An erythroid chaperone that facilitates folding of α-globin subunits for hemoglobin synthesis
Xiang Yu, … , Andrew J. Gow, Mitchell J. Weiss
Xiang Yu, … , Andrew J. Gow, Mitchell J. Weiss
Published July 2, 2007
Citation Information: J Clin Invest. 2007;117(7):1856-1865. https://doi.org/10.1172/JCI31664.
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Research Article

An erythroid chaperone that facilitates folding of α-globin subunits for hemoglobin synthesis

Abstract

Erythrocyte precursors produce abundant α- and β-globin proteins, which assemble with each other to form hemoglobin A (HbA), the major blood oxygen carrier. αHb-stabilizing protein (AHSP) binds free α subunits reversibly to maintain their structure and limit their ability to generate reactive oxygen species. Accordingly, loss of AHSP aggravates the toxicity of excessive free α-globin caused by β-globin gene disruption in mice. Surprisingly, we found that AHSP also has important functions when free α-globin is limited. Thus, compound mutants lacking both Ahsp and 1 of 4 α-globin genes (genotype Ahsp–/–α-globin*α/αα) exhibited more severe anemia and Hb instability than mice with either mutation alone. In vitro, recombinant AHSP promoted folding of newly translated α-globin, enhanced its refolding after denaturation, and facilitated its incorporation into HbA. Moreover, in erythroid precursors, newly formed free α-globin was destabilized by loss of AHSP. Therefore, in addition to its previously defined role in detoxification of excess α-globin, AHSP also acts as a molecular chaperone to stabilize nascent α-globin for HbA assembly. Our findings illustrate what we believe to be a novel adaptive mechanism by which a specialized cell coordinates high-level production of a multisubunit protein and protects against various synthetic imbalances.

Authors

Xiang Yu, Yi Kong, Louis C. Dore, Osheiza Abdulmalik, Anne M. Katein, Suiping Zhou, John K. Choi, David Gell, Joel P. Mackay, Andrew J. Gow, Mitchell J. Weiss

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Figure 6

Identification of transient free αHb and αHb-AHSP complex in erythroid cells.

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Identification of transient free αHb and αHb-AHSP complex in erythroid c...
Reticulocytes from β thalassemia intermedia mice (strain th-3) with wild-type or mutant Ahsp alleles were pulse-labeled with 35S-methionine for 10 minutes, lysed, and then treated with carbon monoxide to stabilize newly synthesized Hbs. Soluble proteins were fractionated by isoelectric focusing and visualized by autoradiography (top panel). Purified human βHb (hβHb; 100 μg/ml) or recombinant mouse AHSP (500 μg/ml) were added separately to different aliquots of each sample 30 minutes prior to electrophoresis, as indicated. The identities of the major bands are shown at the left. Free αHb (prominently visible in lanes 1 and 4) was observed at relatively high level in β thalassemic mice and to a lesser extent in wild-type mice (data not shown); this band shifted to the position of a human-mouse Hb heterotetramer (mα2hβ2) upon addition of human βHb (lanes 2 and 5) and to the position of a putative αHb-AHSP complex upon addition of purified recombinant AHSP (lanes 3 and 6). Note that in Ahsp–/– reticulocytes, the free αHb band was markedly reduced and the αHb-AHSP band was absent (lanes 7 and 8). The identity of the slowest migrating band (asterisk) is unknown. The bottom panel shows the same unstained CAE membrane with steady-state HbA chains visible. Note that free αHb and αHb-AHSP complex were not visualized, indicating that these species are relatively short-lived.