Conservation and Targets of miR-71: A Systematic Review and Meta-Analysis.

MicroRNAs (miRNAs) perform a pivotal role in the regulation of gene expression across the animal kingdom. As negative regulators of gene expression, miRNAs have been shown to function in the genetic pathways that control many biological processes and have been implicated in roles in human disease. First identified as an aging-associated gene in C. elegans, miR-71, a miRNA, has a demonstrated capability of regulating processes in numerous different invertebrates, including platyhelminths, mollusks, and insects. In these organisms, miR-71 has been shown to affect a diverse range of pathways, including aging, development, and immune response. However, the exact mechanisms by which miR-71 regulates these pathways are not completely understood. In this paper, we review the identified functions of miR-71 across multiple organisms, including identified gene targets, pathways, and the conditions which affect regulatory action. Additionally, the degree of conservation of miR-71 in the evaluated organisms and the conservation of their predicted binding sites in target 3' UTRs was measured. These studies may provide an insight on the patterns, interactions, and conditions in which miR-71 is able to exert genotypic and phenotypic influence.

Parkinson's disease and microRNAs - Lessons from model organisms and human studies.

Parkinson's disease (PD) is a progressive, age-associated neurodegenerative disorder that affects an estimated 10 million people worldwide. PD is characterized by proteinaceous, cytoplasmic inclusions containing α-synuclein, called Lewy Bodies, which form in dopaminergic neurons in an age-dependent manner, and are associated with the emergence of characteristic PD symptoms such as resting tremor, rigidity, slow movements and postural instability. Although considerable progress has been made in recent years in identifying genetic and environmental factors that are associated with PD, early diagnosis and therapeutic options remain severely lacking. Recently, microRNAs (miRNAs) have emerged as novel therapeutic targets in various diseases, such as cancer and neurodegenerative diseases. MiRNAs have been shown to play roles in various aging and neurodegenerative disease models across phyla. More recently, studies have identified specific roles for miRNAs and their targets in the pathogenesis and progression of PD in several model organisms. Here, we discuss the evolving field of miRNAs, their association with PD, and the outlook for the future.

A microRNA feedback loop regulates global microRNA abundance during aging.

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.

An investigative graduate laboratory course for teaching modern DNA techniques.

This graduate-level DNA methods laboratory course is designed to model a discovery-based research project and engages students in both traditional DNA analysis methods and modern recombinant DNA cloning techniques. In the first part of the course, students clone the Drosophila ortholog of a human disease gene of their choosing using Gateway® cloning. In the second part of the course, students examine the expression of their gene of interest in human cell lines by reverse transcription PCR and learn how to analyze data from quantitative reverse transcription PCR (qRT-PCR) experiments. The adaptability of the Gateway® cloning system is ideally suited for students to design and create different types of expression constructs to achieve a particular experimental goal (e.g., protein purification, expression in cell culture, and/or subcellular localization), and the genes chosen can be aligned to the research interests of the instructor and/or ongoing research in a department. Student evaluations indicate that the course fostered a genuine excitement for research and in depth knowledge of both the techniques performed and the theory behind them. Our long-term goal is to incorporate this DNA methods laboratory as the foundation for an integrated laboratory sequence for the Master of Science degree program in Molecular and Cellular Biology at Quinnipiac University, where students use the reagents and concepts they developed in this course in subsequent laboratory courses, including a protein methods and cell culture laboratory. © 2017 by The International Union of Biochemistry and Molecular Biology, 45(4):351-359, 2017.

Alzheimer's disease: presence and role of microRNAs.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder that accounts for the most cases of dementia. AD affects more than 25 million people globally and is predicted to affect nearly one in 85 people worldwide by 2050. AD is characterized by the accumulation of dense plaques of ß-amyloid peptide (Aß) and neurofibrillary tangles of hyperphosphorylated tau that cause impairment in memory, cognition, and daily activities. Although early-onset AD has been linked to several mutations, reliable genetic markers for late-onset AD are lacking. Further, the diagnosis of AD biomarkers has its limitations and cannot detect early-stage AD. The identification of accurate, early, and non-invasive biomarkers for AD is, therefore, an unmet challenge. Recently, microRNAs (miRNAs) have emerged as a novel class of gene regulatory elements with conserved roles in development and disease. Recent discoveries have uncovered roles of miRNAs in several model organisms during aging and have identified potential miRNAs biomarkers of AD. Here we will discuss this emerging field of miRNAs associated with AD and prospects for the future.

Discovery of Novel microRNAs in Aging Caenorhabditis elegans.

The rapid development of deep sequencing technologies over the last few years and concomitant increases in sequencing depth and cost efficiencies have opened the door to a ever-widening range of applications in biology-from whole-genome sequencing, to ChIP-seq analysis, epigenomic and RNA transcriptome surveys. Here we describe the application of deep sequencing to the discovery of novel microRNAs and characterization of their differential expression during adulthood in Caenorhabditis elegans.

MicroRNAs mediate dietary-restriction-induced longevity through PHA-4/FOXA and SKN-1/Nrf transcription factors.

