Self-organizing principles in chromatin-templated processes [electronic resource]

The English embryologist Conrad Waddington coined the term "epigenetics" to describe heritable changes in developmental phenotypes that were not encoded in DNA sequence. He was originally fascinated how cells harboring the same genetic code, such as neuronal and muscle lineages derived from the same individual, could exhibit striking divergences in morphology and function and theorized that some regulation must exist to control gene expression in different developmental contexts. Despite the purity of Waddington's original idea, epigenetics has since been popularly repurposed as a blanket term for modifications to histones and DNA, despite conflicting reports on the heritability of these marks or their ability to influence changes in gene expression. Meanwhile, one biologically fascinating mechanism of epigenetic inheritance has been long excluded from serious scientific discussion concerning cellular information flow because of its original association with disease: prions. Discovered by Stanley Prusiner as the cause of bovine spongiform encephalopathy and Creutzfeldt-Jacob disease in humans, prions are proteins endowed with the capacity to adopt an alternative, self-replicating conformation. This conformation converts them into elements that can act as 'protein-based genes' capable of permanently transforming cellular phenotypes in a heritable fashion in the complete absence of DNA. Furthermore, because prion conformations are not encoded in nucleic acid sequence, they are inherited in a non-Mendelian fashion (all progeny in meiosis will inherit the prion, rather than half as would be the case for a mutation or none for most other epigenetic states that are meiotically reset). Despite the paradigm-shifting nature of prion proteins, only a handful of examples have been characterized. Furthermore, most attempts to identify additional prion proteins have relied on bioinformatic algorithms built on only 2 sequences and would not even have been able to identify Prusiner's original prion (PrP). In this thesis, we designed a new type of phenotypic screen to survey the true breadth of protein-based inheritance in Saccharomyces cerevisiae. We discovered almost 50 new prions, expanding the current list of such elements almost five-fold. These were highly enriched in nucleic acid binding proteins with large stretches of predicted disorder, suggesting that cells might use such unstructured domains as a novel mechanism to rewire cellular information flow. I then became interested in prion-based subversion of the central dogma and thus embarked on the biological characterization of 2 such elements recovered from our screen that were able to form protein-based epigenetic memories: Mph1 (a highly conserved DNA helicase) and Rlm1 (a conserved downstream transcription factor in MAPK signaling). I found that Mph1 was able to switch into a catalytically active, heterogeneous prion assembly following an exposure to replication stress that drives a protective DNA damage response and increased phenotypic diversification in meiosis. Rlm1, on the other hand, forms a prion regulated by phosphorylation and its natural MAPK signaling circuitry and drives a heritable memorization of a gene expression network that specializes cellular adaptation to a distinct subset of environmental conditions. In conclusion, the work presented in this thesis demonstrates prion proteins are far more abundant that previously appreciated and can serve as a novel mechanism to rapidly tune adaptation to cellular stressors, rewire patterns of gene expression, and even feedback onto the genetic code itself.

Keywords: Developmental Biology.
Advisor: Jarosz Daniel primary advisor. Thesis advisor
Kingsley David M. (David Mark) advisor. Thesis advisor
Shapiro Lucy advisor. Thesis advisor
Wandless Thomas advisor. Thesis advisor