high-resolution-c-elegans-imaging-across-all-larval-stages

High-Resolution <em>C. elegans</em> Imaging Across All Larval Stages

Caenorhabditis elegans has become one of the most widely studied and best-understood animal models in biology. Three features are key to C. elegans' ...

University Zürich

The anchor cell (AC) in  secretes an epidermal growth factor (EGF) homolog that induces adjacent vulval precursor cells (VPCs) to differentiate. The EGF receptor in the nearest VPC sequesters the limiting EGF amounts released by the AC to prevent EGF from spreading to distal VPCs. Here, we show that not only EGFR localization in the VPCs but also EGF polarity in the AC is necessary for robust fate specification. The AC secretes EGF in a directional manner towards the nearest VPC. Loss of AC polarity causes signal spreading and, when combined with MAPK pathway hyperactivation, the ectopic induction of distal VPCs. In a screen for genes preventing distal VPC induction, we identified  and  as genes specifically required for polarized EGF secretion.  and  mutants exhibit errors in vulval fate specification, reduced precision in VPC to AC alignment and increased variability in MAPK activation.  encodes a seven-pass transmembrane receptor acting in the AC and  a neuropeptide-like protein expressed in the VPCs. SRA-9 and NLP-26 may transduce a feedback signal to channel EGF secretion towards the nearest VPC.

Polarized epidermal growth factor secretion ensures robust vulval cell fate specification in .

Several microfluidic-based methods for Caenorhabditis elegans imaging have recently been introduced. Existing methods either permit imaging across multiple larval stages without maintaining a stable worm orientation, or allow for very good immobilization but are only suitable for shorter experiments. Here, we present a novel microfluidic imaging method that allows parallel live-imaging across multiple larval stages, while maintaining worm orientation and identity over time. This is achieved through an array of microfluidic trap channels carefully tuned to maintain worms in a stable orientation, while allowing growth and molting to occur. Immobilization is supported by an active hydraulic valve, which presses worms onto the cover glass during image acquisition only. In this way, excellent quality images can be acquired with minimal impact on worm viability or developmental timing. The capabilities of the devices are demonstrated by observing the hypodermal seam and P-cell divisions and, for the first time, the entire process of vulval development from induction to the end of morphogenesis. Moreover, we demonstrate feasibility of on-chip RNAi by perturbing basement membrane breaching during anchor cell invasion.

Microfluidic-based imaging of complete Caenorhabditis elegans larval development.

hox genes encode a conserved family of homeodomain transcription factors that are essential to determine the identity of body segments during embryogenesis and maintain adult somatic stem cells competent to regenerate organs. In contrast to higher organisms, somatic cells in C. elegans irreversibly exit the cell cycle after completing their cell lineage and the adult soma cannot regenerate. Here, we show that hox gene expression levels in C. elegans determine the temporal competence of somatic cells to proliferate. Down-regulation of the central hox gene lin-39 in dividing vulval cells results in their premature cell cycle exit, whereas constitutive lin-39 expression causes precocious Pn.p cell and sex myoblast divisions and prolongs the proliferative phase of the vulval cells past their normal point of arrest. Furthermore, ectopic expression of hox genes in the quiescent anchor cell re-activates the cell cycle and induces proliferation until young adulthood. Thus, constitutive expression of a single hox transcription factor is sufficient to prolong somatic cell proliferation beyond the restriction imposed by the cell lineage. The down-regulation of hox gene expression in most somatic cells at the end of larval development may be one cause for the absence of cell proliferation in adult C. elegans.

Prolonging somatic cell proliferation through constitutive hox gene expression in C. elegans.

Vitamin B12 is an essential nutritional co-factor for the folate and methionine cycles, which together constitute one-carbon metabolism. Here, we show that dietary uptake of vitamin B12 modulates cell fate decisions controlled by the conserved RAS/MAPK signaling pathway in C. elegans. A bacterial diet rich in vitamin B12 increases vulval induction, germ cell apoptosis and oocyte differentiation. These effects are mediated by different one-carbon metabolites in a tissue-specific manner. Vitamin B12 enhances via the choline/phosphatidylcholine metabolism vulval induction by down-regulating fat biosynthesis genes and increasing H3K4 tri-methylation, which results in increased expression of RAS/MAPK target genes. Furthermore, the nucleoside metabolism and H3K4 tri-methylation positively regulate germ cell apoptosis and oocyte production. Using mammalian cells carrying different activated KRAS and BRAF alleles, we show that the effects of methionine on RAS/MAPK-regulated phenotype are conserved in mammals. Our findings suggest that the vitamin B12-dependent one-carbon metabolism is a limiting factor for diverse RAS/MAPK-induced cellular responses.

Article	Year
Polarized epidermal growth factor secretion ensures robust vulval cell fate specification in . Development (Cambridge, England)\| PubMed ID: 32439759	2020
Microfluidic-based imaging of complete Caenorhabditis elegans larval development. Development (Cambridge, England)\| PubMed ID: 34170296	2021
Prolonging somatic cell proliferation through constitutive hox gene expression in C. elegans. Nature communications\| PubMed ID: 37891160	2023
Nutritional vitamin B12 regulates RAS/MAPK-mediated cell fate decisions through one-carbon metabolism. Nature communications\| PubMed ID: 39289374	2024

Alex Hajnal

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