Targeting peptide nucleic acid (PNA) oligomers to mitochondria within cells by conjugation to lipophilic cations: implications for mitochondrial DNA replication, expression and disease

The selective manipulation of mitochondrial DNA (mtDNA) replication and expression within mammalian cells has proven difficult. One promising approach is to use peptide nucleic acid (PNA) oligomers, nucleic acid analogues that bind selectively to complementary DNA or RNA sequences inhibiting replication and translation. However, the potential of PNAs is restricted by the difficulties of delivering them to mitochondria within cells. To overcome this problem we conjugated a PNA 11mer to a lipophilic phosphonium cation. Such cations are taken up by mitochondria through the lipid bilayer driven by the membrane potential across the inner membrane. As anticipated, phosphonium–PNA (ph–PNA) conjugates of 3.4–4 kDa were imported into both isolated mitochondria and mitochondria within human cells in culture. This was confirmed by using an ion-selective electrode to measure uptake of the ph–PNA conjugates; by cell fractionation in conjunction with immunoblotting; by confocal microscopy; by immunogold-electron microscopy; and by crosslinking ph–PNA conjugates to mitochondrial matrix proteins. In all cases dissipating the mitochondrial membrane potential with an uncoupler prevented ph–PNA uptake. The ph–PNA conjugate selectively inhibited the in vitro replication of DNA containing the A8344G point mutation that causes the human mtDNA disease ‘myoclonic epilepsy and ragged red fibres’ (MERRF) but not the wild-type sequence that differs at a single nucleotide position. Therefore these modified PNA oligomers retain their selective binding to DNA and the lipophilic cation delivers them to mitochondria within cells. When MERRF cells were incubated with the ph–PNA conjugate the ratio of MERRF to wild-type mtDNA was unaffected, even though the ph–PNA content of the mitochondria was sufficient to inhibit MERRF mtDNA replication in a cell-free system. This unexpected finding suggests that nucleic acid derivatives cannot bind their complementary sequences during mtDNA replication. In summary, we have developed a new strategy for targeting PNA oligomers to mitochondria and used it to determine the effects of PNA on mutated mtDNA replication in cells. This work presents new approaches for the manipulation of mtDNA replication and expression, and will assist in the development of therapies for mtDNA diseases.

‘Quite destitute and … very desirous of going to North America’ : The Roots and Repercussions of Emigration from Sutherland and Caithness

Peer reviewed

La sexualidad como variable importante en la calidad de vida (C.V) de las personas con discapacidad

La afectividad y la sexualidad se han de desarrollar adecuadamente en los individuos aunque no estén en plenas condiciones físicas y psíquicas, indiferentemente si residen en su domicilio o en un centro especializado. La atención a la discapacidad no siempre ha estado ligada al concepto de cuidados de enfermería, pero en la actualidad se considera indispensable hacerlo desde la defensa de los derechos de las personas con Diversidad Funcional (D.F.), ratificados en la Convención de Ginebra de 2008. En este trabajo se presentan datos que forma parte de una tesis doctoral en la que se estudia las alteraciones de la sexualidad en personas con D.F. y como ésta influye en su calidad de vida (CV); Planteado desde el marco enfermero de los Patrones Funcionales de Marjory Gordon, por visualizar los procesos vitales siempre desde una visión integral de salud, guiando de forma lógica al diagnóstico enfermero, contemplando al paciente desde una visión holística y recogiendo toda la información necesaria del paciente, familia y entorno incluyendo su sexualidad.

Poolesville, Maryland and vicinity, 1862 (Raster Image)

This layer is a georeferenced raster image of the untitled historic paper manuscript map: [Map of Poolesville, Maryland and vicinity showing the position of Union brigades]. It was sketched by Union General George Stoneman, Oct. 24th, 1862. Scale not given. Covers a portion of Montgomery County, Maryland from Poolesville to the Potomac River. The image inside the map neatline is georeferenced to the surface of the earth and fit to the Maryland State Plane Coordinate System (in Meters) (Fipszone 1900). All map collar and inset information is also available as part of the raster image, including any inset maps, profiles, statistical tables, directories, text, illustrations, or other information associated with the principal map. This map shows features such as roads, towns and villages, drainage, troop location, and more. This layer is part of a selection of digitally scanned and georeferenced historic maps of the Civil War from the Harvard Map Collection. Many items from this selection are from a collection of maps deposited by the Military Order of the Loyal Legion of the United States Commandery of the State of Massachusetts (MOLLUS) in the Harvard Map Collection in 1938. These maps typically portray both natural and manmade features, in particular showing places of military importance. The selection represents a range of regions, originators, ground condition dates, scales, and purposes.

Jefferson County, Kentucky, 1910 (Raster Image)

This layer is a georeferenced raster image of the historic, topographic paper map entitled: Topography of Jefferson County, Kentucky : from U.S. Geological Survey topographic atlas sheets surveyed in 1904-1910, U.S. Geological Survey ; in cooperation with Kentucky Geological Survey, C. J. Norwood, director. It was published by U.S. Geological Survey in 1912. Scale 1:62,500. The image inside the map neatline is georeferenced to the surface of the earth and fit to the Kentucky North State Plane NAD 1983 coordinate system (in Feet) (Fipszone 1601). All map collar and inset information is also available as part of the raster image, including any inset maps, profiles, statistical tables, directories, text, illustrations, index maps, legends, or other information associated with the principal map. This is a typical topographic map portraying both natural and manmade features. It shows and names works of nature, such as mountains, valleys, lakes, rivers, vegetation, etc. It also identify the principal works of humans, such as roads, railroads, boundaries, transmission lines, major buildings, etc. Relief is shown with standard contour intervals of 20 feet and spot heights. This layer is part of a selection of digitally scanned and georeferenced historic maps from The Harvard Map Collection as part of the Imaging the Urban Environment project. Maps selected for this project represent major urban areas and cities of the world, at various time periods. These maps typically portray both natural and manmade features at a large scale. The selection represents a range of regions, originators, ground condition dates, scales, and purposes.

Tu Fu, the autobiography of a Chinese poet, A.D. 712-770 ... /

Vol. II has title and imprint : Travels of a Chinese poet : Tu Fu, guest of rivers and lakes, A.D. 712-770 by Florence Ayscough ; illustrated from etchings by Lucille Douglass.

An epistolary discourse containing a new resolution and improvement of the grand apocalyptical question, concerning the rise and fall of Rome papal /

Early editions published under title: Apocalyptical key.

Columbus, Ohio; report on a survey of the city government,

Mode of access: Internet.

Extension of the Huffman tree construction algorithm /

Bibliography: p. 42-43.

An investigation of the application of microprocessors to low cost area navigation capabilities for general aviation /

Vita.

The statistical theory of relative errors in floating-point computation /

Thesis (M. S.)--University of Illinois at Urbana-Champaign.

The handling of cherries,

"Awarded second prize."

Volcanoes of Hawaii National Park /

"Special issue."

The Tamworth narrative (New Hampshire)

Includes bibliography.