Identification of the Dombrock blood group glycoprotein as a polymorphic member of the ADP-ribosyltransferase gene family

Blood ◽  
2000 ◽  
Vol 96 (7) ◽  
pp. 2621-2627 ◽  
Author(s):  
Alexander N. Gubin ◽  
J. Muthoni Njoroge ◽  
Urszula Wojda ◽  
Svetlana D. Pack ◽  
Maria Rios ◽  
...  
Keyword(s):  
Gene Family ◽  
Blood Group ◽  
Cell Biology ◽  
Specific Binding ◽  
Fetal Liver ◽  
Erythroid Differentiation ◽  
Single Amino Acid ◽  
Erythroid Cell ◽  
Blood Group System ◽  
Adp Ribosyltransferase

Abstract Identification of the 25 known human blood group molecules is of fundamental importance for the fields of erythroid cell biology and transfusion medicine. Here we provide the first molecular description of the “Dombrock” blood group system. A candidate gene was identified by in silico analyses of approximately 5000 expressed sequence tags (ESTs) from terminally differentiating human erythroid cells. Transfection experiments demonstrated specific binding of anti-Dombrock and confirmed glycosylphosphatidylinositol membrane attachment. Dombrock expression is developmentally regulated during erythroid differentiation and occurs at highest levels in the fetal liver. Homology studies suggest that the Dombrock molecule is a member of the adenosine 5′-diphosphate (ADP)–ribosyltransferase ectoenzyme gene family. Genotypic comparisons suggest Doa versus Dob antigenicity results from a single amino acid substitution within an encoded arginine-glycine-aspartic acid (RGD) motif of the molecule.

Identification of the Dombrock blood group glycoprotein as a polymorphic member of the ADP-ribosyltransferase gene family

Blood ◽  
2000 ◽  
Vol 96 (7) ◽  
pp. 2621-2627
Author(s):  
Alexander N. Gubin ◽  
J. Muthoni Njoroge ◽  
Urszula Wojda ◽  
Svetlana D. Pack ◽  
Maria Rios ◽  
...  
Keyword(s):  
Gene Family ◽  
Blood Group ◽  
Cell Biology ◽  
Specific Binding ◽  
Fetal Liver ◽  
Erythroid Differentiation ◽  
Single Amino Acid ◽  
Erythroid Cell ◽  
Blood Group System ◽  
Adp Ribosyltransferase

Identification of the 25 known human blood group molecules is of fundamental importance for the fields of erythroid cell biology and transfusion medicine. Here we provide the first molecular description of the “Dombrock” blood group system. A candidate gene was identified by in silico analyses of approximately 5000 expressed sequence tags (ESTs) from terminally differentiating human erythroid cells. Transfection experiments demonstrated specific binding of anti-Dombrock and confirmed glycosylphosphatidylinositol membrane attachment. Dombrock expression is developmentally regulated during erythroid differentiation and occurs at highest levels in the fetal liver. Homology studies suggest that the Dombrock molecule is a member of the adenosine 5′-diphosphate (ADP)–ribosyltransferase ectoenzyme gene family. Genotypic comparisons suggest Doa versus Dob antigenicity results from a single amino acid substitution within an encoded arginine-glycine-aspartic acid (RGD) motif of the molecule.

scholarly journals Molecular basis of reduced or absent expression of decay-accelerating factor in Cromer blood group phenotypes

Blood ◽  
1994 ◽  
Vol 84 (4) ◽  
pp. 1276-1282 ◽  
Author(s):  
DM Lublin ◽  
G Mallinson ◽  
J Poole ◽  
ME Reid ◽  
ES Thompson ◽  
...  
Keyword(s):  
Blood Group ◽  
Molecular Basis ◽  
Single Amino Acid ◽  
Single Point Mutation ◽  
Blood Group System ◽  
Single Nucleotide ◽  
Decay Accelerating Factor ◽  
Exon 2 ◽  
Nucleotide Deletion ◽  
Single Amino Acid Change

