Telomerase copies a short template within its integral telomerase RNA onto eukaryotic chromosome ends, compensating for incomplete replication and degradation. Telomerase action extends the proliferative potential of cells, and thus it is implicated in cancer and aging. Nontemplate regions of telomerase RNA are also crucial for telomerase function. However, they are highly divergent in sequence among species, and their roles are largely unclear. Using in silico three-dimensional modeling, constrained by mutational analysis, we propose a three-dimensional model for a pseudoknot in telomerase RNA of the budding yeast Kluyveromyces lactis. Interestingly, this structure includes a U-A ⅐ U major-groove triple helix. We confirmed the triple-helix formation in vitro using oligoribonucleotides and showed that it is essential for telomerase function in vivo. While triplex-disrupting mutations abolished telomerase function, triple compensatory mutations that formed pH-dependent G-C ⅐ C ؉ triples restored the pseudoknot structure in a pH-dependent manner and partly restored telomerase function in vivo. In addition, we identified a novel type of triple helix that is formed by G-C ⅐ U triples, which also partly restored the pseudoknot structure and function. We propose that this unusual structure, so far found only in telomerase RNA, provides an essential and conserved telomerasespecific function.Telomerase, a ribonucleoprotein reverse transcriptase, makes up for losses caused by incomplete DNA replication and degradation, by adding species-specific, 5-to 26-nucleotide (nt) repeats onto the telomere termini (reviewed in reference 2). The telomerase complex contains an RNA subunit (TER) (TLC1 in Saccharomyces cerevisiae), a catalytic reverse transcriptase (TERT) (Est2 in S. cerevisiae), and several other protein components. Unlike other reverse transcriptases, telomerase specializes in repeatedly copying a short RNA template within its integral RNA component.TERs are highly divergent, being conserved in sequence only among closely related species. Phylogenetic covariation was used to predict conserved secondary structures for evolutionarily close species of ciliates (14, 24), vertebrates (4), Kluyveromyces budding yeasts of the K. marxianus cluster (31, 32), and Saccharomyces sensu stricto (3,7,13,37). Limited similarity in the general architecture was observed among these models, consisting of three long arms and a catalytic core domain (4, 13). Although nontemplate regions are essential for the assembly, regulation, and function of telomerase, their specific roles are still unclear (reviewed in reference 30). We hypothesized that important functional elements would exhibit better conservation in their tertiary structures, rather than in their secondary structures or sequences. Solving these tertiary structures may provide insights into their conserved functions.Pseudoknot elements were found to be critical for telomerase function in ciliates (28), vertebrates (4), and Kluyveromyces (32). For S. cerevisiae, alternative pseud...

BackgroundImprovement in the left ventricular ejection fraction (LVEF) may occur in patients with dilated cardiomyopathy (DCM).HypothesisThere are different implications of persistent versus transient improvement in LVEF among DCM patients receiving contemporary therapy.MethodsWe studied 188 patients with nonischemic DCM. Persistent improvement in LVEF (PIEF) was defined as LVEF increase by at least 10% compared to baseline, and found in 2 separate echo‐Doppler exams performed at least 12 months apart. Increased LVEF in echo 2, which was not sustained in echo 3, was defined as transient improvement in LVEF (TIEF).ResultsOver an average follow‐up of 6.8 years, PIEF occurred in 61 (33%) patients, predicting a better long‐term outcome (P < 0.001) in a combined end‐point comprising death, heart transplantation, or the need for a ventricular assist device. The TIEF group had an intermediate course and were closer to nonimprovers (P = 0.003 vs PIEF). Multivariate logistic regression identified the following independent predictors of PIEF: shorter disease duration, pregnancy‐associated disease, left ventricular hypertrophy, and baseline LVEF ≤25%. A score to predict PIEF assigned 1 point to each of the following: disease duration <3 years and no familial cardiomyopathy; pregnancy‐associated presentation; basal LVEF ≤25%; and left ventricular wall thickness ≥12. A score of ≥3 was present in 44% of the patients, reliably predicting PIEF in 91% (P = 0.01).ConclusionsPersistent improvement in LVEF is associated with improved long‐term prognosis. Baseline clinical parameters can be used to identify patients likely to demonstrate PIEF, thereby allowing tailored management in this population.