In This Issue
BIOCHEMISTRY
Roadblocks to DNA repair
The excision and repair of mismatched base pairs is an essential task of cell maintenance. Several models have been proposed to describe how the hemimethylated d(GATC) sequence can direct Escherichia coli mismatch repair while residing on either side of a mismatch, at a separation distance of 1,000 bp or more. Anna Pluciennik and Paul Modrich report that a protein roadblock and breaks in the DNA double helix can disrupt the repair process, which could eliminate some models from consideration. The authors placed a hydrolytically defective form of EcoRI endonuclease on the helix between a d(GATC) sequence and a mismatch to attempt to block signaling along the helix. The blocking protein inhibited MutH endonuclease activation up to 80% of the time. Using a circular DNA strand containing a mismatch and one d(GATC) site, they also showed that a break in the double helix located within the shorter path between the two sites blocked MutH activation, but a break placed within the longer path did not. The authors suggest that signals are transmitted along the helix contour and that it is unlikely that DNA bending is responsible for the signaling. — P.D.
EcoRI experimental system.
“Protein roadblocks and helix discontinuities are barriers to the initiation of mismatch repair” by Anna Pluciennik and Paul Modrich (see pages 12709–12713)
BIOPHYSICS
Fast-scan AFM observations of restriction enzyme
The restriction enzyme EcoP15I cleaves DNA efficiently only when one copy of the enzyme, bound to a restriction site, makes contact with another copy bound to a site as far as 3,500 bp away. Exactly how the two copies arrange to meet has been an open question. When bound to the restriction site, EcoP15I can translocate DNA (which requires ATP), but some scientists believe that DNA supercoiling in the small initial loop would prevent translocation. In addition, consumption of ATP by EcoP15I is remarkably low. Neal Crampton et al. now observe, by fast-scan atomic force microscopy (AFM), that EcoP15I can proceed along DNA by a combination of active translocation and diffusive looping. Two nonspecific sites on the enzyme each allow it to bind a loop of DNA. The two copies of EcoP15I can thus shorten their separation by 2,200 bp and cover the remaining distance by translocation. The authors used AFM cantilevers, fabricated by micromachining, with sharp tips deposited by electron beam lithography. The resulting tips were extraordinarily stiff, with a resonant frequency of 600–1,000 Hz underwater. A scan rate of 1–3 frames per second was achieved, which permitted observation of near-freely diffusing EcoP15I. — K.M.
Translocation and formation of an extruded loop by EcoP15I.
“Fast-scan atomic force microscopy reveals that the type III restriction enzyme EcoP15I is capable of DNA translocation and looping” by Neal Crampton, Masatoshi Yokokawa, David T. F. Dryden, J. Michael Edwardson, Desirazu N. Rao, Kunio Takeyasu, Shige H. Yoshimura, and Robert M. Henderson (see pages 12755–12760)
IMMUNOLOGY
Immune system alters circadian clock
The immune system's first response to many infectious agents is the all too familiar fever and fatigue. Gionata Cavadini et al. show that the fatigue that characterizes some illnesses and autoimmune diseases might be the result of the body's own molecules acting on circadian clock genes. Like other body rhythms, sleep cycles are controlled by the circadian clock. Previous research has implicated the cytokine TNF-α in causing sleepiness. To determine how TNF-α might exert its effects, the authors exposed cells to TNF-α. The cytokine suppressed the expression of several clock genes, an effect that was duplicated in vivo in mice treated with TNF-α. Further investigation showed that TNF-α only suppresses clock genes that have a specific promoter region known as the E-box. Although it is still unclear whether increased rest is a boon to host defense, Cavadini et al. note that increased fatigue decreases quality of life in many autoimmune diseases. Further exploring this functional link between the immune system and fatigue may help explain the effects associated with certain autoimmune diseases, such as multiple sclerosis. — T.H.D.
“TNF-α suppresses the expression of clock genes by interfering with E-box-mediated transcription” by Gionata Cavadini, Saskia Petrzilka, Philipp Kohler, Corinne Jud, Irene Tobler, Thomas Birchler, and Adriano Fontana (see pages 12843–12848)
MEDICAL SCIENCES
Ferreting out HCV's internal accomplices
Viruses are consummate hijackers, co-opting a cell's own machinery to reproduce themselves. The hepatitis C virus (HCV) is particularly adept at hiding inside cells and is able to establish chronic infection 70% of the time. Determining the host cell proteins, or cofactors, that HCV takes advantage of is important in learning how the virus co-opts the cell. Glenn Randall et al. used an RNA interference (RNAi) screen to determine the host proteins that are important for HCV infection. The authors investigated 62 genes, many of which encoded proteins known to interact with HCV. To determine how these genes affect viral replication, Randall et al. systematically silenced each gene by using small interfering RNAs (siRNAs). Twenty-six of the genes appear to modulate the reproduction levels of HCV inside cells. Rather than abrogating infection, the RNAi pathway itself appears to be essential for optimal HCV production. Targeting host genes that unwittingly assist in viral infection could offer an alternate method of treating or preventing HCV infection, the authors say. — T.H.D.
“Cellular cofactors affecting hepatitis C virus infection and replication” by Glenn Randall, Maryline Panis, Jacob D. Cooper, Timothy L. Tellinghuisen, Karen E. Sukhodolets, Sebastien Pfeffer, Markus Landthaler, Pablo Landgraf, Sherry Kan, Brett D. Lindenbach, Minchen Chien, David B. Weir, James J. Russo, Jingyue Ju, Michael J. Brownstein, Robert Sheridan, Chris Sander, Mihaela Zavolan, Thomas Tuschl, and Charles M. Rice (see pages 12884–12889)
SUSTAINABILITY SCIENCE
Humans co-opt earth's biomass production
Measuring human appropriation of net primary production, the aggregate impact of land use on biomass available each year in ecosystems, is one way to quantify the effect that human dominance has on the biosphere. Human land use, such as planting crops, or harvesting, such as clearing forests, alters patterns and pathways of carbon captured by photosynthesis. A recent analysis by Helmut Haberl et al. shows that humans appropriate almost a quarter of the earth's photosynthetic production capacity in this way. Haberl et al. analyzed data on human land use and harvests from 161 countries, which represent 97% of the earth's landmass. The results showed that humans appropriated 24% of the earth's potential production. Over half of the impact is attributable to harvesting crops or other plants. According to the authors, no other single species has such a large impact on the earth's production. The authors caution that, with such an already high human pressure on ecosystems, schemes to replace fossil fuels with biomass fuels should be approached cautiously given their ability to impact the biosphere further. — T.H.D.
Biomass harvesting in Saskatoon, Canada.
“Quantifying and mapping the human appropriation of net primary production in earth's terrestrial ecosystems” by Helmut Haberl, K. Heinz Erb, Fridolin Krausmann, Veronika Gaube, Alberte Bondeau, Christoph Plutzar, Simone Gingrich, Wolfgang Lucht, and Marina Fischer-Kowalski (see pages 12942–12947)
