GAMETE EXPRESSED PROTEIN1 ortholog in Marchantia polymorpha is essential for male pronucleus formation in fertilized egg cells

GAMETE EXPRESSED PROTEIN1 (GEX1) is a gamete nuclear membrane protein essential for nuclear fusion during sexual reproduction in Arabidopsis thaliana. GEX1 orthologs have been identified in several terrestrial plants. The liverwort Marchantia polymorpha, which uses motile gametes (spermatozoids) for fertilization, possesses a GEX1 homolog, M. polymorpha GEX1 (MpGEX1). In the present study, we investigated the function of MpGEX1 in M. polymorpha. MpGEX1 is expressed during both male and female gametogenesis. During male gametogenesis, MpGEX1 was detected in the nuclear envelope of spermatogenous cells and spermatids but not in mature spermatozoids. In female plants, MpGEX1 was detected in the nuclear envelope of mature egg cells. Mpgex1 mutant strains were not defective in egg or spermatozoid​ formation. However, Mpgex1 mutant female plants were defective in fertilization when crossed with wild-type spermatozoids. In fertilized egg cells, the male pronucleus, formed from the sperm nucleus, fuses with the female nucleus. However, fertilized Mpgex1 mutant egg cells lack an egg cell-derived nuclear envelope in their male pronuclei. The aberrant pronuclei were elongated or oval and remained separated from female nuclei. Fertilized Mpgex1 mutant egg cells do not undergo zygotic cell divisions. Female-derived mCitrine-MpGEX1 was transiently localized to the male pronucleus shortly after fertilization in females. These results demonstrate that maternally supplied MpGEX1 is essential for male pronucleus formation in M. polymorpha. Our findings reveal a novel role for the plant GEX1 ortholog in fertilization, suggesting the functional diversification of this conserved protein family during the evolution of plant fertilization mechanisms.