Rachel Kaletsky

The role of insulin/IGF-like signaling in C. elegans longevity and aging

Aging is characterized by general physiological decline over time. A hallmark of human senescence is the onset of various age-related afflictions including neurodegeneration, cardiovascular disease and cancer. Although environmental and stochastic factors undoubtedly contribute to the increased incidence of disease with age, recent studies suggest that intrinsic genetic determinants govern both life span and overall health. Current aging research aims at achieving the ‘longevity dividend’, in which life span extension in humans is accomplished with a concomitant increase in the quality of life (Olshansky et al., 2007). Significant progress has been made using model organisms, especially the nematode worm Caenorhabditis elegans, to delineate the genetic and biochemical pathways involved in aging to identify strategies for therapeutic intervention in humans. In this review, we discuss how C. elegans has contributed to our understanding of insulin signaling and aging.

The C. elegans adult neuronal IIS/FOXO transcriptome reveals adult phenotype regulators

Insulin/insulin-like growth factor signalling (IIS) is a critical regulator of an organism’s most important biological decisions from growth, development, and metabolism to reproduction and longevity. It primarily does so through the activity of the DAF-16 transcription factor (forkhead box O (FOXO) homologue), whose global targets were identified in Caenorhabditis elegans using whole-worm transcriptional analyses more than a decade ago. IIS and FOXO also regulate important neuronal and adult behavioural phenotypes, such as the maintenance of memory and axon regeneration with age, in both mammals and C. elegans, but the neuron-specific IIS/FOXO targets that regulate these functions are still unknown. By isolating adult C. elegans neurons for transcriptional profiling, we identified both the wild-type and IIS/FOXO mutant adult neuronal transcriptomes for the first time. IIS/FOXO neuron-specific targets are distinct from canonical IIS/FOXO-regulated longevity and metabolism targets, and are required for extended memory in IIS daf-2 mutants. The activity of the forkhead transcription factor FKH-9 in neurons is required for the ability of daf-2 mutants to regenerate axons with age, and its activity in non-neuronal tissues is required for the long lifespan of daf-2 mutants. Together, neuron-specific and canonical IIS/FOXO-regulated targets enable the coordinated extension of neuronal activities, metabolism, and longevity under low-insulin signalling conditions.

Genome-wide Functional Analysis of CREB/Long-Term Memory-Dependent Transcription Reveals Distinct Basal and Memory Gene Expression Programs

Induced CREB activity is a hallmark of long-term memory, but the full repertoire of CREB transcriptional targets required specifically for memory is not known in any system. To obtain a more complete picture of the mechanisms involved in memory, we combined memory training with genome-wide transcriptional analysis of C. elegans CREB mutants. This approach identified 757 significant CREB/memory-induced targets and confirmed the involvement of known memory genes from other organisms, but also suggested new mechanisms and novel components that may be conserved through mammals. CREB mediates distinct basal and memory transcriptional programs at least partially through spatial restriction of CREB activity: basal targets are regulated primarily in nonneuronal tissues, while memory targets are enriched for neuronal expression, emanating from CREB activity in AIM neurons. This suite of novel memory-associated genes will provide a platform for the discovery of orthologous mammalian long-term memory components.

Cell-Specific Transcriptional Profiling of Ciliated Sensory Neurons Reveals Regulators of Behavior and Extracellular Vesicle Biogenesis

