RNA Structure and Function 283
example of “antisense” targeting, by which a cellular RNA is manipulated through simple base pairing
with a small oligonucleotide.
7.5.3 Small RNAs Involved in Gene Silencing and Regulation
Eukaryotic cells contain abundant small RNAs of 22 nucleotides in length. These RNAs are used to con-
trol the timing of protein expression, to inhibit the attack of viruses and endogenous transposons, and even
to influence the function of DNA, through chromatin silencing.^64
miRNAsare small pieces of single-stranded RNA that base pair with target genes, thereby modulating
levels of protein synthesis.^64 The miRNAs are encoded by larger, highly conserved transcripts that contain
many long stem-loop structures (Figure 7.34). The functional miRNA molecules are excised by a special-
ized endonuclease called Dicer, which cuts duplex RNA into short segments of 22 nucleotides.
Invading viruses and transposons may also form RNA duplexes that are cleaved by the Dicer enzyme,
which results in short RNA molecules that are called small interfering RNAs (siRNAs).^65 These siRNAs
become incorporated into an RNP called the RNAi-induced silencing complex (RISC), which uses the
siRNA as a guide for identifying and degrading an invading RNA of complementary sequence. A RISC
complex also forms around short miRNAs, and is believed to play a role in translational repression of
endogenous gene expression. RNA interference (RNAi) is therefore a powerful strategy for host defense
and cellular regulation (Section 5.7.2).
RNAi is now widely used for genetic manipulation. To silence(eliminate) expression of a particular
gene, it is often sufficient to transfect cells with an siRNA that is complementary in sequence to an mRNA
target. The siRNA can be chemically synthesized (Section 4.2) or produced from a plasmid (Section 4.3).
Once the siRNA is introduced, the cellular RISC machinery responds to the siRNA and causes cleavage
of the target RNA (Section 5.7.2).
7.6 RNA Structure and Function in Viral Systems
Many of the most important discoveries about the diversity of RNA function have come from studies on
viruses. Due to their small, compact genomes and the rapidity of their evolution, viruses have taken full
advantage of the many chemical and structural capabilities of RNA.
7.6.1 RNA as an Engine Part: The Bacteriophage Packaging Motor
In addition to the many other attributes of RNA, it can also act as a molecular motor. All organisms con-
tain motor enzymes that carry out mechanical work and which undergo conformational changes that are
coupled with ATP binding and hydrolysis. These nanomachines are typically made of protein, but in sev-
eral important cases, they contain essential RNA components. The clearest example of an RNA engine
part is the bacteriophage packaging motor.^66
Bacteriophages are a family of viruses that attack bacteria. After replication, many phages (such as
phi29) have a remarkable mechanism for packagingprogeny DNA into the capsidshell that will encase
a new viral particle. The capsid shell is made up of viral proteins and a collar structure is added, through
which DNA is then sucked rapidly and with great force into the capsid shell (Figure 7.35). Indeed, the phi29
packaging motoris by far the most powerful molecular motor known, capable of pulling against a load of
50 pN.^67 The collar structure contains a number of important components, one of which is a ring of RNA
molecules. Cryoelectron microscopy and biochemical studies have suggested that this RNA ring is an
oligomeric structure that is composed of repeating units of the pRNA, which is encoded by the phage
(Figure 7.36).^66 Currently, the precise role of pRNA and its rotational movement within the collar struc-
ture are undefined. The RNA may play a passive role by creating an anionic corridor that prevents DNA
adhesion to the collar, thereby speeding its passage. Alternatively, pRNA may actively translocate DNA