Essay: Small ribonucleic acid (RNA)

Small ribonucleic acid (RNA) can act as specific regulators of gene expression. Over the years many new small functional RNAs have been found. RNAs are usually thought of as messenger RNA, which serve as templates for the translation of genes into proteins. Encoding of the DNA sequence is often referred to as an RNA gene. Hundreds of genes in our genome encode small functional RNA molecules collectively called microRNAs (miRNAs).
MicroRNAs are a class of non-coding regulatory RNAs, they are approximately 22 nucleotides long, involved in differentiation, development, and metabolism (Castoldi, 2006). Acting at the post-transcriptional level, miRNAs can fine-tune the expression of about 30 percent of mammalian protein-encoding genes.
MicroRNAs (miRNAs) were first discovered by Victor Ambros along with colleagues Roaslind Lee and Rhonda Feinbaum in 1993. Ambros and colleagues conducted a genetic screening of the roundworm Caenorhabditis elegans and identified genes involved in developmental timing (Sassen, 2008). These discoveries along with the discovery of miRNAs in Drosophila were shown to control cell proliferation and apoptosis.
MicroRNAs are shown to control a variety of biological processes, such as cell cycle, apoptosis, and various physiological properties. In addition, highly tissue-specific expression and distinct time-based expression patterns during embryogenesis suggest that microRNAs play a key role in the differentiation and maintenance of tissue identity. MiRNAs have also been implicated in a variety of diseases including various cancers, neurological diseases, and heart disease (Sassen, 2008).
All cancers share a number of characteristics, including an increased ability to proliferate, alterations to the cell cycle, and a loss of cellular identity (Sassen, 2008). Research has shown that miRNAs are able to regulate these processes, suggesting their involvement in cancer development.
In C. elegans, lin-4 and let-7 control the timing of development; mutations in the miRNAs result in abnormalities in the cell cycle along with execution of terminal differentiation program. This prevents the cells from reaching their fully differentiated state. When researchers discovered human microRNAs they noticed that many of the miRNAs were located along fragile sites in the genome or in regions that are commonly amplified or deleted in human cancers (Sassen, 2008). Tumor cell lines and malignant tumors were found to have widespread deregulated miRNA expression compared to normal tissues (Sassen, 2008). Many miRNAs are found to be up- or down-regulated in the cancer samples as they relate to their normal tissue counterparts.
Approximately five years ago miRNAs role in cancer was first reported. Studies show that microRNAs have oncogenic properties, the first was identified was miR-155 (Metzler, 2004). Early studies showed miR-155 to be upregulated in Burkitt’s lymphoma. miR-155 is located on chromosome 21in a host of non-coding RNA called the B cell integration (BIC) and is highly expressed in pediatric Burkitt’s lymphoma. Constinean et al. conducted a study that showed transgenic mice with a pre-B-cell targeted over expression of miR-155. The mice were fertile and viable but developed polyclonal lymphoproliferative disorder which was then followed by pre-B-leukemia. These results provided evidence that if microRNAs are deregulated cancer can develop (Sandhu, 2011).
In 2005, several reports provided the first mechanistic view on how miRNAs may contribute to carcinogenesis. MicroRNA, miR-17-92 polycistron, has been reported to play a major role in lymphomagenesis. miR-17-92 polycistron is located in a region commonly amplified in B-cell lymphomas and is upregulated in approximately 65 percent of lymphoma patients (Sandhu, 2011). Gain and loss-of-function studies of miR-17-92 polycistron have provided an important insight into its mechanism of action and its targets. He et al. demonstrated that virus-mediated overexpression of miR-17-92 in lymphocytes of B-cell transgenic mice accelerated tumor development (He et al., 2005). The pathology indicated lower rates of apoptosis compared to tumors with Myc protein overexpression. Myc is a group of vertebrate oncogenes whose product, a DNA binding protein, is thought to promote the growth of tumor cells (Dictionary.com, 2009). A mutated version of Myc is found in many cancers, which causes Myc to be constitutively expressed. This leads to deregulated expression of many genes. Studies show that miR-17-92 have anti-apoptotic effects through various pathways that promote cell growth and as a mediator of angiogenesis in tumors induced by Myc (Sassen, 2008).
Let-7 a family of microRNAs was the first group of miRNAs revealed to regulate the expression of the proto-oncogene, Ras. Let-7 and its family members are highly conserved across species in sequence and function, misregulation of let-7 leads to a less differentiated cellular state and the development of cell-based diseases such as cancer. Ras is a signaling protein that regulates cell growth and differentiation. miRNAs that control the expression of Ras are predicted to contain tumor suppressor activity (Sassen, 2008). Mutations in Ras oncogene are present in between 15-30 percent of human cancers.
Current research on microRNAs discusses ways that miRNAs are being used as targets for drug development as well as their usage in determining cancer pathways.
One article discusses the uses of miRNAs and their role in determining the function of p53. p53 is a tumor suppressor gene that has been identified as a key player, in the development of multiple types of cancers in its inactive state. p53’s most important role is the prevention of tumor development (Jun-Ming Liao, 2014). The study conducted by Jun-Ming et al. discusses the role p53 driven miRNAs play in human cancer and other diseases.
