Lab Overview
Work in our group is focused on macromolecular electron microscopy
(EM) studies of the structure of complexes involved in transcription
and its regulation. We are interested in the structure and assembly
of the basal transcription machinery (RNA polymerase II and the
general transcription factors), the Mediator complex that enables
regulation during initiation, and the structure of chromatin
modifying and remodeling complexes involved in regulation of
transcription through their effect on chromatin structure.
Top Story of 2006
A ~ 10Å resolution structure of human RNA polymerase II
and implications for transcription initiation
A 10 Å resolution cryo-EM reconstruction of human RNA polymerase II (hRNAPII)
calculated from images of single hRNAPII particles preserved in amorphous ice
using cryo-electron microscopy (cryo-EM) analysis, provides detailed information
about the conformation of the enzyme in solution. As expected from significant
sequence homology, the structure of hRNAPII is remarkably similar to that of
its yeast counterpart and a number of features in the hRNAPII cryo-EM reconstruction
can be correlated to secondary structure elements in the X-ray structure of yeast
RNA polymerase II (yRNAPII). Of particular interest is the structure of the deep
cleft in which the polymerase active site is located. Contrary to what was observed
in X-ray structure of yRNAPII alone, elements in the active site cleft involved
in interaction with the template DNA and nascent RNA strands are ordered in the
hRNAPII cryo-EM structure, their positions corresponding to those of homologous
elements in the X-ray structure of an elongating yRNAPII/DNA/RNA ternary complex.
The conformation of the active site cleft in the hRNAPII cryo-EM reconstruction
would not allow the template strand to reach the polymerase active site, and
provides a structural rational for the observed interaction of eukaryotic RNA
polymerases with blunt and tailed DNA templates. The conformation of the active
site cleft revealed by our structure implies that a critical function of the
general transcription factors TFIIB and TFIIF at initiation must relate to promoting
a rearrangement of the RNAPII active site cleft that permits interaction of the
enzyme with template DNA.
2006 Publications
Takagi, Y., Calero, G., Komori, H., Brown, J. A., Ehrensberger,
A. H., Hudmon, A., Asturias, F. J., and Kornberg, R. D. (2006)
Head Module Control of Mediator Interactions Mol Cell 23:355-364.
Asturias, F. J. 2006. Mammalian Fatty Acid Synthase: X-ray
Structure of a Molecular Assembly Line. ACS Chem Biol 1:135-138.
Asturias, F. J., Cheung, I., Sabouri, N., Chilkova, O., Wepplo,
D., and Johansson, E. 2006. Structure of Saccharomyces cerevisiae
DNA polymerase epsilon by cryo-electron microscopy. Nat Struct
Mol Biol 13:35-43.
