Abstract
The Sox6 transcription factor is crucial for terminal maturation of definitive red blood cells. Sox6-null mouse fetuses present misshapen and nucleated erythrocytes, due to impaired actin assembly and cytoskeleton stability. These defects are accompanied with a reduced survival of Sox6−/− red blood cells, resulting in a compensated anemia. Sox6-overexpression in K562 cells and in human primary ex vivo erythroid cultures enhances erythroid differentiation and leads to hemoglobinization, the hallmark of erythroid maturation. To obtain an overview on processes downstream to Sox6 expression, we performed a differential proteomic analysis on human erythroid K562 cells overexpressing Sox6. Sox6-overexpression induces dysregulation of 64 proteins, involved in cytoskeleton remodeling and in protein synthesis, folding and trafficking, key processes for erythroid maturation. Moreover, 43 out of 64 genes encoding for differentially expressed proteins contain within their proximal regulatory regions sites that are bound by SOX6 according to ENCODE ChIP-seq datasets and are possible direct SOX6 targets. SAR1B, one of the most induced proteins upon Sox6 overexpression, shares a conserved regulatory module, composed by a double SOX6 binding site and a GATA1 consensus, with the adjacent SEC24 A gene. Since both genes encode for COPII components, this element could concur to the coordinated expression of these proteins during erythropoiesis.
Highlights
Twenty Sox transcription factors exist in mice and humans[1,2]
The experiment was performed in quadruplicate on four pairs of samples from cells transduced with either the Sox6-overexpressing vector (Sox6-K562) or the corresponding Empty Vector (EV-K562) as a control, to ensure statistical replication (Fig. 1b)
To reinforce the notion that these genes could be relevant for the erythroid lineage, we looked for GATA1 peaks in the same regulatory regions, according to ENCODE Chromatin Immunoprecipitation (ChIP) datasets (Fig. 5 and Supplementary Fig. S2)
Summary
Twenty Sox transcription factors exist in mice and humans[1,2]. They feature a Sry-type high-mobility-group (HMG box) DNA binding domain. Erythroid differentiation involves the progressive activation of erythroid specific genes, including those encoding for specific RBCs membrane cytoskeleton components and for globin chains, which represent about 95% of RBCs protein content[12,15]. During their terminal maturation, erythroblasts progressively condense their nucleus, which is extruded together with most organelles. Sox[6] overexpression in K562 cells and in human primary ex vivo erythroid cultures enhances erythroid differentiation and a general hemoglobinization[11], the main hallmark of erythroid terminal maturation Overall, these data point to a critical role of SOX6 for cell survival, proliferation and terminal maturation in both normal and stress erythropoiesis. We show that Sox[6] overexpression is associated with a significant change in proteins controlling cytoskeleton remodeling and protein trafficking, two important processes for erythroid terminal differentiation
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