D. R. Higgs

The ATRX syndrome protein forms a chromatin-remodeling complex with Daxx and localizes in promyelocytic leukemia nuclear bodies

ATRX syndrome is characterized by X-linked mental retardation associated with α-thalassemia. The gene mutated in this disease, ATRX, encodes a plant homeodomain-like finger and a SWI2/SNF2-like ATPase motif, both of which are often found in chromatin-remodeling enzymes, but ATRX has not been characterized biochemically. By immunoprecipitation from HeLa extract, we found that ATRX is in a complex with transcription cofactor Daxx. The following evidence supports that ATRX and Daxx are components of an ATP-dependent chromatin-remodeling complex: (i) Daxx and ATRX can be coimmunoisolated by antibodies specific for each protein; (ii) a proportion of Daxx cofractionates with ATRX as a complex of 1 MDa by gel-filtration analysis; (iii) in extract from cells of a patient with ATRX syndrome, the level of the Daxx–ATRX complex is correspondingly reduced; (iv) a proportion of ATRX and Daxx colocalize in promyelocytic leukemia nuclear bodies, with which Daxx had previously been located; and (v) the ATRX complex displays ATP-dependent activities that resemble those of other chromatin-remodeling complexes, including triple-helix DNA displacement and alteration of mononucleosome disruption patterns. But unlike the previously described SWI/SNF or NURD complexes, the ATRX complex does not randomize DNA phasing of the mononucleosomes, suggesting that it may remodel chromatin differently. Taken together, the results suggest that ATRX functions in conjunction with Daxx in a novel chromatin-remodeling complex. The defects in ATRX syndrome may result from inappropriate expression of genes controlled by this complex.

A PCR-based strategy to detect the common severe determinants of α thalassaemia

A rapid and inexpensive polymerase chain reaction (PCR) based strategy is described which detects the three common, severe α thalassaemia determinants observed in southeast Asia (——SEA) and the Mediterranean (——MED and —(α)20.5). Oligonucleotide primers have been chosen which allow specific identification of both normal (αα) and abnormal (——) chromosomes using identical conditions in either the same or parallel PCR reactions. This strategy should be useful in the development of screening programmes to identify carriers of α thalassaemia (——/αα) and prenatal diagnosis of the Hb Barts hydrops fetalis syndrome (——/——) for those populations in which this represents a major cause of perinatal death.

Thalassaemia intermedia in Cyprus: the interaction of alpha and beta thalassaemia.

Summary. Restriction endonuclease analysis has been performed on the α and β globin gene clusters of 57 Cypriots homozygous for β thalassaemia, 30 with the transfusion dependent form of the condition (thalassaemia major) and 27 who are less severely affected (thalassaemia intermedia). There was a significant difference in the incidence of α thalassaemia between the two groups: 14/27 of the patients with thalassaemia intermedia also had deletion forms of a thalassaemia, while only 4/30 of the patients with thalassaemia major were similarly affected. Thus in Cypriot patients who are homozygous for β thalassaemia the co‐inheritance of a thalassaemia is an important factor in determining the clinical course.

Characterization of two deletions that remove the entire human ζ-α globin gene complex (—THAI and —FIL)

We have fully characterized two α thalassaemia mutants that occur in Southeast Asia, —THAI and —FIL. Each mutant is due to a deletion that removes the entire ζ‐α‐globin gene complex. Localization of the 5’breakpoints described here, allows the identification of unique fragments that are specific for each of the two mutations. This information can be used to assess the frequency of these mutants in Southeast Asia and will be of value in prenatal testing for α thalassaemia in this area.

Haemoglobin Constant Spring has an unstable α chain messenger RNA

Haemoglobin Constant Spring (Hb CS) is a variant with an elongated α‐chain associated with an α+ thalassaemia phenotype. The amount of α mRNA relative to β mRNA in reticulocytes was reduced in carriers of Hb CS by an amount equivalent to the reduction observed in carriers of α+ thalassaemia. In a patient with Hb CS‐H disease there was greater α/β mRNA ratio in bone marrow nuclear RNA than in the peripheral blood. Furthermore, all the α mRNA in the patients peripheral blood was derived from the α1 (αA) gene. The data suggest that αCS mRNA is unstable and degraded in the cytoplasm. This instability may be due to destabilization of a specific sequence in the 3’non‐coding region during translation.

