Publications by Author: Frank J Slack

Circular RNAs (circRNA) are a recently described class of RNA molecules that have attracted substantial attention as new components of disease mechanisms and as potential biomarkers in multiple diseases, including cancer. CircRNAs are often highly conserved and exhibit developmental stage- and disease-specific expression. Several studies have reported circRNA expression patterns that are associated with specific cancer types and with patient prognosis. Here, we overview the active registered clinical trials that investigate the value of circRNAs as cancer biomarkers and discuss the potential of circRNAs in clinical cancer care. Taken together, circRNAs are actively being investigated as diagnostic, predictive, and prognostic biomarkers, and their potential to serve as therapeutic intervention points motivates ongoing translational and clinical research.

The accurate inheritance of genetic material is a basic necessity in all domains of life and an unexpectedly large number of RNA processing factors are required for mitotic progression and genome stability. NRDE2 (nuclear RNAi defective-2) is an evolutionarily conserved protein originally discovered for its role in nuclear RNA interference (RNAi) and heritable gene silencing in Caenorhabditis elegans (C. elegans). The function of the human NRDE2 gene remains poorly understood. Here we show that human NRDE2 is an essential protein required for suppressing intron retention in a subset of pre-mRNAs containing short, GC-rich introns with relatively weak 5' and 3' splice sites. NRDE2 preferentially interacts with components of the U5 small nuclear ribonucleoprotein (snRNP), the exon junction complex, and the RNA exosome. Interestingly, NRDE2-depleted cells exhibit greatly increased levels of genomic instability and DNA damage, as well as defects in centrosome maturation and mitotic progression. We identify the essential centriolar satellite protein, CEP131, as a direct NRDE2-regulated target. NRDE2 specifically binds to and promotes the efficient splicing of CEP131 pre-mRNA, and depleting NRDE2 dramatically reduces CEP131 protein expression, contributing to impaired recruitment of critical centrosomal proteins (e.g., γ-tubulin and Aurora Kinase A) to the spindle poles during mitosis. Our work establishes a conserved role for human NRDE2 in RNA splicing, characterizes the severe genomic instability phenotypes observed upon loss of NRDE2, and highlights the direct regulation of CEP131 splicing as one of multiple mechanisms through which such phenotypes might be explained.

As the founding member of the microRNA (miRNA) gene family, insights into lin-4 regulation and function have laid a conceptual foundation for countless miRNA-related studies that followed. We previously showed that a transcriptional lin-4 reporter in C. elegans was positively regulated by a lin-4-complementary element (LCE), and by lin-4 itself. In this study, we sought to (1) identify additional factors required for lin-4 reporter expression, and (2) validate the endogenous relevance of a potential positive autoregulatory mechanism of lin-4 expression. We report that all four core nuclear RNAi factors (nrde-1, nrde-2, nrde-3 and nrde-4), positively regulate lin-4 reporter expression. In contrast, endogenous lin-4 levels were largely unaffected in nrde-2;nrde-3 mutants. Further, an endogenous LCE deletion generated by CRISPR-Cas9 revealed that the LCE was also not necessary for the activity of the endogenous lin-4 promoter. Finally, mutations in mature lin-4 did not reduce primary lin-4 transcript levels. Taken together, these data indicate that under growth conditions that reveal effects at the transgenic locus, a direct, positive autoregulatory mechanism of lin-4 expression does not occur in the context of the endogenous lin-4 locus.

The regulation of gene expression by non-coding RNAs (ncRNAs) has become a new paradigm in biology. RNA-mediated gene silencing pathways have been studied extensively, revealing diverse epigenetic and posttranscriptional mechanisms. In contrast, the roles of ncRNAs in activating gene expression remains poorly understood. In this review, we summarize the current knowledge of gene activation by small RNAs, long non-coding RNAs, and enhancer-derived RNAs, with an emphasis on epigenetic mechanisms.

