NMR Of Red Cells: Plasma Membrane Oxidoreductase, And Cation Transport
Funder
National Health and Medical Research Council
Funding Amount
$192,388.00
Summary
An interesting paradox exists with respect to the 'central' function of the red blood cell (RBC): it delivers the main oxidising capacity to the body (O2), but it also carries the chemically opposite functionality in its membrane, namely reducing capacity. The reduction of many oxidised proteins and metabolites in blood plasma is mediated by a plasma-membrane oxido-reductase (PMOR). Ascorbic acid (vitamin C) dramatically accelerates this rate of reduction but its precise molecular role is unknow ....An interesting paradox exists with respect to the 'central' function of the red blood cell (RBC): it delivers the main oxidising capacity to the body (O2), but it also carries the chemically opposite functionality in its membrane, namely reducing capacity. The reduction of many oxidised proteins and metabolites in blood plasma is mediated by a plasma-membrane oxido-reductase (PMOR). Ascorbic acid (vitamin C) dramatically accelerates this rate of reduction but its precise molecular role is unknown; neither is the immediate source of the reducing equivalents (electrons) known. Novel, non-invasive, 13C NMR methods have been developed, and others are planned in this project, to study the rate of reduction of Otest? compounds, including 13C-ferricyanide, and reactions of 13C-ascorbate. This will provide a quantitative understanding of the kinetics of the redox reactions in the intact cell. The transfer of negative charges (electrons) from the cell, in the longer term (minutes) inevitably must be matched by the movement of cations (positive charges). The main cation flux is mediated by Na+, K+-ATPase, but various cation exchange pathways are also involved in the total Oionic economy? of the cell. Of special interest will be the calcium-activated K+ (or Gardos) channel. This Oopens? inappropriately in malaria, sickle cell anaemia, and under blood bank storage conditions, and this is thought to be the basis of some of the pathological events in these conditions. The alkali-metal cation exchange pathway ( Na+-Li+) is more activate in the red cells of many patients with hypertension. So, multiple-quantum NMR methods will be used to monitor membrane transport and binding of cations to characterise the kinetics and regulation of the K+-channel, and the Na+-Li+ exchange reactions. The significance will lie in a basic understanding of, and possible 'diagnostic methods' for the biochemical processes that occur in red blood cells in health and disease.Read moreRead less
Hemoglobin is the major protein in red blood cells and is essential for the transport of oxygen from the lungs to the tissues. The disorders of hemoglobin production are the commonest genetic diseases world-wide. These diseases can be markedly improved with elevation of the form of hemoglobin produced by the developing fetus, fetal hemoglobin. We have identified a key factor important for fetal gene expression. We will now determine whether manipulation of this factor can cure hemoglobin disorde ....Hemoglobin is the major protein in red blood cells and is essential for the transport of oxygen from the lungs to the tissues. The disorders of hemoglobin production are the commonest genetic diseases world-wide. These diseases can be markedly improved with elevation of the form of hemoglobin produced by the developing fetus, fetal hemoglobin. We have identified a key factor important for fetal gene expression. We will now determine whether manipulation of this factor can cure hemoglobin disorders.Read moreRead less
Characterisation Of Erythropoietic Mutants Identified In A Forward Genetic Screen In Mice.
Funder
National Health and Medical Research Council
Funding Amount
$501,902.00
Summary
The human bone marrow is the pivotal organ in the replacement of the vast numbers of blood cells normally consumed each day. One of the cells regenerated by this organ are the red blood cells which are critical for the transport of oxygen to the tissues. This proposal uses genetically altered mice to identify genes that are critical for the production of normal red blood cells. Mice exposed to a chemical that induces random mutations in their genome are bred and pups with abnormal red blood cell ....The human bone marrow is the pivotal organ in the replacement of the vast numbers of blood cells normally consumed each day. One of the cells regenerated by this organ are the red blood cells which are critical for the transport of oxygen to the tissues. This proposal uses genetically altered mice to identify genes that are critical for the production of normal red blood cells. Mice exposed to a chemical that induces random mutations in their genome are bred and pups with abnormal red blood cells are identified. The responsible genetic mutation is identified and the gene is then studied to determine how it influences red blood cell production. The results of these studies provide insights into a variety of human conditions including anemia, thalassemia and sickle cell disease.Read moreRead less
Identification Of Novel Mechanisms Governing Stage-specific Regulation Of The Human Globin Genes
Funder
National Health and Medical Research Council
Funding Amount
$577,889.00
Summary
Hemoglobin is the major protein in red blood cells and is essential for the transport of oxygen from the lungs to the tissues. The disorders of hemoglobin production are the commonest genetic diseases world-wide. These diseases can be markedly improved with elevation of the form of hemoglobin produced by the developing embryo, embryonic hemoglobin. We have identified a key factor important for fetal gene expression. Our goal is to translate these findings into therapies for the globin disorders.
Sickle cell anaemia and ?-thalassaemia are debilitating diseases for which there is no effective treatment. Patients require lifetime blood transfusions and drugs with significant side effects. These diseases are ameliorated in patients that express foetal haemoglobin into adulthood. The goal of this research is to understand how foetal haemoglobin is normally turned off at birth. This will ultimately help us discover how to switch it back on to treat patients.
