Effects of 5'-fluoro-2-deoxyuridine on mitochondrial biology in Caenorhabditis elegans.

5-Fluoro-2'-deoxyuridine (FUdR) is a DNA synthesis inhibitor commonly used to sterilize Caenorhabditis elegans in order to maintain a synchronized aging population of nematodes, without contamination by their progeny, in lifespan experiments. All somatic cells in the adult nematode are post-mitotic and therefore do not require nuclear DNA synthesis. However, mitochondrial DNA (mtDNA) replicates independently of the cell cycle and thus represents a potential target for FUdR toxicity. Inhibition of mtDNA synthesis can lead to mtDNA depletion, which is linked to a number of diseases in humans. Furthermore, alterations in mitochondrial biology can affect lifespan in C. elegans. We characterized the effects of FUdR exposure on mtDNA and nuclear DNA (nucDNA) copy numbers, DNA damage, steady state ATP levels, nematode size, mitochondrial morphology, and lifespan in the germ line deficient JK1107 glp-1(q244) and PE255 glp-4(bn2) strains. Lifespan was increased very slightly by 25 µM FUdR, but was reduced by 400 µM. Both concentrations reduced mtDNA and nucDNA copy numbers, but did not change their ratio. There was no detectable effect of FUdR on mitochondrial morphology. Although both concentrations of FUdR resulted in smaller sized animals, changes to steady-state ATP levels were either not detected or restricted to the higher dose and/or later timepoints, depending on the method employed and strain tested. Finally, we determined the half-life of mtDNA in somatic cells of adult C. elegans to be between 8 and 13 days; this long half-life very likely explains the small or undetectable impact of FUdR on mitochondrial endpoints in our experiments. We discuss the relative pitfalls associated with using FUdR and germline deficient mutant strains as tools for the experimental elimination of progeny.

A variant in the HS1-BP3 gene is associated with familial essential tremor.

BACKGROUND: Genetic linkage studies have identified two susceptibility loci for essential tremor (ET) on chromosomes 3q13 (ETM1) and 2p24.1 (ETM2). Linkage disequilibrium studies in separate population samples from the United States and Singapore suggest an association between ET and loci at ETM2. METHODS: Fine mapping studies were conducted on multiplex and singleton US families linked to ETM2 using newly detected loci within the candidate interval to establish the minimal critical region (MCR) harboring an ET gene. The genes and transcripts within this interval were systematically analyzed by single-strand conformational polymorphism analysis and DNA sequencing. RESULTS: A 464-kb region between loci D2S2150 and etm1231 was defined as the MCR. The coding regions and flanking intronic splice sites of two genes and seven transcripts in this interval were evaluated for mutations. A missense mutation (828C-->G) in the transcript FLJ14249 (HS1-BP3) was identified in one US family. This mutation was found in another apparently unrelated US family with ET and was absent in 150 control samples (300 chromosomes). The 828C-->G mutation causes a substitution of a glycine for an alanine residue in the HS1-BP3 protein. The HS1-BP3 protein binds to proteins that are highly expressed in motor neurons and Purkinje cells and regulate the Ca2+/calmodulin-dependent protein kinase activation of tyrosine and tryptophan hydroxylase. CONCLUSIONS: A rare variant in the HS1-BP3 gene that is associated with essential tremor (ET) in two families is reported. This finding will facilitate research on the functional role of this gene and related genes in the pathogenesis of ET.

Haplotype analysis at the ETM2 locus in a Singaporean sample with familial essential tremor.

An ancestral haplotype on chromosome 2p24.1 described in an American sample with familial essential tremor (ET) was analyzed in a different ethnic sample from Singapore. Six polymorphic loci (etm1240, etm1231, etm1234, APOB, etm1241, and etm1242) in a 274-kb interval within an ET gene candidate region (ETM2) were analyzed in Singaporean individuals with a family history of ET (n = 52) and compared to Singaporean controls older than age 65 (n = 49). The allele frequencies were significantly different between cases and controls for the loci etm1234 (p = 0.0001) and APOB (p = 0.0320). An extended haplotype formed by the loci etm1231, etm1234, and APOB occurred with a frequency of 31% in Singaporean cases and in 1.8% of elderly Singaporean controls (p = 0.0005). Haplotype studies in two different population samples suggest that a disease locus for ET lies near or within the 100-kb interval between the loci etm1231 and APOB.

Candidate genes for recessive non-syndromic mental retardation on chromosome 3p (MRT2A).

A mild type of autosomal recessive, non-syndromic mental retardation (NSMR) is linked to loci on chromosome 3p. This report delimits the MRT2A minimal critical region to 4.2 Mb between loci D3S3630 and D3S1304. This interval contains nine genes (IL5RA, TRNT1, LRRN1, SETMAR, SUMF1, ITPR1, BHLHB2, EDEM, and MRPS36P1). The results suggest that a mutation does not exist in these genes and that an unknown transcript in the region contributes to the cognitive deficits in NSMR.

A successful attempt to raise goat kids free of infection with caprine arthritis encephalitis virus in an endemically infected goat herd.

Goat kids from a herd endemically infected with caprine arthritis encephalitis (CAE) virus were raised according to 3 methods. One group of ten goat kids was removed from infected does at birth before suckling or licking by the doe could occur (snatch birth technique). Kids were fed on goat colostrum, which had been heated to 57 degrees C for ten minutes and then held in a thermos flask for one hour. Subsequently the kids were fed reconstituted spray dried cows' milk powder. They were raised apart from infected goats with separation maintained by a wire fence. Contact occurred across-the-fence. Passively acquired serum antibody to CAE virus was detected in some kids at two to three months of age. Nine of the ten goats were negative for serum antibody to CAE virus when tested at 5-6, 9 and 12 months of age. One goat was positive at three and nine months of age but was negative when tested at 12 months of age. A second group of four kids was removed at birth and fed heat-treated goat colostrum, followed by milk from CAE virus-infected does. All four kids became infected with CAE virus; they developed serum antibody to CAE virus between 5-6 and 9 months of age. A third group of two kids was not removed from their infected dams. Both kids were infected at 5-6 and 9 months of age.