Dominant role of the niche in melanocyte stem-cell fate determination

Stem cells which have the capacity to self-renew and generate differentiated progeny are thought to be maintained in a specific environment known as a niche. The localization of the niche, however, remains largely obscure for most stem-cell systems. Melanocytes (pigment cells) in hair follicles proliferate and differentiate closely coupled to the hair regeneration cycle. Here we report that stem cells of the melanocyte lineage can be identified, using Dct-lacZ transgenic mice, in the lower permanent portion of mouse hair follicles throughout the hair cycle. It is only the population in this region that fulfils the criteria for stem cells, being immature, slow cycling, self-maintaining and fully competent in regenerating progeny on activation at early anagen (the growing phase of hair follicles). Induction of the re-pigmentation process in K14-steel factor transgenic mice demonstrates that a portion of amplifying stem-cell progeny can migrate out from the niche and retain sufficient self-renewing capability to function as stem cells after repopulation into vacant niches. Our data indicate that the niche has a dominant role in the fate determination of melanocyte stem-cell progeny.     

Modulation of the neuronal glutamate transporter EAAC1 by the interacting protein GTRAP3-18

Excitatory amino-acid carrier 1 (EAAC1) is a high-affinity Na+-dependent L-glutamate/D,L-aspartate cell-membrane transport protein. It is expressed in brain as well as several non-nervous tissues. In brain, EAAC1 is the primary neuronal glutamate transporter. It has a polarized distribution in cells and mainly functions perisynaptically to transport glutamate from the extracellular environment. In the kidney it is involved in renal acidic amino-acid re-absorption and amino-acid metabolism. Here we describe the identification and characterization of an EAAC1-associated protein, GTRAP3-18. Like EAAC1, GTRAP3-18 is expressed in numerous tissues. It localizes to the cell membrane and cytoplasm, and specifically interacts with carboxy-terminal intracellular domain of EAAC1. Increasing the expression of GTRAP3-18 in cells reduces EAAC1-mediated glutamate transport by lowering substrate affinity. The expression of GTRAP3-18 can be upregulated by retinoic acid, which results in a specific reduction of EAAC1-mediated glutamate transport. These studies show that glutamate transport proteins can be regulated potently and that GTRAP can modulate the transport functions ascribed to EAAC1. GTRAP3-18 may be important in regulating the metabolic function of EAAC1.     

A sperm ion channel required for sperm motility and male fertility

Calcium and cyclic nucleotides have crucial roles in mammalian fertilization, but the molecules comprising the Ca2+-permeation pathway in sperm motility are poorly understood. Here we describe a putative sperm cation channel, CatSper, whose amino-acid sequence most closely resembles a single, six-transmembrane-spanning repeat of the voltage-dependent Ca2+-channel four-repeat structure. CatSper is located specifically in the principal piece of the sperm tail. Targeted disruption of the gene results in male sterility in otherwise normal mice. Sperm motility is decreased markedly in CatSper-/- mice, and CatSper-/- sperm are unable to fertilize intact eggs. In addition, the cyclic-AMP-induced Ca2+ influx is abolished in the sperm of mutant mice. CatSper is thus vital to cAMP-mediated Ca2+ influx in sperm, sperm motility and fertilization. CatSper represents an excellent target for non-hormonal contraceptives for both men and women.     

A genome-wide approach accounting for body mass index identifies genetic variants influencing fasting glycemic traits and insulin resistance

Recent genome-wide association studies have described many loci implicated in type 2 diabetes (T2D) pathophysiology and β-cell dysfunction but have contributed little to the understanding of the genetic basis of insulin resistance. We hypothesized that genes implicated in insulin resistance pathways might be uncovered by accounting for differences in body mass index (BMI) and potential interactions between BMI and genetic variants. We applied a joint meta-analysis approach to test associations with fasting insulin and glucose on a genome-wide scale. We present six previously unknown loci associated with fasting insulin at P < 5 × 10(-8) in combined discovery and follow-up analyses of 52 studies comprising up to 96,496 non-diabetic individuals. Risk variants were associated with higher triglyceride and lower high-density lipoprotein (HDL) cholesterol levels, suggesting a role for these loci in insulin resistance pathways. The discovery of these loci will aid further characterization of the role of insulin resistance in T2D pathophysiology.     