Cilia and extracellular vesicles (EVs) are signaling organelles [1]. Cilia act as cellular sensory antennae, with defects resulting in human ciliopathies. Cilia both release and bind to EVs [1]. EVs are sub-micron-sized particles released by cells and function in both short- and long-range intercellular communication. In C. elegans and mammals, the autosomal dominant polycystic kidney disease (ADPKD) gene products polycystin-1 and polycystin-2 localize to both cilia and EVs, act in the same genetic pathway, and function in a sensory capacity, suggesting ancient conservation [2]. A fundamental understanding of EV biology and the relationship between the polycystins, cilia, and EVs is lacking. To define properties of a ciliated EV-releasing cell, we performed RNA-seq on 27 GFP-labeled EV-releasing neurons (EVNs) isolated from adult C. elegans. We identified 335 significantly overrepresented genes, of which 61 were validated by GFP reporters. The EVN transcriptional profile uncovered new pathways controlling EV biogenesis and polycystin signaling and also identified EV cargo, which included an antimicrobial peptide and ASIC channel. Tumor-necrosis-associated factor (TRAF) homologs trf-1 and trf-2 and the p38 mitogen-activated protein kinase (MAPK) pmk-1 acted in polycystin-signaling pathways controlling male mating behaviors. pmk-1 was also required for EV biogenesis, independent of the innate immunity MAPK signaling cascade. This first high-resolution transcriptome profile of a subtype of ciliated sensory neurons isolated from adult animals reveals the functional components of an EVN.

RNA surveillance via nonsense-mediated mRNA decay is crucial for longevity in daf-2 /insulin/IGF-1 mutant C. elegans

Long-lived organisms often feature more stringent protein and DNA quality control. However, whether RNA quality control mechanisms, such as nonsense-mediated mRNA decay (NMD), which degrades both abnormal as well as some normal transcripts, have a role in organismal aging remains unexplored. Here we show that NMD mediates longevity in C. elegans strains with mutations in daf-2/insulin/insulin-like growth factor 1 receptor. We find that daf-2 mutants display enhanced NMD activity and reduced levels of potentially aberrant transcripts. NMD components, including smg-2/UPF1, are required to achieve the longevity of several long-lived mutants, including daf-2 mutant worms. NMD in the nervous system of the animals is particularly important for RNA quality control to promote longevity. Furthermore, we find that downregulation of yars-2/tyrosyl-tRNA synthetase, an NMD target transcript, by daf-2 mutations contributes to longevity. We propose that NMD-mediated RNA surveillance is a crucial quality control process that contributes to longevity conferred by daf-2 mutations.

Transcriptome analysis of adult Caenorhabditis elegans cells reveals tissue-specific gene and isoform expression

The biology and behavior of adults differ substantially from those of developing animals, and cell-specific information is critical for deciphering the biology of multicellular animals. Thus, adult tissue-specific transcriptomic data are critical for understanding molecular mechanisms that control their phenotypes. We used adult cell-specific isolation to identify the transcriptomes of C. elegans’ four major tissues (or “tissue-ome”), identifying ubiquitously expressed and tissue-specific “enriched” genes. These data newly reveal the hypodermis’ metabolic character, suggest potential worm-human tissue orthologies, and identify tissue-specific changes in the Insulin/IGF-1 signaling pathway. Tissue-specific alternative splicing analysis identified a large set of collagen isoforms. Finally, we developed a machine learning-based prediction tool for 76 sub-tissue cell types, which we used to predict cellular expression differences in IIS/FOXO signaling, stage-specific TGF-β activity, and basal vs. memory-induced CREB transcription. Together, these data provide a rich resource for understanding the biology governing multicellular adult animals.

Transcriptome Analysis of Adult C. elegans Cells Reveals Tissue-specific Gene and Isoform Expression

The biology and behavior of adults differ substantially from those of developing animals, and cell-specific information is critical for deciphering the biology of multicellular animals. Thus, adult tissue-specific transcriptomic data are critical for understanding molecular mechanisms that control their phenotypes. We used adult cell-specific isolation to identify the transcriptomes of C. elegans’ four major tissues (or “tissue-ome”), identifying ubiquitously expressed and tissue-specific “super-enriched” genes. These data newly reveal the hypodermis’ metabolic character, suggest potential worm-human tissue orthologies, and identify tissue-specific changes in the Insulin/IGF-1 signaling pathway. Tissue-specific alternative splicing analysis identified a large set of collagen isoforms and a neuron-specific CREB isoform. Finally, we developed a machine learning-based prediction tool for 70 sub-tissue cell types, which we used to predict cellular expression differences in IIS/FOXO signaling, stage-specific TGF-b activity, and basal vs. memory-induced CREB transcription. Together, these data provide a rich resource for understanding the biology governing multicellular adult animals