An erythroid chaperone that facilitates folding of α-globin subunits for hemoglobin synthesis
J. Clin. Invest. Xiang Yu, et al. 117:1856 doi:10.1172/JCI31664 [Go to this article.]

Figure 2
Increased Hb instability in Ahsp^–/–α-globin*^α/αα double-mutant erythrocytes. (A) Steady-state membrane-associated Hbs. TAU gel analysis of membrane skeletons prepared from equal numbers of erythrocytes from mice with genotypes indicated. α- and β-globin chains were stained with Coomassie blue and are indicated. Lane 9 represents an altered β-globin genotype, which results in 2 bands. (B) Analysis of newly synthesized globin chains. Equal numbers of reticulocytes were pulse-labeled with ³⁵S-methionine and ³⁵S-cysteine for 15 minutes, then disrupted by hypotonic lysis. Ahsp and α-globin genotypes are indicated. Soluble cytoplasmic (C) and membrane-associated (M) globins were isolated by differential centrifugation, fractionated on TAU gels, and visualized by autoradiography. The data shown derive from mice with the diffuse β-globin genotype. Results of quantitative analysis of nascent globins in membrane and cytoplasmic fractions from multiple experiments are shown in Table 2. (C) Quantification of ROS. Erythrocytes were incubated with dichlorofluorescin diacetate (DCFH-DA), which enters cells and is converted by ROS to the fluorescent product DCF. Representative flow cytometry data from 2 mice of each genotype are shown. Data from all mice analyzed are summarized in the upper-right corner of each panel, which shows the mean fluorescent intensity of DCF signal with wild-type erythrocytes normalized to 1.0. Three to 6 mice from each group were analyzed.