BACKGROUND: Dietary restriction (DR) has been shown to prolong longevity across diverse taxa, yet the mechanistic relationship between DR and longevity remains unclear. MicroRNAs (miRNAs) control aging-related functions such as metabolism and lifespan through regulation of genes in insulin signaling, mitochondrial respiration, and protein homeostasis. RESULTS: We have conducted a network analysis of aging-associated miRNAs connected to transcription factors PHA-4/FOXA and SKN-1/Nrf, which are both necessary for DR-induced lifespan extension in Caenorhabditis elegans. Our network analysis has revealed extensive regulatory interactions between PHA-4, SKN-1, and miRNAs and points to two aging-associated miRNAs, miR-71 and miR-228, as key nodes of this network. We show that miR-71 and miR-228 are critical for the response to DR in C. elegans. DR induces the expression of miR-71 and miR-228, and the regulation of these miRNAs depends on PHA-4 and SKN-1. In turn, we show that PHA-4 and SKN-1 are negatively regulated by miR-228, whereas miR-71 represses PHA-4. CONCLUSIONS: Based on our findings, we have discovered new links in an important pathway connecting DR to aging. By interacting with PHA-4 and SKN-1, miRNAs transduce the effect of dietary-restriction-mediated lifespan extension in C. elegans. Given the conservation of miRNAs, PHA-4, and SKN-1 across phylogeny, these interactions are likely to be conserved in more-complex species.

Novel microRNAs differentially expressed during aging in the mouse brain.

MicroRNAs (miRNAs) are endogenous small RNA molecules that regulate gene expression post-transcriptionally. Work in Caenorhabditis elegans has shown that specific miRNAs function in lifespan regulation and in a variety of age-associated pathways, but the roles of miRNAs in the aging of vertebrates are not well understood. We examined the expression of small RNAs in whole brains of young and old mice by deep sequencing and report here on the expression of 558 known miRNAs and identification of 41 novel miRNAs. Of these miRNAs, 75 known and 18 novel miRNAs exhibit greater than 2.0-fold expression changes. The majority of expressed miRNAs in our study decline in relative abundance in the aged brain, in agreement with trends observed in other miRNA studies in aging tissues and organisms. Target prediction analysis suggests that many of our novel aging-associated miRNAs target genes in the insulin signaling pathway, a central node of aging-associated genetic networks. These novel miRNAs may thereby regulate aging-related functions in the brain. Since many mouse miRNAs are conserved in humans, the aging-affected brain miRNAs we report here may represent novel regulatory genes that also function during aging in the human brain.

MicroRNAs both promote and antagonize longevity in C. elegans.

BACKGROUND: aging is under genetic control in C. elegans, but the mechanisms of life-span regulation are not completely known. MicroRNAs (miRNAs) regulate various aspects of development and metabolism, and one miRNA has been previously implicated in life span. RESULTS: here we show that multiple miRNAs change expression in C. elegans aging, including novel miRNAs, and that mutations in several of the most upregulated miRNAs lead to life-span defects. Some act to promote normal life span and stress resistance, whereas others inhibit these phenomena. We find that these miRNAs genetically interact with genes in the DNA damage checkpoint response pathway and in the insulin signaling pathway. CONCLUSIONS: our findings reveal that miRNAs both positively and negatively influence life span. Because several miRNAs upregulated during aging regulate genes in conserved pathways of aging and thereby influence life span in C. elegans, we propose that miRNAs may play important roles in stress response and aging of more complex organisms.

Dynamic expression of small non-coding RNAs, including novel microRNAs and piRNAs/21U-RNAs, during Caenorhabditis elegans development.

BACKGROUND: Small non-coding RNAs, including microRNAs (miRNAs), serve an important role in controlling gene expression during development and disease. However, little detailed information exists concerning the relative expression patterns of small RNAs during development of animals such as Caenorhabditis elegans. RESULTS: We performed a deep analysis of small RNA expression in C. elegans using recent advances in sequencing technology, and found that a significant number of known miRNAs showed major changes in expression during development and between males and hermaphrodites. Additionally, we identified 66 novel miRNA candidates, about 35% of which showed transcripts from their 'star sequence', suggesting that they are bona fide miRNAs. Also, hundreds of novel Piwi-interacting RNAs (piRNAs)/21U-RNAs with dynamic expression during development, together with many longer transcripts encompassing 21U-RNA sequences, were detected in our libraries. CONCLUSIONS: Our analysis reveals extensive regulation of non-coding small RNAs during development of hermaphrodites and between different genders of C. elegans, and suggests that these RNAs, including novel miRNA candidates, are involved in developmental processes. These findings should lead to a better understanding of the biological roles of small RNAs in C. elegans development.

Three essential and conserved regions of the group II intron are proximal to the 5'-splice site.

Despite the central role of group II introns in eukaryotic gene expression and their importance as biophysical and evolutionary model systems, group II intron tertiary structure is not well understood. In order to characterize the architectural organization of intron ai5gamma, we incorporated the photoreactive nucleotides s(4)U and s(6)dG at specific locations within the intron core and monitored the formation of cross-links in folded complexes. The resulting data reveal the locations for many of the most conserved, catalytically important regions of the intron (i.e., the J2/3 linker region, the IC1(i-ii) bulge in domain 1, the bulge of D5, and the 5'-splice site), showing that all of these elements are closely colocalized. In addition, we show by nucleotide analog interference mapping (NAIM) that a specific functional group in J2/3 plays a role in first-step catalysis, which is consistent with its apparent proximity to other first-step components. These results extend our understanding of active-site architecture during the first step of group II intron self-splicing and they provide a structural basis for spliceosomal comparison.

A single active-site region for a group II intron.

Despite the biological importance of self-splicing group II introns, little is known about their structural organization. Synthetic incorporation of site-specific photo-cross-linkers within catalytic domains resulted in functional distance constraints that, when combined with known tertiary interactions, provide a three-dimensional view of the active intron architecture. All functionalities important for both steps of splicing are proximal before the first step, suggestive of a single active-site region for group II intron catalysis.