![Figure 2. Expression profiles of the primary erythroid culture cells from patients with KLF1 mutations. An expression profiling of the α(A) and β(B) globin gene clusters using Agilent tiled oligonucleotide array was performed in P2 erythroid cells compared with normal control. (A) A distinctive higher peak of ζ globin gene expression (dark arrow with gray box highlighted) was observed in the patient (bottom) compared with a normal cDNA from a control sample (top). (B) Two unique upregulated expression peaks of γ-globin (arrow 1) and ε-globin (arrow 2) genes in the patient were shown. Another upregulating expression of noncoding mRNA (arrow 3) was observed at the β-globin locus control region (β-globin locus control region, hypersensitive site 2 [HS-2]), all with dark arrows and gray boxes highlighted. However, the meaning of this observation on the downstream globin gene regulation and hereditary persistence of embryonic globin in the patient remains unclear. There was no significant change on expression of other nearby genes on both clusters. (C) Expression profile of erythroid specific genes from primary erythroid cell samples from 5 patients (P) with KLF1 mutations and normal controls (n = 6), using qPCR and Taqman probe hybridization, showing a marked increase of fetal and embryonic globin mRNA expression (HBZ, ζ-globin gene; HBE, ε-globin gene; HBG, γ-globin gene) in all patients with relatively normal expression of KLF1. This suggests that although most patients carry a single nonsense mutation, which could result in haplo-insufficiency, a missense mutation on the other allele might produce stable transcripts that could be detected at similar levels to normal. Alternatively, our mRNA analysis used might not be sensitive enough to demonstrate a minor perturbation in the level of KLF1 expression. With the exception of BCL11A (B-cell CLL/lymphoma 11A [zinc finger protein] from 1 patient [P2]), there was no change in expression compared with normal controls of other erythroid-specific genes, including CD71, SOX6, ERAF, GYPA, glycophorin A, and EPOR (erythropoietin receptor; supplementary Figure 5).](https://ash.silverchair-cdn.com/ash/content_public/journal/blood/123/10/10.1182_blood-2013-09-526087/4/1586f2.jpeg?Expires=1722754247&Signature=SKUOcUtgiWxiZc-yNEyOPQjjx387uRHIIEjn9YE4ZfUb0XS23TplTnSMNzLfrqqOOV-tNBDsbOoZzUVhDp2Y-CPYOwVJgJNM5nFpdOOFP-jqYPc6h5Qhvl8XdwERVCydiZSTsQwnwfo9uhDoNXAtiBzrtCAQUCbxhOOiEpYkCqX18FjHG5s~o1Ey3WK-KX0foSU65C5feRg7UrxaolWlJiilLBuI7MEDmTdlUxqYqkde9IQoGI0x8eMhlAd8U7jwXD6AnxGmea-lbAxJJOfq-XLA6aAB5cMVPmT32O4btyurc-IXKFA1QT4vXKmRc4wNXApv8yXaacVjkDjtSRiEcw__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
Expression profiles of the primary erythroid culture cells from patients with KLF1 mutations. An expression profiling of the α(A) and β(B) globin gene clusters using Agilent tiled oligonucleotide array was performed in P2 erythroid cells compared with normal control. (A) A distinctive higher peak of ζ globin gene expression (dark arrow with gray box highlighted) was observed in the patient (bottom) compared with a normal cDNA from a control sample (top). (B) Two unique upregulated expression peaks of γ-globin (arrow 1) and ε-globin (arrow 2) genes in the patient were shown. Another upregulating expression of noncoding mRNA (arrow 3) was observed at the β-globin locus control region (β-globin locus control region, hypersensitive site 2 [HS-2]), all with dark arrows and gray boxes highlighted. However, the meaning of this observation on the downstream globin gene regulation and hereditary persistence of embryonic globin in the patient remains unclear. There was no significant change on expression of other nearby genes on both clusters. (C) Expression profile of erythroid specific genes from primary erythroid cell samples from 5 patients (P) with KLF1 mutations and normal controls (n = 6), using qPCR and Taqman probe hybridization, showing a marked increase of fetal and embryonic globin mRNA expression (HBZ, ζ-globin gene; HBE, ε-globin gene; HBG, γ-globin gene) in all patients with relatively normal expression of KLF1. This suggests that although most patients carry a single nonsense mutation, which could result in haplo-insufficiency, a missense mutation on the other allele might produce stable transcripts that could be detected at similar levels to normal. Alternatively, our mRNA analysis used might not be sensitive enough to demonstrate a minor perturbation in the level of KLF1 expression. With the exception of BCL11A (B-cell CLL/lymphoma 11A [zinc finger protein] from 1 patient [P2]), there was no change in expression compared with normal controls of other erythroid-specific genes, including CD71, SOX6, ERAF, GYPA, glycophorin A, and EPOR (erythropoietin receptor; supplementary Figure 5).
