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( A ) Schematic overview of the in vitro experimental design to test the Alu insertion’s functional impact in the human induced pluripotent stem cell (iPSC) line, <t>KOLF2.1J.</t> ( B ) Expression of OCA2 and melanocyte marker genes across seven stages of melanocyte differentiation (days 0, 2, 8, 16, 19, 25, and 30). Each dot represents a technical replicate (heterozygous insertion carriers, red; wildtype, gray). Lines indicate the mean across two replicates at each time point. Two OCA2 amplicons are shown: exon 11-13 and exon 16-18 (exon numbering from RefSeq NM_000275.3 as displayed in the UCSC Genome Browser). ( C ) Representative image illustrating pigmentation differences between wild-type (WT; homozygous reference) and heterozygous (HET; one Alu insertion allele) melanocyte cultures. ( D ) Enrichment of histone marks, H3K4me1 and H3K27ac, at days 19 and 30. “IgG” and “Control” indicate negative controls for the enrichment assay and locus-specific signal, respectively. The locus “Control” is derived from the B2M gene. Fold enrichment values were normalized to 1% of the input control. [Figure created with BioRender]
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( A ) Schematic overview of the in vitro experimental design to test the Alu insertion’s functional impact in the human induced pluripotent stem cell (iPSC) line, <t>KOLF2.1J.</t> ( B ) Expression of OCA2 and melanocyte marker genes across seven stages of melanocyte differentiation (days 0, 2, 8, 16, 19, 25, and 30). Each dot represents a technical replicate (heterozygous insertion carriers, red; wildtype, gray). Lines indicate the mean across two replicates at each time point. Two OCA2 amplicons are shown: exon 11-13 and exon 16-18 (exon numbering from RefSeq NM_000275.3 as displayed in the UCSC Genome Browser). ( C ) Representative image illustrating pigmentation differences between wild-type (WT; homozygous reference) and heterozygous (HET; one Alu insertion allele) melanocyte cultures. ( D ) Enrichment of histone marks, H3K4me1 and H3K27ac, at days 19 and 30. “IgG” and “Control” indicate negative controls for the enrichment assay and locus-specific signal, respectively. The locus “Control” is derived from the B2M gene. Fold enrichment values were normalized to 1% of the input control. [Figure created with BioRender]
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( A ) Schematic overview of the in vitro experimental design to test the Alu insertion’s functional impact in the human induced pluripotent stem cell (iPSC) line, <t>KOLF2.1J.</t> ( B ) Expression of OCA2 and melanocyte marker genes across seven stages of melanocyte differentiation (days 0, 2, 8, 16, 19, 25, and 30). Each dot represents a technical replicate (heterozygous insertion carriers, red; wildtype, gray). Lines indicate the mean across two replicates at each time point. Two OCA2 amplicons are shown: exon 11-13 and exon 16-18 (exon numbering from RefSeq NM_000275.3 as displayed in the UCSC Genome Browser). ( C ) Representative image illustrating pigmentation differences between wild-type (WT; homozygous reference) and heterozygous (HET; one Alu insertion allele) melanocyte cultures. ( D ) Enrichment of histone marks, H3K4me1 and H3K27ac, at days 19 and 30. “IgG” and “Control” indicate negative controls for the enrichment assay and locus-specific signal, respectively. The locus “Control” is derived from the B2M gene. Fold enrichment values were normalized to 1% of the input control. [Figure created with BioRender]
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( A ) Schematic overview of the in vitro experimental design to test the Alu insertion’s functional impact in the human induced pluripotent stem cell (iPSC) line, <t>KOLF2.1J.</t> ( B ) Expression of OCA2 and melanocyte marker genes across seven stages of melanocyte differentiation (days 0, 2, 8, 16, 19, 25, and 30). Each dot represents a technical replicate (heterozygous insertion carriers, red; wildtype, gray). Lines indicate the mean across two replicates at each time point. Two OCA2 amplicons are shown: exon 11-13 and exon 16-18 (exon numbering from RefSeq NM_000275.3 as displayed in the UCSC Genome Browser). ( C ) Representative image illustrating pigmentation differences between wild-type (WT; homozygous reference) and heterozygous (HET; one Alu insertion allele) melanocyte cultures. ( D ) Enrichment of histone marks, H3K4me1 and H3K27ac, at days 19 and 30. “IgG” and “Control” indicate negative controls for the enrichment assay and locus-specific signal, respectively. The locus “Control” is derived from the B2M gene. Fold enrichment values were normalized to 1% of the input control. [Figure created with BioRender]
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( A ) Schematic overview of the in vitro experimental design to test the Alu insertion’s functional impact in the human induced pluripotent stem cell (iPSC) line, KOLF2.1J. ( B ) Expression of OCA2 and melanocyte marker genes across seven stages of melanocyte differentiation (days 0, 2, 8, 16, 19, 25, and 30). Each dot represents a technical replicate (heterozygous insertion carriers, red; wildtype, gray). Lines indicate the mean across two replicates at each time point. Two OCA2 amplicons are shown: exon 11-13 and exon 16-18 (exon numbering from RefSeq NM_000275.3 as displayed in the UCSC Genome Browser). ( C ) Representative image illustrating pigmentation differences between wild-type (WT; homozygous reference) and heterozygous (HET; one Alu insertion allele) melanocyte cultures. ( D ) Enrichment of histone marks, H3K4me1 and H3K27ac, at days 19 and 30. “IgG” and “Control” indicate negative controls for the enrichment assay and locus-specific signal, respectively. The locus “Control” is derived from the B2M gene. Fold enrichment values were normalized to 1% of the input control. [Figure created with BioRender]

Journal: bioRxiv

Article Title: A Denisovan-derived Alu insertion in OCA2 contributes to pigmentation diversity in present-day Melanesians

doi: 10.64898/2026.03.18.712481

Figure Lengend Snippet: ( A ) Schematic overview of the in vitro experimental design to test the Alu insertion’s functional impact in the human induced pluripotent stem cell (iPSC) line, KOLF2.1J. ( B ) Expression of OCA2 and melanocyte marker genes across seven stages of melanocyte differentiation (days 0, 2, 8, 16, 19, 25, and 30). Each dot represents a technical replicate (heterozygous insertion carriers, red; wildtype, gray). Lines indicate the mean across two replicates at each time point. Two OCA2 amplicons are shown: exon 11-13 and exon 16-18 (exon numbering from RefSeq NM_000275.3 as displayed in the UCSC Genome Browser). ( C ) Representative image illustrating pigmentation differences between wild-type (WT; homozygous reference) and heterozygous (HET; one Alu insertion allele) melanocyte cultures. ( D ) Enrichment of histone marks, H3K4me1 and H3K27ac, at days 19 and 30. “IgG” and “Control” indicate negative controls for the enrichment assay and locus-specific signal, respectively. The locus “Control” is derived from the B2M gene. Fold enrichment values were normalized to 1% of the input control. [Figure created with BioRender]

Article Snippet: We used the human iPSC line KOLF2.1J (The Jackson Laboratory) for all differentiation experiments.

Techniques: In Vitro, Functional Assay, Expressing, Marker, Control, Derivative Assay