Abstract The human erythrocyte blood group system Cromer consists of high- incidence and low-incidence antigens that reside on decay-accelerating factor (DAF; CD55), a glycosyl-phosphatidylinositol-anchored membrane protein that regulates complement activation on cell surfaces. In the Cromer phenotypes Dr(a-) and Inab there is reduced or absent expression of DAF, respectively. This study investigated the molecular basis of the reduced DAF expression by polymerase chain reaction amplification of genomic DNA and RNA/cDNA obtained from Epstein-Barr virus- transformed lymphoblastoid cell lines. Sequence analysis of the Inab propositus showed a single nucleotide substitution in exon 2 of the DAF gene and at the corresponding position in the cDNA, G314-->A resulting in Trp53-->Stop. This truncation near the amino terminus explains the complete absence of surface DAF in the Inab phenotype. A similar analysis was performed for two Dr(a-) individuals, including KZ, who was previously reported to be Inab phenotype but is now shown by immunochemical and serologic methods to be Dr(a-) phenotype. A single nucleotide change was found in exon 5 of the DAF gene, C649-->T resulting in Ser165-->Leu, which we had previously shown to lead to loss of the Dra epitope. However, two species of cDNA were found, one encoding full-length DAF with the single amino acid change and the more abundant species having a 44-nucleotide deletion. The 44 nucleotide deletion includes the single polymorphic site, which creates a cryptic branch point in the Dr(a-) allele that leads to use of a downstream cryptic acceptor splice site. This shifts the reading frame and leads to a premature stop codon that precludes membrane anchoring. Thus, the single point mutation in the Dr(a-) phenotype results in a novel use of alternative splicing and provides a molecular explanation for both the antigenicity and the reduced DAF expression seen in this phenotype.

Klf1 Regulatory Networks in Primary Erythroid Cells.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1462-1462
Author(s):  
Michael Tallack ◽  
Thomas Whitington ◽  
Brooke Gardiner ◽  
Eleanor Wainwright ◽  
Janelle Keys ◽  
...  
Keyword(s):  
Regulatory Networks ◽  
Conflicts Of Interest ◽  
Fetal Liver ◽  
Es Cells ◽  
Erythroid Differentiation ◽  
Erythroid Cell ◽  
Gene Promoters ◽  
Erythroid Cells ◽  
Red Cell Membrane ◽  
Sequencing Platform

Abstract Abstract 1462 Poster Board I-485 Klf1/Eklf regulates a diverse suite of genes to direct erythroid cell differentiation from bi-potent progenitors. To determine the local cis-regulatory contexts and transcription factor networks in which Klf1 works, we performed Klf1 ChIP-seq using the SOLiD deep sequencing platform. We mapped more than 10 million unique 35mer tags and found ∼1500 sites in the genome of primary fetal liver erythroid cells are occupied by endogenous Klf1. Many reside within well characterised erythroid gene promoters (e.g. b-globin) or enhancers (e.g. E2f2 intron 1), but some are >100kb from any known gene. We tested a number of Klf1 bound promoter and intragenic sites for activity in erythroid cell lines and zebrafish. Our data suggests Klf1 directly regulates most aspects of terminal erythroid differentiation including synthesis of the hemoglobin tetramer, construction of a deformable red cell membrane and cytoskeleton, bimodal regulation of proliferation, and co-ordination of anti-apoptosis and enucleation pathways. Additionally, we suggest new mechanisms for Klf1 co-operation with other transcription factors such as those of the gata, ets and myb families based on over-representation and spatial constraints of their binding motifs in the vicinity of Klf1-bound promoters and enhancers. Finally, we have identified a group of ∼100 Klf1-occupied sites in fetal liver which overlap with Klf4-occupied sites in ES cells defined by Klf4 ChIP-seq. These sites are associated with genes controlling the cell cycle and proliferation and are Klf4-dependent in skin, gut and ES cells, suggesting a global paradigm for Klfs as regulators of differentiation in many, if not all, cell types. Disclosures No relevant conflicts of interest to declare.

