39 research outputs found
Insights into the evolution of mammalian telomerase: Platypus TERT shares similarities with genes of birds and other reptiles and localizes on sex chromosomes
Background The TERT gene encodes the catalytic subunit of the telomerase complex and is responsible for maintaining telomere length. Vertebrate telomerase has been studied in eutherian mammals, fish, and the chicken, but less attention has been paid to other vertebrates. The platypus occupies an important evolutionary position, providing unique insight into the evolution of mammalian genes. We report the cloning of a platypus TERT (OanTERT) ortholog, and provide a comparison with genes of other vertebrates. Results The OanTERT encodes a protein with a high sequence similarity to marsupial TERT and avian TERT. Like the TERT of sauropsids and marsupials, as well as that of sharks and echinoderms, OanTERT contains extended variable linkers in the N-terminal region suggesting that they were present already in basal vertebrates and lost independently in rayfinned fish and eutherian mammals. Several alternatively spliced OanTERT variants structurally similar to avian TERT variants were identified. Telomerase activity is expressed in all platypus tissues like that of cold-blooded animals and murine rodents. OanTERT was localized on pseudoautosomal regions of sex chromosomes X3/Y2, expanding the homology between human chromosome 5 and platypus sex chromosomes. Synteny analysis suggests that TERT co-localized with sex-linked genes in the last common mammalian ancestor. Interestingly, female platypuses express higher levels of telomerase in heart and liver tissues than do males. Conclusions OanTERT shares many features with TERT of the reptilian outgroup, suggesting that OanTERT represents the ancestral mammalian TERT. Features specific to TERT of eutherian mammals have, therefore, evolved more recently after the divergence of monotremes.Radmila Hrdličková, Jiří Nehyba, Shu Ly Lim, Frank Grützner, Henry R Bose J
Yeast telomerase is capable of limited repeat addition processivity
Telomerase is a ribonucleoprotein reverse transcriptase responsible for the maintenance of one strand of telomere terminal repeats. Telomerase-mediated sequence addition is dictated by a short ‘template’ region of the RNA component. Despite the short template segment, telomerases from many organisms have been shown to mediate the synthesis of long extension products. This synthesis presumably depends on two types of translocation events: simultaneous translocation of the RNA–DNA duplex relative to the active site after each nucleotide incorporation (type I or nucleotide addition processivity), and translocation of the RNA relative to the DNA product after each round of repeat synthesis (type II or repeat addition processivity). In contrast, telomerases from yeasts have been shown to synthesize mostly short products, implying a defect in one or both types of translocation. In this report, we analyzed the processivity of yeast telomerase in vitro, and identified two position-specific elongation barriers within the 5′ region of the RNA template that can account for the synthesis of incomplete first round products. These barriers respond differently to variations in nucleotide concentration, primer sequence and mutations in the catalytic protein subunit, consistent with their having distinct mechanistic bases. In addition, by using optimal primers and high concentrations of dGTP, we were able to detect significant type II translocation by the yeast enzyme. Thus, the difference between the elongation property of yeast and other telomerases appears to be quantitative rather than qualitative. Our results suggest that yeast may be a useful system for investigating the physiologic significance of repeat addition processivity
Recommended from our members
Modeling of gold nanocrystal assemblies in superlattices and vesicles, and the synthesis of nanocrystals for low-temperature solar cell fabrication
Recently, nanocrystal (NC) research has grown substantially, due to their unique and diverse properties. Their flexibility has led to a wide set of proposed applications, such as contrast agents in biomedical fields, ordered nanostructures for microelectronics/plasmonics, and as a cheaper alternative to chemical vapor deposition (CVD) methods. While great advancements have been made in utilizing NCs, three challenges often arise – difficulty in characterizing complex nano-systems, a lack of theoretical exploration as to how or why nanocrystals assemble, and challenges in exploiting the benefits of nanocrystals while minimizing disadvantageous properties. This dissertation will address each of these issues in specific systems.
First, thorough work has been done suggesting that hydrophobic gold nanocrystals can be encapsulated in vesicle bilayers. However, the primary characterization method for this system is Cryo-Transmission Electron Microscopy, which cannot provide adequate resolution and contrast to fully characterize nanostructures. Here, small angle x-ray scattering is explored as a method for revealing detailed information regarding the bilayer structure.
vii
Next gold nanocrystal superlattices are explored through molecular dynamics (MD) simulations. While many works have shown crystal structure transitions in a variety of systems, a detailed explanation as to why certain crystal structures are preferred has yet to be provided. This work offers detailed MD simulations to reveal details regarding the packing density of various crystal structures and to estimate diffusion coefficients in various packings. Furthermore, the free energy difference between BCC and FCC configurations for a small set of gold nanocrystals is explored by thermodynamic integration. The simulated properties are also compared to a small set of real systems.
