Liquid droplet organelles (also referred to as liquid-like condensates) are dynamic concentrations of protein and RNA that coalesce spontaneously from the cytoplasm or nucleoplasm via liquid-liquid phase transition. Like oil and water, these liquid droplets and their surrounding cytoplasm are thought to co-exist as separate states of liquid. Current models posit that liquid droplet organelles may exist to concentrate reactants together in space and time in ways that facilitate biochemical reactions such as the complexes processes that underlie gene regulation. Eukaryotic cells possess many non-membrane-enclosed liquid droplet organelles including nucleoli, processing bodies, cajal bodies and germ granules. Germ granules are found in the germ cells of all/most animals where they are thought to contribute to germ cell totipotency. Indeed, orthologous proteins (e.g. VASA) are found in germ granules of many different species of animals, hinting that some aspects of germ granule function may be conserved in all animals. Most epigenetic information is erased at or near the start of each new generation in animals to ensure totipotency of the germline. In some cases, epigenetic information escapes reprogramming and is passed from parent to offspring. The inheritance of epigenetic information for more than one generation is termed trans- generational epigenetic inheritance (TEI). Small non-coding RNAs (e.g. siRNAs, miRNAs, piRNAs, and tRNA fragments) have emerged as important mediators of TEI (cumulatively, RNA-directed TEI). dsRNA-mediated gene silencing (RNAi) in C. elegans is a robust and dramatic example of RNA-directed TEI. My lab is using RNAi inheritance in C. elegans as a model system to explore how epigenetic signals are passed across generations in animals. We recently conducted a genetic screen for RNAi inheritance factors that identified two conserved RNA binding proteins (RBPs). These RBPs contribute directly to inheritance by helping to maintain the expression of silencing RNAs over generations during RNAi inheritance. Interestingly, both RBPs localize to a new C. elegans germ granule whose biogenesis is developmentally regulated and whose positioning relative to other germline condensates is highly ordered. Finally, we find that the two RNAi inheritance factors we have identified are required for maintaining germline immortality. Together, our results have led us to propose that the temporal and spatial ordering of liquid droplet organelles may help germ cells organize and coordinate the complex RNA processing pathways underlying gene regulatory systems, such as RNA-directed TEI and germline immortality. This proposal outlines our efforts to further understand how liquid-like condensates form during development, how they assemble into complex structures with other germline condensates, and, most importantly, why they do so.

Public Health Relevance

Germ granules are liquid droplet organelles (liquid-like condensates) thought to play important roles in germline totipotency in all animals. We have discovered a new type of C. elegans germ granule that we call the Z granule that assembles into ordered multi-condensate structure with two other organelles in germ cells, and which seems to contribute to RNA-directed transgenerational epigenetic inheritance and germline immortality. Currently, we are investigating how Z granules form, what they do, and how they ?know? to assemble into multi-condensates structures with two other liquid droplet organelles during germline development.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
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Development - 1 Study Section (DEV1)
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Salazar, Desiree Lynn
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Harvard Medical School
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