A genetic marker for elevated levels of haemoglobin F in homozygous sickle cell disease

Ten patients with sickle cell (SS) disease from a Jamaican family were found to have unusually high levels of haemoglobin F for this population. Each of them has inherited one sickle cell gene on a chromosome characterized by an arrangement of restriction fragment length polymorphisms (haplotype) which is very rare in the Jamaican population. Genetic analysis of the family suggests that there is a determinant linked to the β‐globin gene cluster, characterized by this haplotype, which is responsible for increased haemoglobin F production in response to anaemia. Interestingly this particular haplotype appears to be common in patients with SS disease in eastern Saudi Arabia in whom a high level of haemoglobin F is the rule rather than the exception. Hence it is possible that this haplotype (++–++) acts as a genetic marker for elevated levels of haemoglobin F in sickle cell disease.

Understanding α‐Globin Gene Regulation: Aiming to Improve the Management of Thalassemia

Abstract: Over the past 50 years, many advances in our understanding of the general principles controlling gene expression during hematopoiesis have come from studying the synthesis of hemoglobin. Discovering how the α‐ and β‐globin genes are normally regulated and documenting the effects of inherited mutations that cause thalassemia have played a major role in establishing our current understanding of how genes are switched on or off in hematopoietic cells. Previously, nearly all mutations causing thalassemia have been found in or around the globin loci, but rare inherited and acquired trans‐acting mutations are being found more often. Such mutations have demonstrated new mechanisms underlying human genetic disease. Furthermore, they are revealing new pathways in the regulation of globin gene expression that, in turn, may open up new avenues for improving the management of patients with common types of thalassemia.

Effect of alpha thalassaemia on the rheology of homozygous sickle cell disease

Summary. A study of rheological determinants (plasma viscosity, whole‐blood viscosity, and erythrocyte deformability) was made in 24 matched pairs of patients with homozygous sickle cell disease, with and without homozygous α‐thalassaemia 2. Patients with coexisting α‐thalassaemia showed a significant increase in erythrocyte deformability measured as filtration of washed erythrocytes through 5 μm diameter pores and also as viscosity of whole blood at high shear rate (230 s−1) and standard haematocrit (0·45). This rheological advantage may explain the beneficial effect of α‐thalassaemia 2 on haematological parameters and clinical events in homozygous sickle cell disease.

The changing face of homozygous sickle cell disease: 102 patients over 60 years

Earlier reports on homozygous sickle cell (SS) disease have been biased by severely affected cases. The Jamaican clinic which seeks to avoid such bias has 102 patients surviving beyond 60 years. The objective of this study was to examine the features of elderly cases and assess factors determining survival and the behaviour of this disease with advancing age. A retrospective review of all cases and prospective assessment in survivors was conducted at The Sickle Cell Clinic at the University of the West Indies, Kingston, Jamaica previously operated by the MRC Laboratories. All patients with SS disease born prior to December 31, 1943 who would, by January 2004, have passed their 60th birthday were traced and their current status ascertained. The molecular and clinical features were assessed and observations on the clinical behaviour of the disease and of haematology and biochemistry are presented. Of the 102 patients, 58 had died, four had emigrated and 40 were alive, resident in Jamaica and aged 60–87 years. Survival was associated with female gender and higher foetal haemoglobin but not with alpha‐thalassaemia or beta‐globin haplotype. A tendency to familial clustering among elderly survivors did not reach statistical significance. Painful crises ameliorated with age and there was a benign course in pregnancy. Mean haemoglobin levels fell with age and were generally associated with rising creatinine levels indicating the importance of renal failure. Elderly survivors present some features of intrinsic mildness but also manifest age‐related amelioration of painful crises and falling haemoglobin levels from progressive renal damage.

Alpha-thalassaemia in Nigeria: Its interaction with sickle-cell disease

We have determined the molecular basis and frequency of α‐thalassaemia in Nigeria. The a‐thalassaemia determinant in this population is caused by only one type of single α globin gene deletion (‐α3.7). Comparison of the haematological features of those patients who have sickle‐cell disease with (‐α/αα, ‐α/‐α) or without (αα/αα) α‐thalassaemia showed similar trends to those reported in Jamaican and U.S. patients with these interactions. However, in contrast to studies in some other African populations we have shown that the frequency of α‐thalassaemia in Nigeria is the same (0.24) in patients with or without homozygous sickle‐cell disease (AA, AS and AC genotypes).