Multiple miRNAs have been identified as new p53 targets over the past decade in an attempt to determine the role of p53 in human diseases such as cancer, this could offer new insights into understanding some of the issues that arise in the gene (Jun-Ming Liao, 2014). p53 plays a major role in the biogenesis of miRNAs, by binding the promoter of the miRNAs, controlling their transcription, or by inhibiting their RNA silencing mechanism. Recent studies suggest that p53-regulated miRNAs regulate not only the p53 pathway, but other human diseases and their respective pathways.
The first p53 target miRNA mR-34 was identified in 2007; it was identified by its role in inducing p53 dependent apoptosis and cell cycle arrest (Jun-Ming Liao, 2014). The study discusses the negative feedback loop between p53 and miRNAs. The loop that normally would regulate cell proliferation and survival can be implemented by p53 target miRNAs either positively or negatively. This is done by altering the p53 antagonists such as MDM2 and MDMX which are major p53 suppressors. The miRNA involved in these pathways are transcriptionally induced to suppress the antagonists, preventing their function and protecting p53 from degradation (Jun-Ming Liao, 2014). This displays miRNAs and their positive role in the p53 feedback regulation. Other miRNAs are responsible for regulating levels of p53 in response to stimuli. For example, miR-29 induced transcriptionally by p53 in response to DNA damage can positively influence p53 levels (Jun-Ming Liao, 2014).
In addition to their roles in the feedback loop of p53, p53 target miRNAs play a major role in mediating cell growth that is p53 dependent. miRNAs that play a role in p53-induced cell cycle arrest include miR-34, miR-107, miR-205, miR-192, and miR-215. miR-34 induces cell cycle arrest by suppressing the expression of multiple cell cycle-associated proteins (Jun-Ming Liao, 2014). The p53 role in the suppression of oncogenes has also presented itself as an area of research due to their role in the under or over expression of each other. The inverse relationship between p53 and c-Myc levels was studied and revealed that miR-145 and miR-34, both p53 targets, were shown to suppress c-Myc expression in response to p53 activation (Jun-Ming Liao, 2014).
Other studies of miRNA show the changes in cancer cells that could be cause by altered expression of miRNAs. A recent study conducted by N. Kohei et al. showed the expression levels of miR-373 in pancreatic cancer cell lines and its effect on the invasiveness of pancreatic cancer (Kohei Nakata, 2014). The study found that the levels of miR-373 expression were low in pancreatic cancer cell line and that miR-373 expression was signi’cantly down-regulated in pancreatic cancer compared with that in healthy pancreas. The study also uncovered that reexpression of the miRNAs represses the invasiveness of pancreatic cancer cells (Kohei Nakata, 2014).
Every miRNA has multiple target sites on different genes, compared to only about two thirds of all mRNAs that have one or more evolutionarily conserved sequences that are predicted to interact with microRNAs. According to Rothschild, the rationale for using miRNAs as therapeutic agents is as follows 1) the cancer phenotype can be changed by targeting miRNA expression and 2) miRNA expression is deregulated in cancer compared to normal tissue (Rothschild, 2014). Therapeutic targeting of microRNAs can be accomplished either by direct inhibition or replacement of miRNAs or by targeting specific genes and therefore regulating the expression of specific miRNAs. microRNA based therapies have several advantages such as the ability to target multiple genes frequently in the context of a network (Rothschild, 2014).
With the discovery of microRNAs being powerful regulators in a wide variety of diseases, it is only a logical that the possibilities of viewing miRNAs as therapeutic entities are being explored. miRNA based therapeutic strategies have been successfully applied in pre-clinical models for several malignancies (Steffy, 2011). Therapeutic application of microRNAs involves two strategies; the first is geared toward a gain-of-function and aims to inhibit oncogenic miRNAs by using miRNA antagonist such as anti-miRs or antagomiRs. These microRNA antagonists are oligonucleotides with sequences complementary to endogenous miRNA. anti-miR oligonucleotides are rationally designed to target stability, target affinity, and tissue uptake. Regulus Therapeutics demonstrated that modulating miRNAs through anti-miR oligonucleotides (AMO) can effectively regulate biological processes and produce therapeutically beneficial results (Steffy, 2011). AMO is a version of steric blocking antisense oligonucleotides that form a duplex with the miRNA guide strand and this binding event is the basis for miRNA inactivation.
The second strategy, miRNA replacement, involves the re-introduction of a tumor suppressor miRNA mimic to restore a loss-of-function. Tumor suppressor miRNAs that display a loss-of-function can be replaced by re-introduction of miRNA into diseased tissues, which in turn reactivates the pathway (Mirna Therapeutics, 2013). Reactivation of miRNA-regulated pathways interferes with the oncogenic properties of cancer cells. This blocks uncontrolled cell proliferation and initiates a cascade of apoptotic events (Rooj, 2012).
Research on miRNAs has developed from discovery to researching roles in disease development to being studied as potential aids in curing disease. miRNAs play a significant role in the process of carcinogenesis and its prevention, aiding the regulatory genes in their fight to maintain the overall health of the cell. miRNAs have been recognized for their role in tumor prevention and protecting essential genes that prevent uncontrolled cell proliferation. p53 employs miRNAs to maintain and protect its function. They have also been shown to play a role in decreasing the invasiveness of some cancers during their development. Given that many miRNAs are deregulated in cancers but have not yet been further studied, it is expected that more miRNAs will emerge as players in the progression of cancer. miRNAs could also be also discussed as a mechanism for cancer therapy. Future trials will provide insights into the safety and efficacy of developed miRNA-based anti-cancer therapies.