action and adjuvants for treating infections and
Note added in proof: After this manuscript was
submitted, groups of F. Sicheri and R. Silverman
published a structural analysis of porcine RNase L.
This work reports a similar crossed homodimer
but observes different KEN/KEN interactions (20)
due to a conformational difference presumably
caused by RNA binding.
References and Notes
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Acknowledgments: We thank F. Hughson (Princeton
University) and A. Korostelev (University of Massachusetts,
Worcester) for reading the manuscript and making valuable
suggestions. We thank the staff of Brookhaven National
Laboratory for providing access to a synchrotron x-ray
source. We are grateful to our colleagues at the
Department of Molecular Biology for stimulating discussions
and an excellent research environment. We thank
Princeton University for supporting and funding our work.
Coordinates have been deposited to the Protein Data
Bank with accession codes 4OAU and 4OAV. Author
contributions: Y.H., J.D., and A.K. determined the structures
of RNase L. S.R. carried out biochemical studies of model
RNA substrates. G.W. and A.C. carried out studies in HeLa
cells. Y.H. and A.K. wrote the manuscript. A.K. supervised
Materials and Methods
Figs. S1 to S12
17 December 2013; accepted 12 February 2014
Published online 27 February 2014;
Fig. 4. Cleavage of biological targets. (A) Site selection by RNase L in
mammalian ribosomes and hepatitis C virus RNA. (B) Total RNA from HeLa
cells cotransfected with plasmids encoding RNase L mutants or not encoding a
protein (–), and 2-5A. Protein expression levels were analyzed by Western blot
(WB). The bar graph shows compiled data from in vitro assays in Fig. 2A and
fig. S4D. (C) Cleavage of substrates DIII and DIV by RNase L and Ire1KR32. Bar
charts show quantification of cleavage. Error bars show mean T SE of a single-
exponential fitting (RNase L) and two time courses (Ire1KR32).