Since their discovery not long ago, microRNAs (miRNAs) have been extensively studied in hundreds of laboratories around the world. Initially thought of as merely cytoplasmic repressors of mRNA expression, it has since become more apparent that they also play regulatory roles in the nucleus. A recent study published in Nature introduces novel concepts in both miRNA regulation and function by showing that the let-7 miRNA regulates its own expression.

MicroRNAs play important roles in animal development, cell differentiation, and metabolism and have been implicated in human cancer. The let-7 microRNA controls the timing of cell cycle exit and terminal differentiation in Caenorhabditis elegans and is poorly expressed or deleted in human lung tumors. Here, we show that let-7 is highly expressed in normal lung tissue, and that inhibiting let-7 function leads to increased cell division in A549 lung cancer cells. Overexpression of let-7 in cancer cell lines alters cell cycle progression and reduces cell division, providing evidence that let-7 functions as a tumor suppressor in lung cells. let-7 was previously shown to regulate the expression of the RAS lung cancer oncogenes, and our work now shows that multiple genes involved in cell cycle and cell division functions are also directly or indirectly repressed by let-7. This work reveals the let-7 microRNA to be a master regulator of cell proliferation pathways.

MicroRNAs (miRNAs) are regulatory RNAs found in multicellular eukaryotes, including humans, where they are implicated in cancer. The let-7 miRNA times seam cell terminal differentiation in C. elegans. Here we show that the let-7 family negatively regulates let-60/RAS. Loss of let-60/RAS suppresses let-7, and the let-60/RAS 3'UTR contains multiple let-7 complementary sites (LCSs), restricting reporter gene expression in a let-7-dependent manner. mir-84, a let-7 family member, is largely absent in vulval precursor cell P6.p at the time that let-60/RAS specifies the 1 degrees vulval fate in that cell, and mir-84 overexpression suppresses the multivulva phenotype of activating let-60/RAS mutations. The 3'UTRs of the human RAS genes contain multiple LCSs, allowing let-7 to regulate RAS expression. let-7 expression is lower in lung tumors than in normal lung tissue, while RAS protein is significantly higher in lung tumors, providing a possible mechanism for let-7 in cancer.

MicroRNAs (miRNAs) are a large family of small regulatory RNAs that are poorly understood. The let-7 miRNA regulates the timing of the developmental switch from larval to adult cell fates during Caenorhabditis elegans development. Expression of let-7 RNA is temporally regulated, with robust expression in the fourth larval and adult stages. Here, we show that, like let-7 RNA, a transcriptional fusion of the let-7 promoter to gfp is temporally regulated, indicating that let-7 is transcriptionally controlled. Temporal upregulation of let-7 transcription requires an enhancer element, the temporal regulatory element (TRE), situated about 1200 base pairs upstream of the start of the mature let-7 RNA. The TRE is both necessary and sufficient for this temporal upregulation. A TRE binding factor (TREB) is able to bind to the TRE, and a 22-base pair inverted repeat within the TRE is necessary and sufficient for this binding. We also find that the nuclear hormone receptor DAF-12 and the RNA binding protein LIN-28 are both required for the correct timing of let-7 RNA and let-7::gfp expression. We speculate that these heterochronic genes regulate let-7 expression through its TRE.

Expression levels of many microRNAs (miRNAs) change during aging, notably declining globally in a number of organisms and tissues across taxa. However, little is known about the mechanisms or the biological relevance for this change. We investigated the network of genes that controls miRNA transcription and processing during C. elegans aging. We found that miRNA biogenesis genes are highly networked with transcription factors and aging-associated miRNAs. In particular, miR-71, known to influence life span and itself up-regulated during aging, represses alg-1/Argonaute expression post-transcriptionally during aging. Increased ALG-1 abundance in mir-71 loss-of-function mutants led to globally increased miRNA expression. Interestingly, these mutants demonstrated widespread mRNA expression dysregulation and diminished levels of variability both in gene expression and in overall life span. Thus, the progressive molecular decline often thought to be the result of accumulated damage over an organism's life may be partially explained by a miRNA-directed mechanism of age-associated decline.

Comment on: Cirera-Salinas D, et al. Cell Cycle 2012; 11:922–33