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East London, South Africa

Uhlmann F.,The Francis Crick Institute
Nature Reviews Molecular Cell Biology | Year: 2016

SMC (structural maintenance of chromosomes) complexes — which include condensin, cohesin and the SMC5–SMC6 complex — are major components of chromosomes in all living organisms, from bacteria to humans. These ring-shaped protein machines, which are powered by ATP hydrolysis, topologically encircle DNA. With their ability to hold more than one strand of DNA together, SMC complexes control a plethora of chromosomal activities. Notable among these are chromosome condensation and sister chromatid cohesion. Moreover, SMC complexes have an important role in DNA repair. Recent mechanistic insight into the function and regulation of these universal chromosomal machines enables us to propose molecular models of chromosome structure, dynamics and function, illuminating one of the fundamental entities in biology. © 2016 Nature Publishing Group, a division of Macmillan Publishers Limited. All Rights Reserved. Source


Grant
Agency: GTR | Branch: BBSRC | Program: | Phase: Research Grant | Award Amount: 314.75K | Year: 2016

This research project is aimed at understanding the mechanism of sex determination in the chicken i.e. the series of molecular events that determine whether the embryonic gonad develops as a testis or as an ovary. It is widely recognized that such primary sex determining mechanisms evolve rapidly, as exemplified by the marked differences between mammals and other vertebrates. Gonadogenesis in mammals is envisaged as a linear process that is dependent on a switch mechanism based on the Y-chromosome gene Sry. If Sry is expressed appropriately, then the developing gonad becomes a testis: without Sry the gonad becomes an ovary. With only limited exceptions, extensive studies to identify similar master switch genes in other vertebrate species have been unsuccessful. In our studies, we have demonstrated that chicken cells acquire an inherent sex identity at fertilisation or shortly thereafter and believe that this is key to avian sex determination; i.e. the testis forms because the genital ridge is composed of male cells or the ovary forms because the genital ridge is composed of female cells. This suggests that, in birds, rather than gonadal sex determination depending on a sex-specific switch mechanism, testis and ovary differentiation represent two separate pathways. It was widely accepted that once gonadal fate had been determined, it was permanent, but some surprising recent findings suggest that this is not the case. In fact, the adult mammalian gonad displays a great degree of plasticity and testicular and ovarian identity has to be maintained throughout life. It appears that this maintenance depends on the expression of two genes, DMRT1 in males or FOXL2 in female. Dmrt1 has also been shown to be necessary for the proper development and survival of male germ cells. Dmrt1 and Foxl2 are not thought to be important for primary sex determination events in the mouse embryo, but they have been shown to play key roles in gonadogenesis in several vertebrate species including the chick and some mammals. It may be that the requirement for DMRT1 and FOXL2 to maintain adult mammalian gonads represent an evolutionary residue of their major roles in primary sex determining mechanisms in lower vertebrates, where plasticity is often evident during embryonic stages. It is also possible that, unlike the mouse, a number of mammalian species retain elements or this earlier primary sex determining system. We will investigate the possibility that Dmrt1 and Foxl2 balance of expression determines the sexual fate of the embryonic gonads in birds. To do this, we will use cutting edge methods of genetic manipulation to delete copies of Dmrt1 and Foxl2 from the genome of chicken germ cells (PGC) and use these germ cells to derive birds with these genetic mutations. We will assess the effects of these deletions on PGC growth in culture and on germ cell development after injection of PGCs into embryos. Injected embryos will be hatched and raised to sexual maturity and crossed with wild-type birds: by selective crossing we can generate birds with either one or no copies of Dmrt1 or Foxl2. We will compare the development of the gonads and germ cells in these manipulated birds with that in wild-type male and female birds, and so determine the effect of Dmrt1 and Foxl2 on primary sex determination and germ cell development in birds. We will also carry out a series of molecular analyses to determine the networks of genes regulated by Dmrt1 and Foxl2, and identify the genes affected by manipulating the expression levels/ balance of these transcription factors.


Grant
Agency: GTR | Branch: BBSRC | Program: | Phase: Research Grant | Award Amount: 318.76K | Year: 2016

Understanding the biology of the metabolic network is key for biotechnology, where single cellular organisms such as budding yeast are used to produce proteins, vaccines or antibiotics. A metabolic network formed from similar reactions operates in mammalian cells, and changes during a lifetime being considered a main driver of ageing and age-associated disorders. Here we are applying for an industrial/academic partnership that will bring a new level into the understanding of this largest of all cellular systems, by creating an enzyme-centric quantitative map that spans the yeast genome. With our industrial partner Sciex, we establish a unique technological platform that can quantify 80% of metabolic enzymes in less than 30 minutes. We will apply this platform to measure enzymes in a collection of ~4800 yeast strains, each of which is lacking one gene at a time. In this way, we connect the majority of all genes in the genome with the metabolites and metabolic enzymes they affect. This map will be the most comprehensive investigation into a eukaryotic proteome conducted so far, and address both already known genes, and genes for which there is only little or no functional information so far available. We will learn about the function of new genes in two ways, first by studying their direct impact on the proteome and metabolism, and by associating them with the already known genes on the basis of their proteomic footprint. For these reasons, the project is of unique value to the mass spec manufacturing industry, that seeks possibilities to bring proteomic technology into environmental analytics, to biotechnology, that lacks information about metabolic networks so that they can exploit it for improving production cycles, and for basic science, that will gain unique insights into the function of novel genes and can use it to develop new strategies for addressing ageing-associated disease.


