Missense mutations in PIEZO1, which encodes the Piezo1 mechanosensor protein, define Er red blood cell antigens

We welcome the comments of Bianchi et al. and share the authors’ enthusiasm for extended investigation of Piezo1 variant function in different physiological contexts, brought to the fore by our attribution of the Er blood group antigens to this protein (1).
We do however wish to record some clarifications, as we feel the suggestion that this discovery disrupts our understanding of blood group antigens and their phenotypic effects may be overstated. Piezo1/Er appears analogous to other ion channel/membrane transporters carrying blood groups, but also known to be disease-associated, e.g. Band 3/Diego and RhAG. In these cases, pathogenic mutations are largely transmembrane/internal (2) whilst antigenic mutations are, by nature, external and generally benign. This does not exclude a mutation being both antigenic and disease-causing, and it is possible that Er antigens (including those yet to be discovered) may encompass both. For example, the homozygous PIEZO1 mutation encoding M2225R in the protein’s extracellular domain (3), could yet prove to be associated with loss of a high incidence Er antigen, in addition to the documented DHS pathology.
We agree that only limited conclusions can be drawn from apparent absence of Piezo1 functional effects in erythroid BEL-A cells overexpressing Er variants. Such caveats are recognised in many Piezo1 functional studies in non-native cellular environments (4), and true assessment requires interrogation of channel activity in fresh red cells (5). The historic nature of the samples included in our study precludes such experiments, however available clinical data showed no typical DHS characteristics or obvious clinical manifestations. Together with unaltered channel activity observed by patch clamping of BEL-A cells, this suggests at least, absent or mild functional disruption. Whilst not excluding the possibility of clinical manifestations in either mature red cells or other in vivo cellular environments, our evidence does not support the suggestion that Er antigenic variants define particular health risks.

1. Karamatic Crew V, Tilley LA, Satchwell TJ, et al. Missense mutations in PIEZO1, encoding the Piezo1 mechanosensor protein, define the Er red blood cell antigens. Blood 2022; blood.2022016504. doi: 10.1182/blood.2022016504.
2. Flatt JF, Bruce LJ. The Molecular Basis for Altered Cation Permeability in Hereditary Stomatocytic Human Red Blood Cells. Front Physiol. 2018 Apr 16;9:367.
3. Zarychanski R, Schulz VP, Houston BL, et al. Mutations in the mechanotransduction protein PIEZO1 are associated with hereditary xerocytosis. Blood. 2012;120(9):1908-1915.
4. Yamaguchi Y, Allegrini B, Rapetti-Mauss R, et al. Hereditary Xerocytosis: Differential Behavior of PIEZO1 Mutations in the N-Terminal Extracellular Domain Between Red Blood Cells and HEK Cells. Front. Physiol. 2021 Oct 18; 12:736585.

Discovering that Piezo1 defines its own blood group is not only the resolvement of a 40-year mystery of the Er antigen (1) but also a new and disruptive definition of blood groups. To date, blood groups have been considered the result of rather minor modifications of the membrane outer surface, all linked to agglutination effects. Thus, further physiological features have been undervalued. Therefore, a Piezo1-based blood group is a game changer. In addition to being an integral membrane protein, Piezo1 is an ion channel. In particular, its Ca(2+)-conductance makes it a potent signal mediator even if abundant in a low copy number.
The fact that Crew et al. (1) did not find functional differences in Piezo1 variants overexpressed in the BEL-A cell line does not mean much. First, mechanosensitive ion channels such as Piezo1 are extremely sensitive to their nanoenvironment, resulting in variable properties depending on the host cell type (2). Second, the mutation in the Piezo1 "head" may affect protein trafficking in hematopoiesis (3) and therefore the extent of surface expression, resulting in different whole-cell currents.
However, the detection of putative normal channel activity in variants with antigenic effects stresses the need for functional evidence in assessing the pathogenic role of novel variants in patients suspected to have hereditary xerocytosis.
Moreover, due to the high signaling capacity of Piezo1 and the strong functional dependence on the cell type, its variants defined by the Er blood group may result in modulated functions in other Piezo1-expressing tissues (4). Thus, the Er blood group likely defines particular health risks.
The newly reported antigenic/pathogenic role of Piezo1, which has been demonstrated for other transmembrane proteins, might be extended further, clarifying the conditions of unexplained alloimmunization. This may be of particular interest in the transfusion-dependent cases affected by erythrocyte disorders known to be associated with resistance to malaria infection (5) and possibly coinherited with the newly reported Er antigens.

1. Crew VK, Tilley LA, Satchwell TJ, et al. Missense mutations in PIEZO1, encoding the Piezo1 mechanosensor protein, define the Er red blood cell antigens. Blood. 2022;
2. Yamaguchi Y, Allegrini B, Rapetti-Mauss R, et al. Hereditary Xerocytosis: Differential Behavior of PIEZO1 Mutations in the N-Terminal Extracellular Domain Between Red Blood Cells and HEK Cells. Front Physiol. 2021;12:736585.
3. Caulier A, Garçon L. PIEZO1, sensing the touch during erythropoiesis. Curr Opin Hematol. 2022;29(3):112–118.
4. Song S, Zhang H, Wang X, et al. The role of mechanosensitive Piezo1 channel in diseases. Prog Biophysics Mol Biology. 2022;172:39–49.
5. Ma S, Cahalan S, LaMonte G, et al. Common PIEZO1 Allele in African Populations Causes RBC Dehydration and Attenuates Plasmodium Infection. Cell. 2018;173(2):443-455.e12.