scholarly journals Molecular basis of reduced or absent expression of decay-accelerating factor in Cromer blood group phenotypes

Blood ◽  
1994 ◽  
Vol 84 (4) ◽  
pp. 1276-1282 ◽  
Author(s):  
DM Lublin ◽  
G Mallinson ◽  
J Poole ◽  
ME Reid ◽  
ES Thompson ◽  
...  
Keyword(s):  
Blood Group ◽  
Molecular Basis ◽  
Single Amino Acid ◽  
Single Point Mutation ◽  
Blood Group System ◽  
Single Nucleotide ◽  
Decay Accelerating Factor ◽  
Exon 2 ◽  
Nucleotide Deletion ◽  
Single Amino Acid Change

The human erythrocyte blood group system Cromer consists of high- incidence and low-incidence antigens that reside on decay-accelerating factor (DAF; CD55), a glycosyl-phosphatidylinositol-anchored membrane protein that regulates complement activation on cell surfaces. In the Cromer phenotypes Dr(a-) and Inab there is reduced or absent expression of DAF, respectively. This study investigated the molecular basis of the reduced DAF expression by polymerase chain reaction amplification of genomic DNA and RNA/cDNA obtained from Epstein-Barr virus- transformed lymphoblastoid cell lines. Sequence analysis of the Inab propositus showed a single nucleotide substitution in exon 2 of the DAF gene and at the corresponding position in the cDNA, G314-->A resulting in Trp53-->Stop. This truncation near the amino terminus explains the complete absence of surface DAF in the Inab phenotype. A similar analysis was performed for two Dr(a-) individuals, including KZ, who was previously reported to be Inab phenotype but is now shown by immunochemical and serologic methods to be Dr(a-) phenotype. A single nucleotide change was found in exon 5 of the DAF gene, C649-->T resulting in Ser165-->Leu, which we had previously shown to lead to loss of the Dra epitope. However, two species of cDNA were found, one encoding full-length DAF with the single amino acid change and the more abundant species having a 44-nucleotide deletion. The 44 nucleotide deletion includes the single polymorphic site, which creates a cryptic branch point in the Dr(a-) allele that leads to use of a downstream cryptic acceptor splice site. This shifts the reading frame and leads to a premature stop codon that precludes membrane anchoring. Thus, the single point mutation in the Dr(a-) phenotype results in a novel use of alternative splicing and provides a molecular explanation for both the antigenicity and the reduced DAF expression seen in this phenotype.

scholarly journals The mouse Klf1 Nan variant impairs nuclear condensation and erythroid maturation

2018 ◽  
Author(s):  
Ileana Cantú ◽  
Harmen J.G. van de Werken ◽  
Nynke Gillemans ◽  
Ralph Stadhouders ◽  
Steven Heshusius ◽  
...  
Keyword(s):  
Fetal Liver ◽  
Chromatin Condensation ◽  
Erythroid Differentiation ◽  
Liver Cells ◽  
Single Amino Acid ◽  
Erythroid Cells ◽  
Nuclear Condensation ◽  
Similar Phenotype ◽  
Fetal Liver Cells ◽  
Erythroid Maturation

ABSTRACTKrüppel-like factor 1 (KLF1) is an essential transcription factor for erythroid development, as demonstrated by Klf1 knockout mice which die around E14 due to severe anemia. In humans, >65 KLF1 variants, causing different erythroid phenotypes, have been described. The Klf1 Nan variant, a single amino acid substitution (p.E339D) in the DNA binding domain, causes hemolytic anemia and is dominant over wildtype KLF1. Here we describe the effects of the Nan variant during fetal development. We show that Nan embryos have defects in erythroid maturation. RNA-sequencing of the Nan fetal liver cells revealed that Exportin 7 (Xpo7) was among the ~780 deregulated genes. This nuclear exportin is implicated in terminal erythroid differentiation; in particular it is involved in nuclear condensation. Indeed, KLF1 Nan fetal liver cells had larger nuclei and reduced chromatin condensation. Knockdown of XPO7 in wildtype erythroid cells caused a similar phenotype. We conclude that reduced expression of XPO7 is partially responsible for the erythroid defects observed in Nan erythroid cells.

