Page 70 - Hematology_ Basic Principles and Practice ( PDFDrive )

42 Part I Molecular and Cellular Basis of Hematology

nuclear export of mRNA is for correct hematopoietic development, poly(A) tail is thought to promote mRNA degradation by facilitating
mutations or deletions in Nups can result in MDS and leukemia. For attack by both the exosome complex and the decapping complex.
example, point mutations of Nup98 in hematopoietic precursors Rapid mRNA degradation via AU-rich elements is a critical mecha-
results in myelodysplasia and eventual AML. Furthermore, multiple nism for preventing the overproduction of potent cytokines such as
translocations involving Nup98 (up to 29 recognized partners) have tumor necrosis factor (TNF) and GM-CSF. AU-rich elements also
been found in patients with MDS and AML as the sole cytogenetic regulate the synthesis of mRNA for proto-oncogenic transcription
abnormality. factors like c-Jun and c-Fos. The AU elements in the mRNA of these
Naked RNA cannot be exported through NPC channels. Rather, genes mediate destruction of their transcripts in quiescent cells,
RNA export from the nucleus requires that newly synthesized RNAs preventing inappropriate cell proliferation that would occur if Fos/
undergo the previously described processing steps: 5′ capping, splic- Jun were still active.
ing, and 3′ polyadenylation. In addition, RNA binding proteins are Eukaryotic mRNA messages are also subject to surveillance for
required to fold and shuttle the modified RNA through NPCs. accuracy by a mechanism termed nonsense-mediated decay (NMD).
Several of these RNA binding proteins have been identified as The NMD complex surveys the transcript for the presence of prema-
important in hematopoiesis. For example, the eukaryotic translation ture stop codons (nonsense codons) in the message. These premature
initiation factor 4E (eIF4E) enhances nuclear export of specific RNA stop codons can arise via either incomplete splicing mutations in
transcripts and is critical for proper granulocyte differentiation. DNA, transcription errors, or leaky scanning by the ribosome,
Overexpression of eIF4E impedes myeloid maturation and can result causing frame shifts. Detection of a premature stop codon by NMD
in AML. Inhibiting eIF4E with ribavirin has shown activity in early- triggers mRNA degradation by 5′ decapping, 3′ poly(A) tail removal,
phase clinical trials of AML and may represent a promising novel or endonucleolytic cleavage.
class of leukemia therapy. Translational efficiency can be regulated by cellular factors that
bind mRNA in a sequence-specific manner. Iron metabolism is an
excellent example of how cells coordinate uptake and sequestration
RNA METABOLISM of an essential metabolite in response to availability. Transferrin is a
plasma protein that carries iron. Receptors for transferrin (TfR) are
RNA does not live forever, and that is a good thing. In mammalian expressed on cells requiring iron for maturation, such as erythroid
cells, mRNA lifetimes range from several minutes to days. The progenitor cells. They mediate internalization of transferrin loaded
limited lifetime of mRNA enables a cell to alter protein synthesis in with iron into the cytoplasm through receptor-mediated endocytosis.
response to its changing needs. The stability of mRNA is regulated When a cell becomes iron deficient, a Kreb cycle enzyme, aconitase,
by the untranslated regions (UTRs) of mRNA. UTRs are sections of is structurally altered, becoming an iron-responsive protein (IRP) so
the mRNA before the start codon (5′) and after the stop codon (3′) that it can bind to iron-responsive elements (IREs) in the UTR of
that are not translated. These regions govern mRNA half-life, local- transferrin receptor (TfR) mRNA (Fig. 4.6). UTR binding leads to
ization, and translational efficiency. Translational efficiency—both stabilization of the TfR mRNA transcript, thus allowing greater
enhancement and inhibition—can be controlled by UTRs. Both availability for translation, which results in increased protein expres-
proteins and small RNA species can bind to either the 5′ or 3′ UTRs, sion. However, when a cell has sufficient iron, aconitase is not altered,
and these can either regulate translation or influence survival of the and TfR mRNA becomes unstable and prone to degradation. There-
transcript. There are several fascinating mechanisms by which this fore, in that situation, TfR receptor expression is low and the fewer
occurs, and these will be described later. UTR sequence regulation of receptors import less iron.
mRNA survival is essential for proper hematopoietic differentiation.
