Genetische regulatoren van dooiereiwitproductie in de rondworm Caenorhabditis elegans

While invertebrate reproduction has been actively researched during thepast decades, its neuroendocrine control mechanisms have somehow escaped attention. In sharp contrast, the key features of the neuroendocrine hypothalamus-pituitary-gonad axis, which is central to vertebrate reproductive control, are well known. One downstream process driven by this regulatory system is maternal yolk formation, a process called vitellogenesis, which is thought to be a critical step of reproduction in not only vertebrate but all egg-laying animals.This work aimed to gain more insight into the (neuroendocrine) regulation of nematode reproduction by specifically exploring the molecular genetic control of yolk protein production in the model organism Caenorhabditis elegans . In the first and major part of this work, a simple forward genetic screen (~7,500 mutagenized haploid genomes) was performed to identify mutants with aberrant expression of a green fluorescent protein (GFP)-tagged yolk protein transgene. Twenty-five such mutants were recovered andten selected for further identification of the causative mutations via a combined mapping and whole-genome sequencing strategy. This genome-wide screening effort identified one allele each of three previously identified genes involved in trafficking of yolk its receptor as well as one involved in transgene silencing, validating the screen. Further analysis using transgene rescue confirmed that the remaining six mutant alleles corresponding to four novel regulatory loci indeed rather robustly disrupt yolk protein synthesis in C. elegans. The transcriptional and translational data on each of these latter molecular players support the view that parallel pathways, some of which contain evolutionarily unique regulators, exert control over yolk protein production in C. eleg ans. In the second part of this work, a set of ~200, mainly neuropeptidergic butalso some steroidogenic, candidate genes was interrogated bymeans of RNA interference to be involved in the regulation of the expression of different GFP-tagged yolk protein transgenes. As such, twenty-four novel potential regulators, believed to rather subtly stimulate yolk protein production in C. elegans , were identified, including fourteen neuropeptide G protein-coupled receptors, three neuropeptide-encoding precursors and two steroidogenic enzymes. The revelation of these latter components truly motivates to continue to address the interaction between steroid signaling and nematode reproductive control, similar to many other (in)vertebrate systems. By analogy with their far better characterized(putative) counterparts in other animals, several of the newly discovered potential yolk protein regulators in C. elegans emphasize possible striking direct/indirect similarities between the reproductive control machinery in Nematoda and diverse other invertebrate and vertebrate phyla. It is not so much the central regulatory axis that appears tobe conserved but in particular the peripheral (e.g. metabolic) controlling factors. Additionally, some potential ecdysozoan-, nematode- and Caenorhabditis -specific genetic determinants of C. elegans yolk protein production were uncovered as well. Taken together, both the core genetic regulatory network controlling C. elegans yolk protein production proposed in this work and all implicated robust yolk protein regulating mutants offer a platform which rather easily enables to further unambiguously fit in the possible subtle regulators of yolk protein synthesis, for example by obtaining sensitive transcriptional data. Revealing such crosstalk and/or interdependency between robust and subtle regulators will allow to further elucidate the (neuroendocrine) control of yolk protein production - and thus reproduction -in nematodes.status: publishe