ExosomeIn eukaryotic
cells, stable RNAs are transcribed with a 3’ extension that is
subsequently trimmed from 3’-to-5’, whereas aberrantly processed RNAs
are subject to rapid degradation. A conserved 300 – 400 kDa
exoribonuclease protein complex, the exosome, plays crucial roles in
both the RNA 3’-end formation and turnover processes (Fig. 1). The
nuclear exosome is required for the 3’ end formation of 5.8S rRNA,
small nuclear and nucleolar RNAs; and involved in the degradation of
inefficiently spliced and hyper– or hypo-adenylated pre-mRNAs. The
cytoplasmic exosome is required for the degradation of normal mRNAs as
well as those containing premature termination codons, lacking
termination codons, or bearing AU-rich elements near the 3’
untranslated region. Using X-ray crystallography and biochemical tools,
we aim to understand the architecture, working mechanism, and the
regulation of this multi-subunit machinery.
Riboswitch
Riboswitches are structured RNAs that recognize specific
small molecules, usually key metabolites inside the cell, and “switch” on or
off gene expression at either transcription or translation level. The discovery
of these short cis-acting RNA elements has drastically changed our
understanding of gene regulation. Majority of riboswitches were found in
prokaryotic genomes, while only a few examples of eukaryotic riboswitches have been
reported. Riboswitches are especially prevalent in Gram-positive bacteria,
exemplified by Bacillus subtilis as a model organism, but also in a number of important
pathogens such as Bacillus anthracis, Staphylococcus, Enterococcus,
Streptococcus, Listeria, Clostridium, and Mycobacterium. This and other characteristics attract increasing
attention to target riboswitches for antibiotic development.
Of the twelve different riboswitch classes that
recognize nine different key metabolites, S-adenosyl methionine (SAM)
riboswitches are the most commonly found. For example, nearly half of all known
riboswitches identified in B. anthracis
bear consensus primary sequence and secondary structures of the S box motif.
Three classes of SAM riboswitches have been found. They contain completely
different secondary structures, representing nature’s three independent RNA
solutions to achieve specific SAM recognition. We recently determined the crystal
structures of two (out of three) SAM riboswitch classes: the E. faecalis SMK and the B. subtilis S-box.
Bacteriophage Phi29 Packaging RNA
How
bacteriophage f29 packages its genome into protein capsids is a
mystery and a marvel. The DNA genome is packed to near crystalline density
inside the capsid, against internal pressure of up to 2000 psi, through a
molecular motor composed of a dodecameric head-tail protein connector, and a
pentameric prohead RNA (pRNA) and ATPase gp16. We are working on the structures
of the packaging motor components to explain how this natural nano-machine
works and why RNA is needed as a component.
Signal Recognition Particle (SRP) RNA-protein (ribonucleoprotein) complexes, such as the ribosome, signal recognition particle (SRP), spliceosome, and telomerase, carry out essential functions inside cells. We are currently investigating the structural mechanism of SRP-mediated co-translational translocation of proteins across or into cell membranes. In this vital cellular process, SRP recognizes the hydrophobic signal sequence of the nascent polypeptide emerging from the ribosome, resulting in transient elongation arrest in eukaryotes, and targets the ribosome to the membrane via a GTP-dependent interaction with the SRP receptor (SR). |
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