Grant
Agency: GTR | Branch: BBSRC | Program: | Phase: Research Grant | Award Amount: 417.48K | Year: 2014

The gastrointestinal tract is a vital organ that converts our diet into useful digestible nutrients, contributes to the maintenance of water balance and protects our body from pathogenic microorganisms that are present within the lumen of the gut, along with large numbers of beneficial bacteria. In order for the gut to carry out its essential functions, it contains exquisitely specialised cells, including epithelial cells, immune cells, nerve cells and muscle cells. Intestinal epithelial cells are tightly connected to each other to form a sophisticated gatekeeping system that allows the selective transport of nutrients and water but keeps away harmful toxins or pathogenic bacteria. Immune cells constantly monitor the lumen and the wall of the gut and respond in case the essential intestinal barrier is breached. Finally, complex networks of nerve cells within the gut wall are responsible for generating intestinal movements that are essential for proper digestive function by activating the musculature of the gut wall. Since the intestinal epithelium is constantly exposed to harmful substances and pathogenic microorganisms, it is quite vulnerable and is often damaged. Normally this does not have detrimental consequences for an organism since all cells of the intestinal epithelium are continuously replenished by stem cells that are dedicated to producing constantly fresh epithelial cells. Although the continuous regeneration of the intestinal epithelium is essential for maintaining it in good working order, other cell types play a major role in keeping them healthy. In particular, glial cells, which normally accompany and support nerve cells in all parts of the nervous system, are also found in the vicinity of intestinal epithelial cells and release substances that are essential for maintaining the intestinal epithelial barrier; if these enteric glial cells are eliminated in experimental conditions, the barrier breaks down and animals die from acute inflammation of the small intestine. In addition, several studies have suggested that the inflammation that accompanies common gut diseases, such as Crohns disease or ulcerative colitis, may also involve the abnormal interaction of glial cells with intestinal epithelial cells and immune cells. These observations support the idea that despite their specialised functions, the different cell types that make up the gut wall (and indeed any organ) need to work in concert in order to support its physiological roles. Despite the important roles of the intestinal glial cells in supporting the critical functions of the nerve cells and the epithelium of the gut, very little is known about their biology in healthy individuals and in disease situations. In this proposal we will aim at filling this knowledge gap by building on some of our own recent observations. In particular, we will identify and characterise the properties of the gliogenic stem cells which generate new glial cells throughout life. We will also identify conditions and signals that modulate the behaviour of intestinal glial cells. Finally, we plan to characterise molecules which are located within the nucleus and are important for these cells to maintain their properties and continue to generate new glial cells throughout adult life. Normal digestive function depends on the fine balance between the loss of old and the production of new cells in the different gut tissues and the optimal cross talk between the different cell types. Breakdown of such an equilibrium results in uncontrolled growth of cells (cancer), severe inflammation of the gut wall (inflammatory bowel disease-IBD) or inability of the gut wall to protect the internal environment of an organisms from toxic substances or pathogenic bacteria. Understanding how local glial cells contribute to the integrity and normal function of gut tissues, we can ultimately use these cells as a means to alter the course of common debilitating gastrointestinal disorders.


Grant
Agency: GTR | Branch: BBSRC | Program: | Phase: Research Grant | Award Amount: 349.11K | Year: 2014

White blood cells are a key part of the bodys immune system, playing a critical role in combating infections by viruses, bacteria and other pathogens. Our research is focussed on a type white blood cell called a T cell, which can detect the presence of an infectious agent such as a virus inside other cells. We are interested in understanding the mechanism by which a T cell recognises whether another cell contains an infectious agent within it. As part of this process, the T cell and the infected cell have to adhere tightly to each other and we are studying this process of adhesion. In recent work we have identified that a protein called Wnk1 plays an important role in regulating T cell adhesion, and in the research we are now proposing, we will extend this work by examining how the T cells function when they lack the protein, and whether this makes them defective in contributing to normal immune responses. We suspect that Wnk1 may also control how the T cells migrate through the body. Normal T cells are continuously on the move, trafficking between the blood and lymph nodes and the spleen. This continuous movement is a vital feature of the T cells as it allows them to patrol the body looking for signs of infection. We will investigate this potential function of the Wnk1 protein in controlling T cell migration. We suspect that Wnk1 functions as a signalling protein, transmitting signals from the surface of the cell to adhesion molecules. We will investigate this hypothesis further by looking for other proteins to which Wnk1 transmits a signal. Overall our aim is to understand how Wnk1 controls T cell adhesion and migration. Such knowledge will reveal new opportunities for the design of rational drugs that could modulate the function of T cells and thereby the whole immune system.

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