Identification of a Defect in the Intracellular Trafficking of a Kell Blood Group Variant

Blood ◽  
1999 ◽  
Vol 94 (1) ◽  
pp. 310-318 ◽  
Author(s):  
Karina Yazdanbakhsh ◽  
Soohee Lee ◽  
Qian Yu ◽  
Marion E. Reid
Keyword(s):  
Membrane Proteins ◽  
Cell Surface ◽  
Blood Group ◽  
Molecular Mechanisms ◽  
Expression System ◽  
Single Amino Acid ◽  
Cell Surface Expression ◽  
Blood Group System ◽  
Structural Constraints ◽  
Low Incidence

Blood group polymorphisms have been used as tools to study the architecture of the red blood cell (RBC) membrane. Some blood group variants have reduced antigen expression at the cell surface. Understanding the underlying mechanism for this reduced expression can potentially provide structural information and help to elucidate protein trafficking pathways of membrane proteins. The Kp(a+) phenotype is a variant in the Kell blood group system that is associated with a single amino acid substitution (R281W) in the Kell glycoprotein and serologically associated with a weakened expression of other Kell system antigens by an unknown mechanism. We found by immunoblotting of RBCs that the weakening of Kell antigens in this variant is due to a reduced amount of total Kell glycoprotein at the cell surface rather than to the inaccessibility of the antigens to Kell antibodies. Using a heterologous expression system, we demonstrate that the Kpa mutation causes retention of most of the Kell glycoprotein in a pre-Golgi compartment due to differential processing, thereby suggesting aberrant transport of the Kell protein to the cell surface. Furthermore, we demonstrated that single nucleotide substitutions into the coding region of the common KEL allele, as predicted by the molecular genotyping studies, was sufficient to encode three clinically significant low incidence antigens. We found that two low incidence antigens can be expressed on a single Kell protein, thus showing that the historical failure to detect such a variant is not due to structural constraints in the Kell protein. These studies demonstrate the power of studying the molecular mechanisms of blood group variants for elucidating the intracellular transport pathways of membrane proteins and the requirements for cell surface expression.

scholarly journals Homeodomain-interacting protein kinase 2 plays an important role in normal terminal erythroid differentiation

Blood ◽  
2010 ◽  
Vol 115 (23) ◽  
pp. 4853-4861 ◽  
Author(s):  
Shilpa M. Hattangadi ◽  
Karly A. Burke ◽  
Harvey F. Lodish
Keyword(s):  
Cell Cycle Progression ◽  
Fetal Liver ◽  
Erythroid Differentiation ◽  
Erythroid Cell ◽  
Cycle Progression ◽  
Interacting Protein ◽  
Primary Mouse ◽  
Proliferation And Apoptosis ◽  
Hemoglobin Biosynthesis ◽  
Terminal Erythroid Differentiation