Inherited disorders of the blood, such as sickle-cell anaemia and thalassaemia, result from mutations in the genes that produce haemoglobin. Current treatments can partially alleviate some of the debilitating symptoms of these diseases but these treatments have significant side effects, and despite the best efforts of clinicians, many patients succumb to their conditions at an early age. It has been observed that certain individuals exhibit a milder form of the disease, as a consequence of the r ....Inherited disorders of the blood, such as sickle-cell anaemia and thalassaemia, result from mutations in the genes that produce haemoglobin. Current treatments can partially alleviate some of the debilitating symptoms of these diseases but these treatments have significant side effects, and despite the best efforts of clinicians, many patients succumb to their conditions at an early age. It has been observed that certain individuals exhibit a milder form of the disease, as a consequence of the reactivation of their foetal haemoglobin genes, (a distinct set of genes that would have been active in utero but are normally silenced around the time of birth). It is widely accepted that if pharmaceutical means can be found for reactivating the foetal haemoglobin genes then many patients would benefit. The regulation of the foetal globin genes, like most human genes, is complicated and there are few obvious means of increasing their activity. Nevertheless, it is believed that by investigating the molecular mechanisms by which they are controlled it will be possible to devise therapeutic agents that mimic these mechanisms or to develop agents that prevent the shutdown of the foetal genes around birth. To this end we have been working on the molecules that regulate the activity of the haemoglobin genes. We have recently cloned a number of DNA-binding proteins, and their co-factors, that appear to be involved in silencing foetal globin gene expression. This grant proposal is concerned with learning how these new molecules operate to silence gene expression as a first step towards designing agents that will prevent the silencing.Read moreRead less
Functional Characterisation Of Regulators Of Human Globin Gene Switching
Funder
National Health and Medical Research Council
Funding Amount
$232,131.00
Summary
Red blood cells produce haemoglobin, a tetramer of two alpha globin chains and two beta-globin chains. Haemoglobin reversibly interacts with oxygen in such a way that it efficiently shuttles oxygen between the lungs and the rest of the body. Integrity of the hemoglobin molecule, and red cells which carry it, is essential for life of all organisms with blood. The alpha-globin and beta-globin chains that make up haemoglobin are prodcued by red cell precursors in the bone marrow according to the ge ....Red blood cells produce haemoglobin, a tetramer of two alpha globin chains and two beta-globin chains. Haemoglobin reversibly interacts with oxygen in such a way that it efficiently shuttles oxygen between the lungs and the rest of the body. Integrity of the hemoglobin molecule, and red cells which carry it, is essential for life of all organisms with blood. The alpha-globin and beta-globin chains that make up haemoglobin are prodcued by red cell precursors in the bone marrow according to the genetic blueprint (genes) that are inherited. Genetic disorders resulting from defects in the beta-globin gene are the most common inherited disorders of man. Children who fail to make beta-globin have a disease known as beta-thalassaemia. They are transfusion dependent from ~ 6 months of age and need intensive chelation therapy (infusions) to avoid the serious consequnces of iron overload. The average life expectancy in Western cultures is ~ 30 years. There is no cure. In third world countries where a reliable blood supply is unavailable, death occurs earlier. Patients are aften infected with blood born viruses such as hepatitis B, hepatitis C and the AIDS virus, HIV. Sickle cell anaemia is also a very common disease. It is due to a single DNA base mutation at in the beta-globin gene that results in production of normal amounts of a defective beta-globin molecule (HbS). In low oxygen, HbS molecules polymerize in red cells and irreversibly damage them. These red cells get trapped in small blood capillaries throughout the circulation causing small infarcts which results in severe pain and organ damage. The life expectancy is <2 years in the thrid world and ~20-30 years in the west. The irony of these two diseases is that there is a perfectly normal fetal globin gene that has been silenced during fetal life. This grant aims to understand the mechanism of the switch from fetal to adult globin gene usage so it can be reversed in adults with b-thalassemia and sickle cell diseaseRead moreRead less
Determining The Function Of Parasite Proteins At The Membrane Skeleton Of Malaria-infected Red Blood Cells
Funder
National Health and Medical Research Council
Funding Amount
$392,036.00
Summary
Malaria is a serious disease that frequently kills its victim after a bout of high fever and coma. The most vicious form of malaria is caused by a minute parasite called Plasmodium falciparum that lives inside red blood cells. As these parasites grow, they make some dramatic renovations to their red blood cell home that make it become very stiff and sticky. Instead of flowing around the body like normal red blood cells, the infected cells become trapped in small veins and can no longer carry out ....Malaria is a serious disease that frequently kills its victim after a bout of high fever and coma. The most vicious form of malaria is caused by a minute parasite called Plasmodium falciparum that lives inside red blood cells. As these parasites grow, they make some dramatic renovations to their red blood cell home that make it become very stiff and sticky. Instead of flowing around the body like normal red blood cells, the infected cells become trapped in small veins and can no longer carry out their normal job. The ability of the parasite to make red blood cells stiff and sticky is what makes this type of malaria so dangerous, particularly when red cells get stuck in the brain. We plan to look at certain proteins that malaria parasites place on the walls of red blood cells because we think this is what makes them stiff and sticky. We hope this will help with the development of and urgently required ways to cure malaria.Read moreRead less