The Human Obesity Gene Map: The 1996 Update

An update of the human obesity gene map up to October 1996 is presented. Evidence from Mendelian disorders exhibiting obesity as a clinical feature, single-gene mutation rodent models, quantitative trait loci uncovered in crossbreeding experiments with mouse, rat, and pig models, association and case-control studies with candidate genes, and linkage studies with genes and other markers is reviewed. All chromosomal locations of the animal loci are converted into human genome locations based on syntenic relationships between the genomes. A complete listing of all these loci reveals that only 4 of the 24 human chromosomes are not yet represented, i.e., 9, 18, 21, and Y. Several chromosome arms are characterized by the presence of several putative loci. The following arms include at least three such loci: 1p, 1q, 3p, 4q, 6p, 7q, 8p, 8q, 11p, 11q, 15q, 20q, and Xq. Studies with negative association and linkage results are also reviewed.     

Saikosaponin-d,a novel SERCA inhibitor,induces autophagic cell death in apoptosis-defective cells

Autophagy is an important cellular process that controls cells in a normal homeostatic state by recycling nutrients to maintain cellular energy levels for cell survival via the turnover of proteins and damaged organelles. However, persistent activation of autophagy can lead to excessive depletion of cellular organelles and essential proteins, leading to caspase-independent autophagic cell death. As such, inducing cell death through this autophagic mechanism could be an alternative approach to the treatment of cancers. Recently, we have identified a novel autophagic inducer, saikosaponin-d (Ssd), from a medicinal plant that induces autophagy in various types of cancer cells through the formation of autophagosomes as measured by GFP-LC3 puncta formation. By computational virtual docking analysis, biochemical assays and advanced live-cell imaging techniques, Ssd was shown to increase cytosolic calcium level via direct inhibition of sarcoplasmic/endoplasmic reticulum Ca²⁺ ATPase pump, leading to autophagy induction through the activation of the Ca²⁺/calmodulin-dependent kinase kinase–AMP-activated protein kinase–mammalian target of rapamycin pathway. In addition, Ssd treatment causes the disruption of calcium homeostasis, which induces endoplasmic reticulum stress as well as the unfolded protein responses pathway. Ssd also proved to be a potent cytotoxic agent in apoptosis-defective or apoptosis-resistant mouse embryonic fibroblast cells, which either lack caspases 3, 7 or 8 or had the Bax-Bak double knockout. These results provide a detailed understanding of the mechanism of action of Ssd, as a novel autophagic inducer, which has the potential of being developed into an anti-cancer agent for targeting apoptosis-resistant cancer cells.     

Substrate-trapping techniques in the identification of cellular PTP targets

Tyrosine phosphorylation is negatively regulated by the protein-tyrosine phosphatases (PTPs). In order to find the physiological substrates of these enzymes, diverse PTP mutants that do not possess any catalytic activities but appear to bind tightly to their tyrosine phosphorylated substrates have been designed. Hence, they can be used as tools to pull out their respective substrates from heterogeneous extracts. Named PTP "substrate-trapping" mutants by the Tonks laboratory, they represent a diverse variety of defective PTPs that are epitomized by the Cys to Ser mutant (C/S) where the active cysteine residue of the signature motif is mutated to a serine residue. In addition, new mutants have been developed which are expected to help characterize novel and less abundant substrates. In this article, we review and describe all the different substrate-trapping mutants that have successfully been used or that hold interesting promises. We present their methodology to identify substrates in vivo (co-immunoprecipitation) and in vitro (GST pulldown), and provide a current list of substrates that have been identified using these technologies.     

Chromosomal anomalies induced by the organic phosphate pesticide guthion in Chinese hamster cells

Chromosomal anomalies associated with the use of the organic phosphate pesticide guthion were investigated in Chinese hamster cells (line CHO-K1). Most commonly observed were chromatid breaks and exchanges. Infrequently, mild failure of condensation, despiralization, secondary constriction, gaps, pulverization, ring and dicentric chromosomes were noted. The mean number of chromosome breaks per cell was significantly higher in treated cells than in control cells. Autoradiographic studies revealed that while higher dosages of the chemical (80–120 μg/ml) arrested cells and prevented their movement out of S phase, a lower dosage (60 μg/ml) caused a progression delay. In relation to the relative length of no. 1 and no. 2 chromosomes, no apparent difference existed in the incidence of total breaks between them. However, significant differences in the nonrandom distribution of breaks in no. 1 and no. 2 chromosomes indicated linear differentiation of breakage susceptibility along the chromosomes. An increased concentration of breaks was observed in the long arms of both no. 1 and no. 2 chromosomes. The experimental results suggest that guthion, as well as inducing chromosomal anomalies, may produce a viable mutant.     

