Exploiting Oxytricha trifallax nanochromosomes to screen for non-coding RNA genes

We took advantage of the unusual genomic organization of the ciliate Oxytricha trifallax to screen for eukaryotic non-coding RNA (ncRNA) genes. Ciliates have two types of nuclei: a germ line micronucleus that is usually transcriptionally inactive, and a somatic macronucleus that contains a reduced, fragmented and rearranged genome that expresses all genes required for growth and asexual reproduction. In some ciliates including Oxytricha, the macronuclear genome is particularly extreme, consisting of thousands of tiny ‘nanochromosomes’, each of which usually contains only a single gene. Because the organism itself identifies and isolates most of its genes on single-gene nanochromosomes, nanochromosome structure could facilitate the discovery of unusual genes or gene classes, such as ncRNA genes. Using a draft Oxytricha genome assembly and a custom-written protein-coding genefinding program, we identified a subset of nanochromosomes that lack any detectable protein-coding gene, thereby strongly enriching for nanochromosomes that carry ncRNA genes. We found only a small proportion of non-coding nanochromosomes, suggesting that Oxytricha has few independent ncRNA genes besides homologs of already known RNAs. Other than new members of known ncRNA classes including C/D and H/ACA snoRNAs, our screen identified one new family of small RNA genes, named the Arisong RNAs, which share some of the features of small nuclear RNAs

Deficiency of a Niemann-Pick, Type C1-related Protein in Toxoplasma Is Associated with Multiple Lipidoses and Increased Pathogenicity

Several proteins that play key roles in cholesterol synthesis, regulation, trafficking and signaling are united by sharing the phylogenetically conserved ‘sterol-sensing domain’ (SSD). The intracellular parasite Toxoplasma possesses at least one gene coding for a protein containing the canonical SSD. We investigated the role of this protein to provide information on lipid regulatory mechanisms in the parasite. The protein sequence predicts an uncharacterized Niemann-Pick, type C1-related protein (NPC1) with significant identity to human NPC1, and it contains many residues implicated in human NPC disease. We named this NPC1-related protein, TgNCR1. Mammalian NPC1 localizes to endo-lysosomes and promotes the movement of sterols and sphingolipids across the membranes of these organelles. Miscoding patient mutations in NPC1 cause overloading of these lipids in endo-lysosomes. TgNCR1, however, lacks endosomal targeting signals, and localizes to flattened vesicles beneath the plasma membrane of Toxoplasma. When expressed in mammalian NPC1 mutant cells and properly addressed to endo-lysosomes, TgNCR1 restores cholesterol and GM1 clearance from these organelles. To clarify the role of TgNCR1 in the parasite, we genetically disrupted NCR1; mutant parasites were viable. Quantitative lipidomic analyses on the ΔNCR1 strain reveal normal cholesterol levels but an overaccumulation of several species of cholesteryl esters, sphingomyelins and ceramides. ΔNCR1 parasites are also characterized by abundant storage lipid bodies and long membranous tubules derived from their parasitophorous vacuoles. Interestingly, these mutants can generate multiple daughters per single mother cell at high frequencies, allowing fast replication in vitro, and they are slightly more virulent in mice than the parental strain. These data suggest that the ΔNCR1 strain has lost the ability to control the intracellular levels of several lipids, which subsequently results in the stimulation of lipid storage, membrane biosynthesis and parasite division. Based on these observations, we ascribe a role for TgNCR1 in lipid homeostasis in Toxoplasma

Hypoxanthine-guanine phosophoribosyltransferase (HPRT) deficiency: Lesch-Nyhan syndrome

Deficiency of hypoxanthine-guanine phosphoribosyltransferase (HPRT) activity is an inborn error of purine metabolism associated with uric acid overproduction and a continuum spectrum of neurological manifestations depending on the degree of the enzymatic deficiency. The prevalence is estimated at 1/380,000 live births in Canada, and 1/235,000 live births in Spain. Uric acid overproduction is present inall HPRT-deficient patients and is associated with lithiasis and gout. Neurological manifestations include severe action dystonia, choreoathetosis, ballismus, cognitive and attention deficit, and self-injurious behaviour. The most severe forms are known as Lesch-Nyhan syndrome (patients are normal at birth and diagnosis can be accomplished when psychomotor delay becomes apparent). Partial HPRT-deficient patients present these symptoms with a different intensity, and in the least severe forms symptoms may be unapparent. Megaloblastic anaemia is also associated with the disease. Inheritance of HPRT deficiency is X-linked recessive, thus males are generally affected and heterozygous female are carriers (usually asymptomatic). Human HPRT is encoded by a single structural gene on the long arm of the X chromosome at Xq26. To date, more than 300 disease-associated mutations in the HPRT1 gene have been identified. The diagnosis is based on clinical and biochemical findings (hyperuricemia and hyperuricosuria associated with psychomotor delay), and enzymatic (HPRT activity determination in haemolysate, intact erythrocytes or fibroblasts) and molecular tests. Molecular diagnosis allows faster and more accurate carrier and prenatal diagnosis. Prenatal diagnosis can be performed with amniotic cells obtained by amniocentesis at about 15–18 weeks' gestation, or chorionic villus cells obtained at about 10–12 weeks' gestation. Uric acid overproduction can be managed by allopurinol treatment. Doses must be carefully adjusted to avoid xanthine lithiasis. The lack of precise understanding of the neurological dysfunction has precluded development of useful therapies. Spasticity, when present, and dystonia can be managed with benzodiazepines and gamma-aminobutyric acid inhibitors such as baclofen. Physical rehabilitation, including management of dysarthria and dysphagia, special devices to enable hand control, appropriate walking aids, and a programme of posture management to prevent deformities are recommended. Self-injurious behaviour must be managed by a combination of physical restraints, behavioural and pharmaceutical treatments