Abstract Gene-targeting experiments report that the homeodomain-interacting protein kinases 1 and 2, Hipk1 and Hipk2, are essential but redundant in hematopoietic development because Hipk1/Hipk2 double-deficient animals exhibit severe defects in hematopoiesis and vasculogenesis, whereas the single knockouts do not. These serine-threonine kinases phosphorylate and consequently modify the functions of several important hematopoietic transcription factors and cofactors. Here we show that Hipk2 knockdown alone plays a significant role in terminal fetal liver erythroid differentiation. Hipk1 and Hipk2 are highly induced during primary mouse fetal liver erythropoiesis. Specific knockdown of Hipk2 inhibits terminal erythroid cell proliferation (explained in part by impaired cell-cycle progression as well as increased apoptosis) and terminal enucleation as well as the accumulation of hemoglobin. Hipk2 knockdown also reduces the transcription of many genes involved in proliferation and apoptosis as well as important, erythroid-specific genes involved in hemoglobin biosynthesis, such as α-globin and mitoferrin 1, demonstrating that Hipk2 plays an important role in some but not all aspects of normal terminal erythroid differentiation.

scholarly journals Cellular immune responses in red blood cell alloimmunization

Hematology ◽  
2016 ◽  
Vol 2016 (1) ◽  
pp. 452-456 ◽  
Author(s):  
James C. Zimring ◽  
Krystalyn E. Hudson
Keyword(s):  
Blood Cell ◽  
Blood Group ◽  
Red Blood Cell ◽  
Cell Biology ◽  
Antigen Processing ◽  
The United States ◽  
Single Amino Acid ◽  
Solid Organ ◽  
Blood Group Antigens ◽  
Transfusion Therapy

Abstract In excess of 340 blood group antigens have now been described that vary between individuals. Thus, any unit of blood that is nonautologous represents a significant dose of alloantigen. Most blood group antigens are proteins, which differ by a single amino acid between donors and recipients. Approximately 1 out of every 70 individuals are transfused each year (in the United States alone), which leads to antibody responses to red blood cell (RBC) alloantigens in some transfusion recipients. When alloantibodies are formed, in many cases, RBCs expressing the antigen in question can no longer be safely transfused. However, despite chronic transfusion, only 3% to 10% of recipients (in general) mount an alloantibody response. In some disease states, rates of alloimmunization are much higher (eg, sickle cell disease). For patients who become alloimmunized to multiple antigens, ongoing transfusion therapy becomes increasingly difficult or, in some cases, impossible. While alloantibodies are the ultimate immune effector of humoral alloimmunization, the cellular underpinnings of the immune system that lead to ultimate alloantibody production are complex, including antigen consumption, antigen processing, antigen presentation, T-cell biology, and B-cell biology. Moreover, these cellular processes differ to some extent with regard to transfused RBCs as compared with other better-studied immune barriers (eg, infectious disease, vaccines, and solid organ transplantation). The current work focuses on illustrating the current paradigm of humoral immunity, with a specific focus on particulars of RBC alloimmunization and recent advances in the understanding thereof.

ADAR1 Is Essential For Erythroid Development

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 9-9
Author(s):  
Brian Liddicoat ◽  
Jochen Hartner ◽  
Alistair Chalk ◽  
Jun Lu ◽  
Stuart H. Orkin ◽  
...  
Keyword(s):  
Cell Fate ◽  
Rna Editing ◽  
Fetal Liver ◽  
Erythroid Differentiation ◽  
Negative Regulator ◽  
Erythroid Cell ◽  
Compensatory Mechanism ◽  
Type I ◽  
Hematopoietic Stem ◽  
Erythroid Development

