Cancer Beyond the Coding Frontier: miRNAs, piRNAs, and the Epigenome in Human Oncogenesis

The advent of small noncoding RNAs as important modulators of cancer susceptibility and progression has gained widespread recognition in recent years. This understanding was precipitated by the 1993 discovery of a 22--nucleotide RNA in C. elegant capable of translational repression but was itself never translated into protein. Since the unveiling of this first microRNA (miRNA), over 1,000 additional members of the miRNA superfamily have been uncovered in humans, together accounting for the regulation of an estimated 60% of the human genome. The near functional ubiquity of miRNAs has in turn provided the catalyst for a considerable number of molecular and epidemiological undertakings to elucidate the role of miRNAs in human cancers, leading to an enhanced understanding of miRNA function in human carcinogenesis. Indeed, the role of miRNAs in the oncogenic process has, in a relatively short period of time, evolved from being novel to canon, w ith implications in a wide spectrum of cancer types.;In contrast to the voluminous body of experimental and observational evidence that underpin the miRNA-cancer connection, knowledge of PIWI-interacting RNAs (piRNAs) in the carcinogenic process remains in its infancy. Although purportedly existing in over 20,000 unique species in the human genome, piRNAs enjoy few of the epistemiological advantages that allow miRNAs to be so readily studied in cancer. Elements of the biogenic process of piRNAs remain unelucidated, including the mechanism of transcription and the structure of the primary transcript, and we are only now starting to gain an appreciation of the potentially vast array of functions associated with the piRNA regulatory domain. Although it was believed that the primary role of piRNAs is to silence active transposable elements in the germ line and thereby guard the genome against transposon-induced insertional mutations, mounting evidence now point towards an active role for piRNAs in somatic gene regulation through sequencespecific histone modification and DNA methylation. Despite the broad implications this development holds for the study of piRNAs in cancer, current research of the PIWI/piRNA axis in human cancers is largely restricted to expression profiling of PIWI proteins and a narrow selection of individual piRNAs.;In this thesis, we use functional genetics to explicate the molecular behavior that underlies observed genetic associations between two small non coding RNAs and their respective cancer types. Chapter 1 details the results of our efforts to elucidate the functional link between a single nucleotide polymorphism (SNP) within the miR-61S precursor sequenceand its association with follicular non-Hodgkins lymphoma. Our findings reveal that the presence of the variant allele may negatively impact the production of mature effector miR--618 by interfering with the post-transcriptional miRNA biogenic process, in turn leading to the dysrcgulation of lymphoma-relevant miR-618 targets. In Chapter 2, we demonstrate the ability of certain piRNAs to affect - potentially by binding to DNA in a sequence-specific manner the DNA methylation of protein-coding genes in somatic cancer cell lines. This finding helps to establish a possible mechanism by which piRNAs may be able to alter one s susceptibility to cancer development, namely through piRNA-mediated DNA methylation of oncogenes and tumor suppressors. Lastly, in Chapter 3, we attem pt to reconcile an association between a SNP in piR-021285 and breast cancer with gene-specific DNA methylation differences found between wild type and variant piR-021285 mimic-transfected breast cancer cells. Our findings reveal that predominance of the variant allele may be associated with attenuated methylation and enhanced expression of the cell motility-related ARHGAPI1A gene, which may confer a hyper-invasive phenotype on breast tumor cells. Together, these findings help characterize a little studied miRNA in the lymphomagenic process by capitalizing upon a clearly defined biogenic process and mechanism of action, as well as reveal possible mechanisms through which a piRNA may able to alter breast cancer susceptibility and disease progression.