Altered purine and pyrimidine metabolism in erythrocytes with purine nucleoside phosphorylase deficiency

Purine and pyrimidine metabolism was compared in erythrocytes from three patients from two families with purine nucleoside phosphorylase deficiency and T-cell immunodeficiency, one heterozygote subject for this enzyme deficiency, one patient with a complete deficiency of hypoxanthine-guanine phosphoribosyltransferase, and two normal subjects. The erythrocytes from the heterozygote subject were indistinguishable from the normal erythrocytes. The purine nucleoside phosphorylase deficient erythrocytes had a block in the conversion of inosine to hypoxanthine. The erythrocytes with 0.07% of normal purine nucleoside phosphorylase activity resembled erythrocytes with hypoxanthine-guanine phosphoribosyltransferase deficiency by having an elevated intracellular concentration of PP-ribose-P, increased synthesis of PP-ribose-P, and an elevated rate of carbon dioxide release from orotic acid during its conversion to UMP. Two hypotheses to account for the associated immunodeficiency—that the enzyme deficiency leads to a block of PP-ribose-P synthesis or inhibition of pyrimidine synthesis—could not be supported by observations in erythrocytes from both enzyme-deficient families.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/44134/1/10528_2004_Article_BF00484238.pd

THE EFFECTS OF PLYOMETRIC TRAINING ON MUSCLE ACTIVATION CHARACTERISTICS IN POST- PUBESCENT ADOLESCENT FEMALES

J. Savage1, J. Seegmiller1, C. McGowan1, R. Baker1, J. McNeal2, J. May1 1University of Idaho, Moscow, ID; 2Eastern Washington University, Cheney, WA Previous research has demonstrated that females are at an increased risk for anterior cruciate ligament (ACL) injury. One modifiable risk factor proposed is neuromuscular control. Coactivation of the quadriceps and hamstrings is suggested to be a favorable movement strategy, thereby, reducing the risk of ACL injury. PURPOSE: To determine how a 6-week plyometric training intervention may influence muscle activation onset, duration, and time to peak activation in the right (R) and left (L) vastus lateralis (VL), vastus medialis (VM), biceps femoris (BF) and semitendinosus (SEM), during a jump-landing task in post-pubescent adolescent females. METHODS: Electromyography was used to record onset, duration, and time to peak muscle activation of the right and left limbs during a jump-landing task, prior to and following a 6-week plyometric training intervention in 19 post-pubescent adolescent females. Repeated-measures ANOVAs were used to identify if significant differences in activation characteristics. RESULTS: Following a six-week plyometric training intervention, participants demonstrated decreased time to peak activation. Time to peak activation was approximately 0.55 ms earlier in the RSEM than in the RVL (2.67±.691, p = .017), 0.39 ms earlier in the RBF than in the RVM (2.81±.770, p = .049), .053 ms earlier in the RSEM than in the RVM (2.67±.691, p = .029), 0.85 ms earlier in the LBF than in the LVL (2.69±.942, p = .006), 0.89 ms earlier in the LSEM than in the LVL (2.40+1.08, p = .019), and 1.03 ms earlier in the LBF than in the LVM (2.69±.942, p = .006). CONCLUSION: Following plyometric training, participants exhibited a shortened time to peak activation of the hamstrings, an antagonist, generally responsible for terminal deceleration. The noticeable reduction in time to peak activation as seen in the hamstrings following plyometric training, is indicative of a modified recruitment pattern and likely a result of the prescribed neuromuscular training

THE EFFECTS OF PLYOMETRIC TRAINING ON MUSCLE ACTIVATION CHARACTERISTICS IN POST- PUBESCENT ADOLESCENT FEMALES

J. Savage1, J. Seegmiller1, C. McGowan1, R. Baker1, J. McNeal2, J. May1 1University of Idaho, Moscow, ID; 2Eastern Washington University, Cheney, WA Previous research has demonstrated that females are at an increased risk for anterior cruciate ligament (ACL) injury. One modifiable risk factor proposed is neuromuscular control. Coactivation of the quadriceps and hamstrings is suggested to be a favorable movement strategy, thereby, reducing the risk of ACL injury. PURPOSE: To determine how a 6-week plyometric training intervention may influence muscle activation onset, duration, and time to peak activation in the right (R) and left (L) vastus lateralis (VL), vastus medialis (VM), biceps femoris (BF) and semitendinosus (SEM), during a jump-landing task in post-pubescent adolescent females. METHODS: Electromyography was used to record onset, duration, and time to peak muscle activation of the right and left limbs during a jump-landing task, prior to and following a 6-week plyometric training intervention in 19 post-pubescent adolescent females. Repeated-measures ANOVAs were used to identify if significant differences in activation characteristics. RESULTS: Following a six-week plyometric training intervention, participants demonstrated decreased time to peak activation. Time to peak activation was approximately 0.55 ms earlier in the RSEM than in the RVL (2.67±.691, p = .017), 0.39 ms earlier in the RBF than in the RVM (2.81±.770, p = .049), .053 ms earlier in the RSEM than in the RVM (2.67±.691, p = .029), 0.85 ms earlier in the LBF than in the LVL (2.69±.942, p = .006), 0.89 ms earlier in the LSEM than in the LVL (2.40+1.08, p = .019), and 1.03 ms earlier in the LBF than in the LVM (2.69±.942, p = .006). CONCLUSION: Following plyometric training, participants exhibited a shortened time to peak activation of the hamstrings, an antagonist, generally responsible for terminal deceleration. The noticeable reduction in time to peak activation as seen in the hamstrings following plyometric training, is indicative of a modified recruitment pattern and likely a result of the prescribed neuromuscular training