Abstract Understanding the mechanisms controlling erythroid differentiation may allow therapeutic regulation of erythropoiesis. Focus so far has concentrated on the roles of transcription factors in determining erythroid fate, however one mechanism of gene regulation that has been poorly explored is that of direct modification of RNA. It is now becoming apparent that RNA sequences can be widely modified by RNA editing. Adenosine-to-Inosine (A-to-I) RNA editing is a posttranscriptional process resulting in sequence alterations to RNA transcripts, which is catalyzed by members of the adenosine deaminase acting on RNA (ADAR) family of enzymes. We have previously reported that targeted disruption of ADAR1 in hematopoietic stem cells (HSC) resulted in a global upregulation of type I and II interferon-inducible transcripts and rapid apoptosis (Hartner et al., Nature Immunology 2009). This study identified that ADAR1 is essential for maintenance of both the fetal and adult HSC compartment in a cell-autonomous fashion. Although ADAR1 is dispensable for myeloid development and B-lymphopoiesis, investigation of its role in erythropoiesis is granted owing to the severe loss erythrocytes in ADAR1 null mice. To determine the role of ADAR1 in erythropoiesis, we generated mice with an erythroid-restricted deletion of ADAR1 (EpoR-Cre Adar1fl/-). These animals were found to die in utero at ∼E14.5. The fetal liver (FL) was small and had significantly lower cellularity than in control littermates. Analysis of FL erythropoiesis demonstrated increased apoptosis and a loss of cells after the phenotypic polychromatic erythroblast stage of erythroid differentiation. When transplantation studies were done with EpoR-Cre Adar1fl/- FL and control EpoR-Cre Adar1fl/+ FL, recipients of EpoR-Cre Adar1fl/- FL could not restore erythropoiesis and were anemic in contrast to the controls. They also developed splenomegaly suggestive of extramedullary hemopoiesis occurring as a compensatory mechanism. To understand the mechanism through which ADAR1 regulates erythroid development, gene expression arrays and miRNA profiling were performed. As with HSCs, loss of ADAR1 caused a significant upregulation of IFN signatures in EpoR-Cre Adar1fl/- FL. Interestingly, despite evidence related to regulation of miRNA function by ADARs, there were only subtle changes to the expression pattern and levels of miRNAs in Adar1-deficient erythroid cells. These results demonstrate that ADAR1 is an essential in vivo mediator of erythroid cell fate and a critical negative regulator of the IFN response in cells specifically committed to erythropoiesis. Disclosures: Hartner: TaconicArtemis: Employment.

Exportin 7 (RanBP16) Plays An Essential Role In Terminal Erythroid Differentiation

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3877-3877
Author(s):  
Shilpa Hattangadi ◽  
Karly Burke ◽  
Harvey Lodish
Keyword(s):  
Cellular Proliferation ◽  
Conflicts Of Interest ◽  
Fetal Liver ◽  
Chromatin Condensation ◽  
Erythroid Differentiation ◽  
Erythroid Cell ◽  
Activating Transcription Factor 4 ◽  
Activating Transcription Factor ◽  
Terminal Erythroid Differentiation

Abstract Abstract 3877 Members of the nuclear transport receptor family of importins and exportins regulate the passage of proteins between the nucleus and cytoplasm. Although evolutionarily conserved across several species, Exportin 7 (Xpo7 or RanBP16) and its cargo are not well understood. In our study, we find that Xpo7 is highly erythroid-specific, as all other exportins are downregulated during terminal erythroid differentiation, a process including the induction of a highly specialized erythroid expression program, a set number of 3–5 terminal cell divisions, and chromatin condensation and eventual enucleation. Xpo7, in contrast, is highly induced during terminal erythropoiesis. Using retroviral shRNA knockdown of Xpo7 in in vitro fetal liver erythroid cell cultures, we demonstrate that exportin 7 is necessary for normal cellular proliferation and terminal erythroid differentiation, specifically for normal enucleation. Through microarray and biocomputational analysis of mRNA isolated from the knockdown cultures, we have found that the promoters of genes that are dysregulated after Xpo7 knockdown are enriched for binding sites for the activating transcription factor 4 (ATF4). Given that the erythroid phenotype of the ATF4 knockout mouse is very similar to the specific erythroid defects we observe in our in vitro knockdown analysis, our data suggests that either ATF4 or its binding protein may be Xpo7's cargo during terminal erythroid differentiation. Ongoing studies aimed at confirming this mechanism, the interaction between ATF4 and Xpo7, and the role and cargo of Xpo7 in terminal erythroid differentiation, are underway. Disclosures: No relevant conflicts of interest to declare.

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