Biomedical subjects

M McCormick-Graham

Publications and source records attributed to M McCormick-Graham.

4 recordsLinked to original sources

PubMed1997

Variable telomeric repeat synthesis in Paramecium tetraurelia is consistent with misincorporation by telomerase.

Telomeric DNA at the ends of chromosomes consist of short, tandem repeat sequences. The telomeres of Paramecium tetraurelia are made up of variable repeats, whereas Paramecium caudatum telomeric repeats are largely invariant. To investigate variable repeat synthesis in P. tetraurelia, mutated telomerase RNA genes were expressed in vivo. We demonstrate that the P. caudatum telomerase RNA can participate in telomere synthesis when expressed in the P. tetraurelia macronucleus, despite 24% primary sequence divergence of the RNAs between the two species. De novo telomeric repeats from transformants indicate that P. tetraurelia telomerase fidelity is dramatically affected by template substitutions and that misincorporation at a single templating position is likely to account for the majority of P. tetraurelia telomeric DNA variability. Furthermore, we show that fidelity is not solely a function of the RNA moiety, as the P. caudatum telomerase RNA does not impart high fidelity to the chimeric enzyme.

Animals↗

PubMed1996

Characterization of homologous DNA recombination activity in normal and immortal mammalian cells.

Homologous DNA recombination levels were measured in normal and spontaneously immortalized murine and human fibroblasts, and in a number of primate and murine established fibroblast cell lines. Immortal cell lines and tumor-derived clones homologously recombined extrachromosomal plasmid substrates at frequencies approximately 100-fold higher than did normal cells. To further explore the mechanism responsible for this phenotype, homologous recombination frequency was measured using nuclear extracts derived from normal and immortalized murine and human fibroblasts. Extracts prepared from immortal cells catalyzed high levels of homologous recombination, whereas very little recombination activity was detected in extracts prepared from normal fibroblasts. Similarly, only extracts derived from immortal cells contained strand-transferase activity as measured by the recently described pairing-on-membrane assay. Mixing experiments indicated that a recombination enhancing factor or factors present in immortal cells, rather than a recombination inhibitor in normal cells, was responsible for the enhanced homologous recombination activity observed using extracts derived from the former.

Animals↗

PubMed1996

A single telomerase RNA is sufficient for the synthesis of variable telomeric DNA repeats in ciliates of the genus Paramecium.

Paramecium telomeric DNA consists largely of a random distribution of TTGGGG and TTTGGG repeats. Given the precise nature of other ciliate telomerases, it has been postulated that there are two distinct types of the Paramecium enzyme, each synthesizing perfect telomeric repeats: one with a template RNA that specifies the addition of TTTGGG and the second dictating the synthesis of TTGGGG repeats. We have cloned and sequenced telomerase RNA genes from Paramecium tetraurelia, P. primaurelia, P. multimicronucleatum, and P. caudatum. Surprisingly, a single gene encodes telomerase RNA in all four species, although an apparently nontranscribed pseudogene is also present in the genome of P. primaurelia. The overall lengths of the telomerase RNAs range between 202 and 209 nucleotides, and they can be folded into a conserved secondary structure similar to that derived for other ciliate RNAs. All Paramecium telomerase RNAs examined include a template specific for the synthesis of TTGGGG telomeric repeats, which has not been posttranscriptionally edited to account for the conventional synthesis of TTTGGG repeats. On the basis of these results, possible mechanisms for the synthesis of variable telomeric repeats by Paramecium telomerase are discussed.

Animals↗

PubMed1995

Ciliate telomerase RNA structural features.

Telomerase RNA is an integral part of telomerase, the ribonucleoprotein enzyme that catalyzes the synthesis of telomeric DNA. The RNA moiety contains a templating domain that directs the synthesis of a species-specific telomeric repeat and may also be important for enzyme structure and/or catalysis. Phylogenetic comparisons of telomerase RNA sequences from various Tetrahymena spp. and hypotrich ciliates have revealed two conserved secondary structure models that share many features. We have cloned and sequenced the telomerase RNA genes from an additional six Tetrahymena spp. (T. vorax, T. borealis, T. australis, T. silvana, T. capricornis and T. paravorax). Inclusion of these sequences, most notably that from T. paravorax, in a phylogenetic comparative analysis allowed us to more narrowly define structural elements that may be necessary for a minimal telomerase RNA. A primary sequence element, positioned 5' of the template and conserved between all previously known ciliate telomerase RNAs, has been reduced from 5'-(C)UGUCA-3' to the 4 nt sequence 5'-GUCA-3'. Conserved secondary structural features and the impact they have on the general organization of ciliate telomerase RNAs is discussed.

Animals↗