The Human Obesity Gene Map: The 1997 Update

An update of the human obesity gene map incorporating published results up to October 1997 is presented. Evidence from Mendelian disorders exhibiting obesity as a clinical feature; single-gene mutation rodent models; quantitative trait loci uncovered in human genome-wide scans and in crossbreeding experiments with mouse, rat, and pig models; association and case-control studies with candidate genes; and linkage studies with genes and other markers is reviewed. All chromosomal locations of the animal loci are converted into human genome locations based on syntenic relationships between the genomes. A complete listing of all of these loci reveals that all but chromosome Y of the 24 human chromosomes are represented. Some chromosomes show at least three putative loci related to obesity on both arms (1, 2, 6, 8, 11, and 20) and several on one chromosome arm only (3p, 4q, 5q, 7q, 12q, 13q, 15q, 15p, 22q, and Xq). Studies reporting negative association and linkage results are also listed, with the exception of the unlinked markers from genome-wide scans.     

The Na(+)-K(+)-ATPase alpha2 gene and trainability of cardiorespiratory endurance: the HERITAGE family study.

The Na(+)-K(+)-ATPase plays an important role in the maintenance of electrolyte balance in the working muscle and thus may contribute to endurance performance. This study aimed to investigate the associations between genetic variants at the Na(+)-K(+)-ATPase alpha2 locus and the response (Delta) of maximal oxygen consumption (VO(2 max)) and maximal power output (W(max)) to 20 wk of endurance training in 472 sedentary Caucasian subjects from 99 families. VO(2 max) and W(max) were measured during two maximal cycle ergometer exercise tests before and again after the training program, and restriction fragment length polymorphisms at the Na(+)-K(+)-ATPase alpha2 (exons 1 and 21-22 with Bgl II) gene were typed. Sibling-pair linkage analysis revealed marginal evidence for linkage between the alpha2 haplotype and DeltaVO(2 max) (P = 0.054) and stronger linkages between the alpha2 exon 21-22 marker (P = 0.005) and alpha2 haplotype (P = 0.003) and DeltaW(max). In the whole cohort, DeltaVO(2 max) in the 3.3-kb homozygotes of the exon 1 marker (n = 5) was 41% lower than in the 8.0/3.3-kb heterozygotes (n = 87) and 48% lower than in the 8.0-kb homozygotes (n = 380; P = 0.018, adjusted for age, gender, baseline VO(2 max), and body weight). Among offspring, 10.5/10.5-kb homozygotes (n = 14) of the exon 21-22 marker showed a 571 +/- 56 (SE) ml O(2)/min increase in VO(2 max), whereas the increases in the 10.5/4.3-kb (n = 93) and 4.3/4.3-kb (n = 187) genotypes were 442 +/- 22 and 410 +/- 15 ml O(2)/min, respectively (P = 0.017). These data suggest that genetic variation at the Na(+)-K(+)-ATPase alpha2 locus influences the trainability of VO(2 max) in sedentary Caucasian subjects.     

Persistent infection of human oligodendrocytic and neuroglial cell lines by human coronavirus 229E

Human coronaviruses (HuCV) cause common colds. Previous reports suggest that these infectious agents may be neurotropic in humans, as they are for some mammals. With the long-term aim of providing experimental evidence for the neurotropism of HuCV and the establishment of persistent infections in the nervous system, we have evaluated the susceptibility of various human neural cell lines to acute and persistent infection by HuCV-229E. Viral antigen, infectious virus progeny and viral RNA were monitored during both acute and persistent infections. The astrocytoma cell lines U-87 MG, U-373 MG, and GL-15, as well as neuroblastoma SK-N-SH, neuroglioma H4, and oligodendrocytic MO3.13 cell lines, were all susceptible to an acute infection by HuCV-229E. The CHME-5 immortalized fetal microglial cell line was not susceptible to infection by this virus. The MO3.13 and H4 cell lines also sustained a persistent viral infection, as monitored by detection of viral antigen and infectious virus progeny. Sequencing of the S1 gene from viral RNA after approximately 130 days of infection showed two point mutations, suggesting amino acid changes during persistent infection of MO3.13 cells but none for H4 cells. Thus, persistent in vitro infection did not generate important changes in the S1 portion of the viral spike protein, which was shown for murine coronaviruses to bear hypervariable domains and to interact with cellular receptor. These results are consistent with the potential persistence of HuCV-229E in cells of the human nervous system, such as oligodendrocytes and possibly neurons, and the virus's apparent genomic stability.