The best example of this is globin synthesis, where its mRNA is quite
stable because of UTR sequences. This long half-life meets the needs MICRO-RNA
of reticulocytes to synthesize globin for up to 2 days after terminally
mature erythroblasts lose the ability to make new mRNA. In the last two decades another powerful mechanism of regulation of
Some of the elements contained in UTRs form a characteristic gene expression at the RNA level has been discovered. In this mecha-
secondary structure that alters the survival of the mRNA transcript. nism small RNA molecules, termed micro-RNA (miRNA), bind to
One class of these mRNA elements, the riboswitches, directly bind complementary sequences on target mRNA transcripts. This binding
the small molecules that their mRNA encodes enzymes that regulate results in either degradation or inhibition of translation, and conse-
its synthesis. For example, the mRNA for several enzymes in the quent silencing of gene expression. There are roughly 1000 miRNA
cobalamine pathway has riboswitches that bind adenosylcobalamine, molecules coded in the human genome, indicating how robust this
and this regulates the survival of these mRNAs. Thus, in states of regulatory mechanism is. These miRNAs usually contain 18 to 25
high cobalamine, there is decreased survival of the mRNA for enzymes nucleotides, and each miRNA has the potential to target about 500
used in this synthetic pathway. genes. Conversely, an estimated 60% of all mRNAs have one or more
Another class of UTR secondary structures that regulate stability sequences that are predicted to interact with miRNAs. This principle,
is exemplified by the prothrombin 3′ UTR. This mRNA is constitu- often termed RNA interference (RNAi), has also been very useful in
tively polyadenylated at seven or more positions, and the 3′ UTR the laboratory, allowing investigators to repress the expression of
folds into at least two distinct stem-loop conformations. These specific genes to study artificially induced phenotypes. In these
alternate structures expose a consensus binding site for trans-acting studies, small interfering RNAs (siRNA) are synthetically created to
factors, like heterogeneous nuclear ribonucleoprotein 1 (hnRNP-I), bind to homologous sequences within specific mRNAs. These are
polypyrimidine tract-binding protein-1 (PTB-1), and nucleolinin, then transfected into cells, where they mediate destruction of their
with translational regulatory properties. Another type of 3′ UTR target mRNA through endogenous ribonucleases. Repression of gene
regulatory sequence involves selenocysteine insertion sequence expression in this manner has become known as “gene knock-down,”
(SECIS) elements. These represent another stem-loop RNA structure and is widely used to define the function of genes by assessing what
found in mRNA transcripts that serve as protein binding sites on function the cell lacks in the absence of the expression of the target
UTR segments that direct the ribosome to translate the codon UGA gene.
as selenocysteines rather than as a stop codon. An example of this miRNAs are produced from transcripts that form stem-loop
regulation can be found in selenoprotein P in plasma. structures, whereas siRNAs are produced from long double-stranded
Another class of UTR binding site that affects the stability of the RNA (dsRNA) precursors (Fig. 4.7). Similarly, both miRNAs and
mRNA is the AU-rich elements (AREs). AREs are lengths of mRNA siRNAs are processed in the nucleus by a multiprotein complex called
consisting mostly of adenine and uracil nucleotides. These sequences the RNA-induced silencing complex (RISC), which contains the
destabilize those transcripts attached to them through the action of RNase III enzyme Dicer, DGCR8, and Argonaute. The specificity of
riboendonucleases that stimulate poly(A) tail removal. Loss of the miRNA and siRNA interactions with their target mRNAs mediates