Regulation and function of microRNAs and other noncoding RNAs in normal physiology and disease

The Mendell laboratory is interested in mechanisms of post-transcriptional regulation of gene expression and how these pathways influence normal physiology and disease. In particular, we have focused on the regulation and functions of noncoding RNAs with an emphasis on microRNAs (miRNAs). miRNAs are an abundant and diverse family of ~20- to 23-nucleotide RNAs that recognize sites of complementarity in target mRNAs, resulting in decay and reduced translation of target transcripts. Approximately 500-1000 miRNAs are encoded in mammalian genomes. Our work on the miRNA pathway focuses mainly on three broad questions: What functions do miRNAs perform in normal physiologic states? How does aberrant miRNA activity contribute to diseases such as cancer? How is miRNA abundance regulated in normal physiology and disease?

Functional Integration of miRNAs into Oncogenic and Tumor-Suppressor Pathways

From numerous miRNA-profiling studies, it is apparent that miRNA expression is frequently dysregulated in diverse human malignancies. In interpreting these data, however, it is important to consider that miRNA expression is tightly regulated during development, across tissues, and during cellular differentiation. Therefore, abnormal miRNA expression patterns in cancer cells are likely to be in part a consequence of the loss of normal cellular identity that accompanies malignant transformation. Thus, a major challenge emerges from these studies: how can we identify the key miRNAs whose altered function directly contributes to neoplastic transformation and tumorigenesis? To address this question, we reasoned that the miRNAs that are directly controlled by critical oncogenic and tumor-suppressor pathways might be highly enriched for the miRNAs that functionally contribute to oncogenesis. Moreover, these same pathways that become aberrantly hyper- or hypoactive in cancer cells are often critical for normal physiology and development. By characterizing miRNAs that have been functionally integrated into these pathways, we will likely gain insight into fundamental aspects of normal miRNA function as well as their activities in disease states. This idea has formed the basis for a number of our studies.

We are currently investigating the roles of miRNAs in several cancer-relevant pathways. Our work in this area is well illustrated by our studies on the c-Myc oncogenic transcription factor (Myc). Through the direct activation or repression of a large network of target genes, Myc potently drives cellular proliferation and tumorigenesis. We hypothesized that miRNAs are likely to be important downstream targets of this oncoprotein, given their known roles in the regulation of cellular proliferation and apoptosis. We used novel microarrays developed in our laboratory and multiple in vitro and in vivo model systems with regulatable Myc activity to test this hypothesis. These studies revealed that Myc broadly reprograms miRNA expression to favor cellular proliferation and survival. For example, a group of six cotranscribed miRNAs (the miR-17-92 cluster) are highly induced by Myc. A large body of evidence generated by our laboratory and others has documented that the miR-17-92 cluster has potent oncogenic activity, and therefore its activation by Myc contributes to tumorigenesis. These findings not only highlighted the significance of miRNAs in Myc-mediated tumorigenesis but also established the principle that miRNAs are likely to be important components of other well-known signaling pathways.

In addition to up-regulation of the oncogenic miR-17-92 cluster, we have observed that, surprisingly, the predominant consequence of Myc activation is widespread reduction of expression of numerous miRNAs. We demonstrated that enforced expression of several individual Myc-repressed miRNAs completely suppressed the ability of cells to form tumors in a model of Myc-induced B cell lymphoma. These findings indicate that Myc-mediated repression of miRNAs provides a powerful selective advantage to cancer cells.

The functional integration of miRNAs into signaling cascades that function in normal physiology and cancer is not unique to the Myc pathway. We have shown that miRNAs also provide critical functions downstream of the Kras oncogenic pathway and the p53 tumor suppressor pathway and we are continuing to study how miRNAs contribute to other signaling networks. Moreover, a major effort is underway in the laboratory to generate and characterize novel loss- and gain-of-function models in mice and zebrafish to elucidate the roles of these miRNAs in both normal physiology and tumor development in vivo.

Therapeutic Manipulation of the miRNA Pathway

In light of our demonstration that expression of individual miRNAs can strongly block tumorigenesis, we hypothesized that miRNA replacement strategies designed to restore physiologic expression levels of these molecules to cancer cells might represent an effective therapeutic paradigm. We are using multiple mouse cancer models to explore this possibility. Recently, we provided strong proof-of-concept support for a miRNA-based therapeutic strategy for liver cancer in which we used adeno-associated virus (AAV) to deliver a miRNA to a mouse model of hepatocellular carcinoma. Remarkably, delivery of a single miRNA dramatically inhibited disease progression and induced apoptosis specifically in tumors without any associated toxicity. With these promising results, we are using AAV and other delivery systems to investigate the efficacy of miRNA-based therapeutics in a variety of disease models.

Mechanisms of miRNA Regulation in Normal Physiology and Disease

We have become increasingly interested in the molecular mechanisms through which miRNA abundance is controlled. Emerging data indicate that there is surprising complexity in the post-transcriptional regulation of miRNA biogenesis. We have demonstrated that Myc deploys both transcriptional and post-transcriptional mechanisms to reprogram miRNA expression, and we are investigating how Myc controls miRNA biogenesis at these distinct levels. More broadly, accumulating data demonstrate that widespread post-transcriptional control of miRNA biogenesis is common in many physiologic and pathophysiologic settings. Given that re-expression of even a single miRNA in tumor cells can have dramatic therapeutic benefit, reversal of this broad repression of miRNA expression in cancer cells might be beneficial for cancer therapy. We now using in vitro and in vivo model systems to biochemically characterize the miRNA biogenesis machinery under high- and low-efficiency states. Our long-term goal is to understand the mechanisms and consequences of dynamic regulation of miRNA biogenesis and to develop tools to influence the efficiency of the pathway for therapeutic benefit.