nematode and water bear

Research: Evolution of Germline Development

Nearly all sexually-reproducing animals produce germ cells (cells that give rise to eggs and sperm). Germ cells are totipotent - they can produce all cell types in organisms. Proper specification of germ cells is essential for life; failure to make germ cells can lead to sterility. To preserve their totipotency, germ cells must be protected from factors that would cause them to differentiate into somatic cells (such as muscle, neurons, skin, etc.).

Germ cells in multiple organisms have common traits, including the expression of a highly-conserved DEAD-box RNA helicase Vasa, the presence of specific chromatin methylation patterns, and cessation of transcription. However, organisms appear to use a variety of mechanisms to ensure that germ cells acquire the correct fate at the correct time. How diverse are these mechanisms? Are there common rules that govern germline development? To better understand how germ cell fate is specified, and how germline development has evolved among animals, I am studying this question in two invertebrate species.

Tardigrades as a new model system for germline development

Ecdysozoan phylogeny (left) and adult tardigrade (right) I have been developing the tardigrade Hypsibius dujardini as a model for understanding germ cell fate specification. Tardigrades (also known as water bears) are small, segmented invertebrates that have become popular organisms for studying survival in extreme conditions. Tardigrades comprise a phylum closely related to arthropods, nematodes and other Ecdysozoans (invertebrates that molt) (Fig. 1), and their evolutionary position can help us to better understand how similarities and differences in germline development have arisen in well-studied model organisms.

I recently published a protocol to disrupt gene function by RNA interference (RNAi). RNAi is a well-conserved mechanism by which messenger RNA encoding a known gene is degraded, preventing translation of the encoded protein, and producing "loss-of-function" phenotypes (Fig. 2). Dr. Bob Goldstein at the University of North Carolina-Chapel Hill has initiated a genome sequencing project for H. dujardini, and as the genome becomes available, we will eventually be able to target any gene for disruption by RNAi.

Fig. 2: RNAi results in target-specific depletion of gene function. a-c Representative images of wild-type embryos. a Stage 13 embryo (~24 hours after egg-laying), showing elongation along the anterior-posterior axis, with ectodermal segmentation (red arrows). b Same embryo as in (a), at late Stage 15 (~48 hrs after egg-laying), showing three developing limb buds (asterisks). Intestinal birefringent granules are visible in a higher focal plane. c Stage 19 embryo (~4 days after egg laying), prior to hatching. The pharynx and intestine are outlined (white dotted lines). Yellow arrows in this panel and in panel (e) mark birefringent granules, a marker of intestine differentiation. Note that this embryo is different from the one shown in a, b. d Hd-act-1(RNAi) embryo, ~24 hours after egg-laying. e Hd-mag-1(RNAi) embryo, ~48 hrs after egg-laying. The embryo has not elongated along the anterior-posterior axis, but birefringent granules are visible. f Hd-dlc-1(RNAi) embryo, ~4 days after egg-laying. The pharynx and part of the intestine are outlined (white dotted lines). The intestine appears to lack birefringent granules. A=anterior; P=posterior.

Some of the projects that I hope to have SPU students working on include:

Germline development in nematodes

As another approach to understanding the evolution of germline development, I am interested in the extent to which mechanisms of germline development are conserved within the nematode phylum. Much of what we know about cell fate specification and early germline development in nematodes comes from studies in C. elegans. However, it is not clear how cell fates are specified in other nematode species. Analysis of multiple nematode genome sequences suggests that some germline-specific proteins required for C. elegans development are absent in non-Caenorhabditis species. What mechanisms do non-Caenorhabditis species use to ensure the proper segregation of somatic and germ cell fates?

Some of the projects that I hope to have SPU students working on (both in my lab, and in lab courses that I teach) include:

Note: I am open to mentoring other projects if students have specific questions they are interested in studying using tardigrades or nematodes, and that can be studied using the resources available in my lab and at SPU.

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