A conditional knockout resource for the genome-wide study of mouse gene function

Gene targeting in embryonic stem cells has become the principal technology for manipulation of the mouse genome, offering unrivalled accuracy in allele design and access to conditional mutagenesis. To bring these advantages to the wider research community, large-scale mouse knockout programmes are producing a permanent resource of targeted mutations in all protein-coding genes. Here we report the establishment of a high-throughput gene-targeting pipeline for the generation of reporter-tagged, conditional alleles. Computational allele design, 96-well modular vector construction and high-efficiency gene-targeting strategies have been combined to mutate genes on an unprecedented scale. So far, more than 12,000 vectors and 9,000 conditional targeted alleles have been produced in highly germline-competent C57BL/6N embryonic stem cells. High-throughput genome engineering highlighted by this study is broadly applicable to rat and human stem cells and provides a foundation for future genome-wide efforts aimed at deciphering the function of all genes encoded by the mammalian genome.     

An ancient recipe for flood-basalt genesis

Large outpourings of basaltic lava have punctuated geological time, but the mechanisms responsible for the generation of such extraordinary volumes of melt are not well known. Recent geochemical evidence suggests that an early-formed reservoir may have survived in the Earth's mantle for about 4.5 billion years (ref. 2), and melts of this reservoir contributed to the flood basalt emplaced on Baffin Island about 60 million years ago. However, the volume of this ancient mantle domain and whether it has contributed to other flood basalts is not known. Here we show that basalts from the largest volcanic event in geologic history--the Ontong Java plateau--also exhibit the isotopic and trace element signatures proposed for the early-Earth reservoir. Together with the Ontong Java plateau, we suggest that six of the largest volcanic events that erupted in the past 250 million years derive from the oldest terrestrial mantle reservoir. The association of these large volcanic events with an ancient primitive mantle source suggests that its unique geochemical characteristics--it is both hotter (it has greater abundances of the radioactive heat-producing elements) and more fertile than depleted mantle reservoirs-may strongly affect the generation of flood basalts.     

Anaerobic microbial metabolism can proceed close to thermodynamic limits.

Many fermentative bacteria obtain energy for growth by reactions in which the change in free energy (DeltaG') is less than that needed to synthesize ATP. These bacteria couple substrate metabolism directly to ATP synthesis, however, by classical phosphoryl transfer reactions. An explanation for the energy economy of these organisms is that biological systems conserve energy in discrete amounts, with a minimum, biochemically convertible energy value of about -20 kJ mol-1 (refs 1, 2, 3). This concept predicts that anaerobic substrate decay ceases before the minimum free energy value is reached, and several studies support this prediction. Here we show that metabolism by syntrophic associations, in which the degradation of a substrate by one species is thermodynamically possible only through removal of the end product by another species, can occur at values close to thermodynamic equilibrium (DeltaG' approximately 0 kJ mol-1). The free energy remaining when substrate metabolism halts is not constant; it depends on the terminal electron-accepting reaction and the amount of energy required for substrate activation. Syntrophic associations metabolize near thermodynamic equilibrium, indicating that bacteria operate extremely efficient catabolic systems.     

Molecular evidence for new mutation at the hprt locus in Lesch-Nyhan patients

Hypoxanthine-guanine phosphoribosyltransferase (HPRT; EC2.4.2.8), which functions in the metabolic salvage of purines, is encoded by an X-linked gene in man. Partial HPRT deficiencies are associated with gouty arthritis, while absence of activity results in Lesch-Nyhan syndrome (L-N). L-N patients fail to reproduce and the heterozygous state appears to confer no selective advantage. Thus, Haldane's principle predicts that new mutations at the hprt locus must occur frequently in order for L-N syndrome to be maintained in the population. This constant introduction of new mutations would be expected to result in a heterogeneous collection of genetic lesions, some of which may be novel. As we report here, the mutations in the hprt gene of seven L-N patients, selected from an initial survey of 28 patients, have been characterized and all were found to be distinctly different, as predicted. The origin of one unusual mutation has been identified by analysis of DNA from four generations of family members. Further molecular analysis of the origin of new mutations at the hprt locus should aid in resolving the issue of an apparent difference in the frequency of hprt mutations in males and females.