterozygous variant c.516C>A detected in PCG049 resulting in p.Ser172Arg. C) Variant c.722T>A detected in homozygous condition in PCG052 leading to protein change p.Val241Glu. D) A heterozygous variant c.740T>A leading to p.Leu247Gln detected in PCG060. E) Sequence chromatogram of heterozygous variant c.1263T>A detected in PCG062 resulting in p.Phe421Leu. F) A homozygous variant c.724G>C leading to p.Asp242His detected in PCG067.</p

List of reported and a previously unreported single nucleotide polymorphism detected in <i>CYP1B1</i> gene in PCG patients analyzed in this study.

List of reported and a previously unreported single nucleotide polymorphism detected in CYP1B1 gene in PCG patients analyzed in this study.</p

DNA13-07610.hcdiffs

DNA13-07610.hcdiff