Poster Presentation The Annual Scientific Meeting of the Endocrine Society of Australia and the Society for Reproductive Biology 2014

Disruption of Wnt/β-catenin and ERK/MAPK signalling in the male fetal gonad impacts on the core germ cell pluripotency network (#304)

Kirsten Hogg 1 , Sonja Gustin 1 , Jessica M Stringer 1 , Patrick S Western 1
  1. MIMR-PHI Institute, Clayton, VIC, Australia

The Wnt/β-catenin and ERK/MAPK signalling pathways are major regulators of primordial germ cell (PGC) function, playing crucial roles in maintenance of pluripotency, cell cycle progression and germline fate decisions. Individual and dual inhibition of these pathways enhance the formation of murine embryonic germ (EG) cells from PGCs in vitro, presenting an informative model for efficient conversion from a partially committed to a fully pluripotent state. We studied the transcriptional response of core germ cell genes involved in pluripotency and development, and examined germ cell cycle dynamics, following inhibition of the Wnt/β-catenin and ERK/MAPK signalling pathways. We used a whole gonad culture system to assess transcriptional profiles in the context of an ex vivo environment in which somatic and germ cell interactions are maintained. Male embryonic day (E)11.5 and E12.5 whole gonads were cultured for 72 h at 37°C in the presence of Chir 99021, an inhibitor of glycogen synthase kinase 3 (preventing degradation of β-catenin), PD 0325901, an inhibitor of ERK/MAPK signalling, a combination of both inhibitory molecules (2i), or vehicle control (DMSO). Transcriptional profiling of genes regulating germ cell pluripotency and differentiation revealed differential effects of Chir and PD culture conditions. Maintenance of β-catenin signalling resulted in up-regulation of germ cell pluripotency markers Nanog and Sox2 (p<0.01) at E11.5/E12.5 time-points. In contrast, inhibition of ERK/MAPK signalling led to down-regulation of the pluripotency/male germline marker Dppa4 (p<0.05) and an increase in Stra8 (p<0.01) expression, a gene involved in entry of germ cells into meiosis. Dual inhibition using 2i revealed an additive effect of Chir and PD treatments on Dppa4 and Stra8 expression. Consistent with blocking male germline development, treatment with PD or 2i repressed the entry of male germ cells into mitotic arrest. These data indicate potentially differential effects of Wnt/β-catenin and ERK/MAPK signalling on germ cell development.