Abstract
Alpha-hemoglobin stabilizing protein (AHSP) is a molecular chaperone that can reduce the damage caused by excess free α-globin to erythroid cells in patients with impaired β-globin chain synthesis. We assessed the effect of sodium phenylbutyrate and sodium valproate, two histone deacetylase inhibitors (HDIs) that are being studied for the treatment of hemoglobinopathies, on the expression of AHSP, BCL11A (all isoforms), γ-globin genes (HBG1/2), and some related transcription factors including GATA1, NFE2, EKLF, KLF4, and STAT3. For this purpose, the K562 cell line was cultured for 2, 4, and 6 days in the presence and absence of sodium phenylbutyrate and sodium valproate. Relative real-time qRT-PCR analysis of mRNA levels was performed to determine the effects of the two compounds on gene expression. Expression of all target mRNAs increased significantly (p < 0.05), except for the expression of BCL11A, which was down-regulated (p < 0.05) in the cells treated with both compounds relative to the levels measured for untreated cells. The findings indicated that sodium valproate had a more considerable effect than sodium phenylbutyrate (p < 0.0005) on BCL11A repression and the up-regulation of other studied genes. γ-Globin and AHSP gene expression continuously increased during the culture period in the treated cells, with the highest gene expression observed for 1 mM sodium valproate after 6 days. Both compounds repressed the expression of BCL11A (-XL, -L, -S) and up-regulated GATA1, NFE2, EKLF, KLF4, STAT3, AHSP, and γ-globin genes expression. Moreover, sodium valproate showed a stronger effect on repressing BCL11A and escalating the expression of other target genes. The findings of this in vitro experiment could be considered in selecting drugs for clinical use in patients with β-hemoglobinopathies.
Highlights
Alpha-hemoglobin stabilizing protein (AHSP) is an erythroid-specific protein that acts as a molecular chaperone for α-globin chains and forms a stable but reversible complex with the free alpha chains of hemoglobin, stabilizing them to prevent precipitation in cells [1].AHSP-alpha globin interactions could be a potential β-thalassemia modifier and lead to diversity in hematological and clinical symptoms of patients
The findings indicated that sodium valproate had a more considerable effect than sodium phenylbutyrate (p < 0.0005) on BCL11A repression and the up-regulation of other studied genes. γ-Globin and AHSP gene expression continuously increased during the culture period in the treated cells, with the highest gene expression observed for 1 mM sodium valproate after 6 days
The discovery of AHSP has delivered new perceptions into its role as a chaperone-like molecule specialized for erythroid series which is involved in the formation of functional hemoglobin
Summary
AHSP-alpha globin interactions could be a potential β-thalassemia modifier and lead to diversity in hematological and clinical symptoms of patients. The effect of HDIs on AHSP expression with β-thalassemia trait are asymptomatic and show insignificant symptoms, possibly as a result of excess free α-globin chains stabilization by AHSP. Simultaneous mutations that affect AHSP function or expression in β-thalassemia could result in more disease severity [2, 3]. It has been found that an increase in the α-globin to β-globin chain synthesis ratio leads to AHSP overexpression. AHSP can bind to more free alpha chains, decreasing the severity of clinical symptoms caused by sedimentation of excess unbound α-globin chains [4]
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