Germ Plasm Aggregation and Compaction in the Early Zebrafish Embryo

Eno, Celeste

Abstract

Germ plasm movement and early acto-myosin movement in the Zebrafish embryo. One of the earliest cell-fate decisions in animal development is the determination of primordial germ cell (PGCs) differentiation, which occurs via inductive cell-cell interactions (e.g. mammals) or preformation (e.g. teleosts, Drosophila, C. elegans). Preformative (maternally inherited) germ plasm is a specialized structure made up of ribonucleoparticles (RNPs), proteins and often associated with cytoskeletal components. The zebrafish model is excellent for studying the movement of germ plasm in early embryogenesis. The transparent embryos are easily manipulated using drugs, as well as morpholino and RNA injections. Recently, members of our laboratory have refined a protocol for manipulation of maternal factors acting in the embryo by injection of reagents into oocytes undergoing in vitro maturation, which has opened the possibility of manipulation prior to and immediately after fertilization. My preliminary data suggest that phosphomyosin is present in germ plasm RNPs during their multimerization prior to and during furrow formation, and that these RNPs reside on F- actin. We propose a mechanism by which myosin II-driven F-actin sliding aggregates germ plasm particles and moves the aggregates into the first two forming furrows. Later, during furrow maturation, germ plasm RNPs continue to undergo multimerization until they form compact masses at the distal ends of these furrows. A maternal effect lethal mutant, aura, does not have the concentric actin rings seen in wild type embryos and do not properly aggregate germ plasm RNPs. This suggests that the protein aura codes for, Mid1ip1L, is involved in the cytoskeletal dynamics of germ plasm early movement. I plan to address these hypotheses by characterizing germ plasm aggregation and recruitment on actin filaments prior to and at furrow initiation, and determining the role Mid1ip1L plays in early cytoskeletal dynamics. The proposed hypothesis of GP RNP movement will provide details on a novel mechanism for the movement of such cellular determinants via the action of a myosin motor on an actin cytoskeletal network. The mechanisms studied will lead to understanding the proper movement of PGC-determining factors, relating to cell stemness, reproduction, carcinogenesis and pluripotency.

Public Health Relevance

Proposed aims will define the functional requirements of germ plasm multimerization during the early zebrafish cell cycles. Because germ plasm factors share functions involved in not only germ cell determination but also pluripotency, understanding the movement of germ plasm in zebrafish may result in technologies to better manipulate germ cells in vertebrate species. In addition, most studied cases of intracellular movement of cell determinants involves transport along microtubules, and the proposed hypothesis of GP RNP movement will provide mechanistic details on a novel mechanism for the movement of such cellular determinants via the action of a myosin motor on an actin in the cytoskeletal network.