6 research outputs found
Molecular basis of RNA polymerase III transcription repression by Maf1 & Structure of human mitochondrial RNA polymerase
Topic I
Molecular basis of RNA polymerase III transcription repression by Maf1
RNA polymerase III (RNAP III) is a conserved 17-subunit enzyme that transcribes genes encoding short
untranslated RNAs such as transfer RNAs (tRNAs) and 5S ribosomal RNA (rRNA). These genes are essential
and involved in fundamental processes like protein biogenesis; hence RNAP III activity needs to be tightly
regulated. RNAP III is repressed upon stress and this is regulated by Maf1, a protein conserved from yeast to
humans. Many stress pathways were shown to converge on Maf1 and result in its phosphorylation, followed by
its nuclear import and eventual repression of RNAP III activity. However, the molecular mechanisms of this
repression activity were not known at the beginning of these studies.
This work establishes the mechanism of RNAP III specific transcription repression by Maf1. The
crystal structure of Maf1 was solved. It has a globular fold with surface accessible NLS sequences, which sheds
new light on already published results and explains how stress-induced phopshorylation leads to import of Maf1
into the nucleus. Additionally, cryo EM studies and competition assays show that Maf1 binds RNAP III at its
clamp domain and thereby induces structural rearrangements of RNAP III, which inhibits the interaction with
Brf1, a subunit of the transcription initiation factor TFIIIB. This specifically impairs recruitment of RNAP III to
its promoters and implies that Maf1 is a repressor of transcription initiation. Competition and transcription
assays show that Maf1 also binds RNAP III that is engaged in transcription, leaving RNAP III activity intact but
preventing re-initiation.
Topic II
Structure of human mitochondrial RNA polymerase
The nuclear-encoded human mitochondrial RNAP (mitoRNAP) transcribes the mitochondrial genome, which
encodes rRNA, tRNAs and mRNAs. MitoRNAP is a single subunit (ss) polymerase, related to T7 bacteriophage
and chloroplast polymerases. All share a conserved C-terminal core, whereas the N-terminal parts of mitoRNAP
do not show any homology to other ss RNAPs. Unlike phage RNAPs, which are self-sufficient, human
mitoRNAP needs two essential transcription factors for initiation, TFAM and TFB2M. Both of these factors are
likely to control the major steps of transcription initiation, promoter binding and melting. Thus human
mitoRNAP has evolved a different mechanism for transcription initiation and exhibits a unique transcription
system. Structural studies thus far concentrated on the nuclear enzymes or phage RNAPs, whereas the structure
of mitochondrial RNA polymerase remained unknown. The structural organization of human mitoRNAP and the
molecular mechanisms of promoter recognition, binding and melting were subject of interest in these studies.
In this work the crystal structure of human mitoRNAP was solved at 2.4 Å resolution and reveals a
T7-like C-terminal catalytic domain, a N-terminal domain that remotely resembles the T7 promoter-binding
domain (PBD), a novel pentatricopeptide repeat (PPR) domain, and a flexible N-terminal extension.
MitoRNAP specific adaptions in the N-terminus include the sequestering of one of the key promoter binding
elements in T7 RNAP, the AT-rich recognition loop, by the PPR domain. This sequestration and repositioning of
the N-terminal domain explain the need for the additional initiation factor TFAM. The highly conserved active
site within the C-terminal core was observed to bind a sulphate ion, a well known phosphate mimic, and thereby
suggests conserved substrate binding and selection mechanisms between ss RNAPs. However, conformational
changes of the active site were observed due to a movement of the adjacent fingers subdomain. The structure
reveals a clenching of the active site by a repositioned fingers subdomain and an alternative position of the
intercalating -hairpin. This explains why the conserved transcription factor TFB2M is required for promoter
melting and initiation. A model of the mitochondrial initiation complex was build to further explore the initiation
mechanism, and to rationalize the available biochemical and genetic data.
The structure of mitoRNAP shows how this enzyme uses mechanisms for transcription initiation that
differ from those used by phage and cellular RNAPs, and which may have enabled regulation of mitochondrial
gene transcription and adaptation of mitochondrial function to changes in the environment
The James Webb Space Telescope Mission
Twenty-six years ago a small committee report, building on earlier studies,
expounded a compelling and poetic vision for the future of astronomy, calling
for an infrared-optimized space telescope with an aperture of at least .
With the support of their governments in the US, Europe, and Canada, 20,000
people realized that vision as the James Webb Space Telescope. A
generation of astronomers will celebrate their accomplishments for the life of
the mission, potentially as long as 20 years, and beyond. This report and the
scientific discoveries that follow are extended thank-you notes to the 20,000
team members. The telescope is working perfectly, with much better image
quality than expected. In this and accompanying papers, we give a brief
history, describe the observatory, outline its objectives and current observing
program, and discuss the inventions and people who made it possible. We cite
detailed reports on the design and the measured performance on orbit.Comment: Accepted by PASP for the special issue on The James Webb Space
Telescope Overview, 29 pages, 4 figure
The James Webb Space Telescope Mission
Twenty-six years ago a small committee report, building on earlier studies, expounded a compelling and poetic vision for the future of astronomy, calling for an infrared-optimized space telescope with an aperture of at least 4 m. With the support of their governments in the US, Europe, and Canada, 20,000 people realized that vision as the 6.5 m James Webb Space Telescope. A generation of astronomers will celebrate their accomplishments for the life of the mission, potentially as long as 20 yr, and beyond. This report and the scientific discoveries that follow are extended thank-you notes to the 20,000 team members. The telescope is working perfectly, with much better image quality than expected. In this and accompanying papers, we give a brief history, describe the observatory, outline its objectives and current observing program, and discuss the inventions and people who made it possible. We cite detailed reports on the design and the measured performance on orbit