The outcome of poliovirus infections in K562 cells is cytolytic rather than persistent after hemin-induced differentiation.

K562-Mu erythroleukemia cells readily establish a long-term persistent poliovirus infection characterized by continuous virus production in the absence of complete p220 cleavage and host translation shutoff (R. E. Lloyd and M. Bovee, Virology 194:200-209, 1993). The mechanism of resistance appears to be modulated at the intracellular level and to be related to decreased virus-mediated cytopathic effects (P. A. Benton, J. W. Murphy, and R. E. Lloyd Virology 213:7-18, 1995). It is well documented that hemin induces the differentiation of K562 cells and alters the expression of several host proteins. We report here that growth of K562 cells in hemin prior to poliovirus infection results in a dose-dependent increase in virus-induced cell lysis and thereby alters the normally persistent outcome of infection to a more lytic phenotype. K562 cells infected after hemin treatment displayed increased host translation shutoff, p220 cleavage, viral protein synthesis, and viral RNA accumulation compared with nontreated cells. Since hemin treatment of K562 cells also induced the increased expression of several heat shock proteins (Hsp70, Hsc70, Hsp90, and cohort p60), we tested the hypothesis that their increased expression may play a role in altering poliovirus infection in hemin-treated K562 cells. However, neither heat stress nor oxidative stress, inducers of heat shock protein synthesis, altered the outcome (of virus infections. In addition, we report the novel finding that subunits of two translation initiation factors, p220 (eIF-4G) and eIF-2alpha, are cleaved as a result of hemin treatment of K562 cells. It is proposed that hemin alters the expression of specific host proteins in K562 cells, probably other than heat shock proteins, which changes the initial response to poliovirus infections from persistent to lytic.     

Torque generation in the flagellar motor of Escherichia coli: evidence of a direct role for FliG but not for FliM or FliN.

Among the many proteins needed for assembly and function of bacterial flagella, FliG, FliM, and FliN have attracted special attention because mutant phenotypes suggest that they are needed not only for flagellar assembly but also for torque generation and for controlling the direction of motor rotation. A role for these proteins in torque generation is suggested by the existence of mutations in each of them that produce the Mot- (or paralyzed) phenotype, in which flagella are assembled and appear normal but do not rotate. The presumption is that Mot- defects cause paralysis by specifically disrupting functions essential for torque generation, while preserving the features of a protein needed for flagellar assembly. Here, we present evidence that the reported mot mutations in fliM and fliN do not disrupt torque-generating functions specifically but, instead, affect the incorporation of proteins into the flagellum. The fliM and fliN mutants are immotile at normal expression levels but become motile when the mutant proteins and/or other, evidently interacting flagellar proteins are overexpressed. In contrast, many of the reported fliG mot mutations abolish motility at all expression levels, while permitting flagellar assembly, and thus appear to disrupt torque generation specifically. These mutations are clustered in a segment of about 100 residues at the carboxyl terminus of FliG. A slightly larger carboxyl-terminal segment of 126 residues accumulates in the cells when expressed alone and thus probably constitutes a stable, independently folded domain. We suggest that the carboxyl-terminal domain of FliG functions specifically in torque generation, forming the rotor portion of the site of energy transduction in the flagellar motor.     

Genetic mapping studies of 40 loci and 23 cosmids in chromosome 11p13-11g13, and exclusion of μ-calpain as the multiple endocrine neoplasia type 1 gene

Forty loci (16 polymorphic and 24 non-polymorphic) together with 23 cosmids isolated from a chromosome 11-specific library were used to construct a detailed genetic map of 11p13-11g13. The map was constructed by using a panel of 13 somatic cell hybrids that sub-divided this region into 19 intervals, a meiotic mapping panel of 33 multiple endocrine neoplasia type 1 (MEN1) families (134 affected and 269 unaffected members) and a mitotic mapping panel that was used to identify loss of heterozygosity in 38 MENI-associated tumours. The results defined the most likely order of the 16 loci as being: 11pter-D11S871(D11S288, D11S149)-11cen-CNTF-PGA-ROM1-D11S480-PYGM-SEA-D11S913-D115970-D11S97-D11S146-INT2-D11S971-D11S533-11gter. The meiotic mapping studies indicated that the most likely location of the MEN1 gene was in the interval flanked by PYGM and D11S97, and the results of mitotic mapping suggested a possible location of the MEN1 gene telomeric to SEA. Mapping studies of the gene encoding μ-calpain (CAPN1) located CAPN1 to llg13 and in the vicinity of the MEN1 locus. However, mutational analysis studies did not detect any germ-line CAPN1 DNA sequence abnormalities in 47 unrelated MEN1 patients and the results therefore exclude CAPN1 as the MEN1 gene. The detailed genetic map that has been constructed of the 11p13-11g13 region should facilitate the construction of a physical map and the identification of candidate genes for disease loci mapped to this region.     

Mapping of Unconventional Myosins in Mouse and Human

Myosins are molecular motors that move along filamentous actin. Seven classes of myosin are expressed in vertebrates: conventional myosin, or myosin-II, as well as the 6 unconventional myosin classes -I, -V, -VI, -VII, -IX, and -X. We have mapped in mouse 22 probes encompassing all known unconventional myosins and, as a result, have identified 16 potential unconventional myosin genes. These genes include 7 myosins-I, 2 myosins-V, 1 myosin-VI, 3 myosins-VII, 2 myosins-IX, and 1 myosin-X. The map location of 5 of these genes was identified in human chromosomes by fluorescencein situhybridization.     

Analysis of glycosphingolipid-derived oligosaccharides by high pH anion exchange chromatography

As an adjunct to existing thin layer and column chromatographic methods for the identification of glycolipids a method that utilizes the high pH anion chromatographic (HPAEC) analysis of the oligosaccharides released from the glycolipids by endoglycoceramidase has been developed. Using a Dionex Carbo Pak PA1 column and elution with a linear gradient of sodium acetate in 0.2M NaOH, the elution times of eight neutral and fourteen acidic oligosaccharides derived from glycolipids were determined. Under these conditions the neutral oligosaccharides were well separated from each other but some of the acidic oligosaccharides had overlapping elution times. The ganglioside-derived oligosaccharides could be further identified by treating them with sialidase or by mild acid hydrolysis and reanalysing the products by HPAEC. The method was applied to the analysis of mixed bovine brain gangliosides. The procedure provides an additional approach for the initial identification of glycolipids by analysing the component oligosaccharides rather than the intact glycolipids.     

Biomass and carbon storage of the North American deciduous forest

Field measures of tree and shrub dimensions were used with established biomass equations in a stratified, two-stage cluster sampling design to estimate above-ground ovendry woody biomass and carbon storage of the eastern deciduous forest of North America. Biomass averaged 8.1 ± 1.4 (95% C.I.) kg/m² and totaled 18.1 ± 3.1 (95% C.I.) gigatons. Carbon storage averaged 3.6 ± 0.6 (95% C.I.) kg/m² and totaled 8.1 ± 1.4 (95% C.I.) gigatons. These values are lower than previous estimates commonly used in the analysis of the global carbon budget which range from 17.1 to 23.1 kg/m² for biomass and 7.7 to 10.4 kg/m² for carbon storage. These new estimates for the deciduous forest, together with earlier work in the boreal forest begin to reveal a pattern of overestimation of global carbon storage by vegetation in analyses of the global carbon budget. We discuss reasons for the differences between the new and earlier estimates, as well as implications for our understanding of the global carbon cycle.