The second half of this dissertation addresses practical applications of NCs for photovoltaics. Despite manufacturing benefits, it is well known that the small NC grains and insulating capping ligands make it difficult to produce efficient solar cells. Therefore, two approaches to removing these ligands and growing nanocrystal grains are explored. The first approach focuses on further studying CuInSe2 synthesis as a window into grain growth. The second offers an example of a material with favorable properties for grain growth – Cu3BiS3 – and addresses difficulties in producing it.Chemical Engineerin
Self-Assembly and Thermal Stability of Binary Superlattices of Gold and Silicon Nanocrystals
Simple
hexagonal (sh) AB<sub>2</sub> binary superlattices (BSLs)
of organic ligand-capped silicon (A; 5.40(±9.8%) nm diameter)
and gold (B; 1.88(±10.1%) nm diameter) nanocrystals were assembled
by evaporation of colloidal dispersions and characterized using transmission
electron microscopy (TEM) and grazing incidence small-angle X-ray
scattering (GISAXS). When deposited on tilted substrates by slow evaporation,
the sh-AB<sub>2</sub> superlattice contracted slightly toward the
substrate with centered orthorhombic structure. Heating the BSL to
200 °C in air led to gold coalescence and segregation to the
surface of the assembly without disrupting the Si nanocrystal sublattice,
thus creating a sh superlattice of Si nanocrystals
Chains, Sheets, and Droplets: Assemblies of Hydrophobic Gold Nanocrystals with Saturated Phosphatidylcholine Lipid and Squalene
Assemblies of saturated 1,2-diacylphosphatidylcholine
lipid and
hydrophobic dodecanethiol-capped 1.8 nm diameter gold nanocrystals
were studied as a function of lipid chain length and the addition
of the naturally occurring oil, squalene. The gold nanocrystals formed
various lipid-stabilized agglomerates, sometimes fusing with lipid
vesicle bilayers. The nanocrystal assembly structure depended on the
hydrocarbon chain length of the lipid fatty acids. The lipid with
the shortest fatty acid length studied, dilauroylphosphatidylcholine,
created extended chains of gold nanocrystals. The lipid with slightly
longer fatty acid chains created planar sheets of nanocrystals. Further
increases of the fatty acid chain length led to spherical agglomerates.
The inclusion of squalene led to lipid- and nanocrystal-coated oil
droplets
Chloroform-Enhanced Incorporation of Hydrophobic Gold Nanocrystals into Dioleoylphosphatidylcholine (DOPC) Vesicle Membranes
Vesicles of dioleoylphosphatidylcholine (DOPC) formed
by extrusion
(liposomes) with hydrophobic alkanethiol-capped Au nanocrystals were
studied. Dodecanethiol-capped 1.8-nm-diameter Au nanocrystals accumulate
in the lipid bilayer, but only when dried lipid–nanocrystal
films were annealed with chloroform prior to hydration. Without chloroform
annealing, the Au nanocrystals phase separate from DOPC and do not
load into the liposomes. Au nanocrystals with slightly longer capping
ligands of hexadecanethiol or with a larger diameter of 4.1 nm disrupted
vesicle formation and created lipid assemblies with many internal
lamellar attachments
The Role of Ligand Packing Frustration in Body-Centered Cubic (bcc) Superlattices of Colloidal Nanocrystals
This paper addresses the assembly
of body centered-cubic (bcc)
superlattices of organic ligand-coated nanocrystals. First, examples
of bcc superlattices of dodecanethiol-capped Au nanocrystals and oleic
acid-capped PbS and PbSe nanocrystals are presented and examined by
transmission electron microscopy (TEM) and grazing incidence small-angle
X-ray scattering (GISAXS). These superlattices tend to orient on their
densest (110) superlattice planes and exhibit a significant amount
of {112} twinning. The same nanocrystals deposit as monolayers with
hexagonal packing, and these thin films can coexist with thicker bcc
superlattice layers, even though there is no hexagonal plane in a
bcc lattice. Both the preference of bcc in bulk films over the denser
face-centered cubic (fcc) superlattice structure and the transition
to hexagonal monolayers can be rationalized in terms of packing frustration
of the ligands. A model is presented to calculate the difference in
entropy associated with capping ligand packing frustration in bcc
and fcc superlattices