Transcriptomic analyses in the gametophytes of the apomictic fern Dryopteris affinis

Abstract

The gametophyte of ferns reproduces either by sexual or asexual means. In the latter, apogamy represents a peculiar case of apomixis in which an embryo is formed from somatic cells. A proteomic and physiological approach was applied to the apogamous fern Dryopteris affinis ssp. affinis and its sexual relative D. oreades. The proteomic analysis compared apogamous versus female gametophytes, whereas the phytohormone study included, in addition to females, three apogamous stages (filamentous, spatulate and cordate). The proteomic profiles revealed a total of 879 proteins and, after annotation, differential regulation was found in 206 proteins of D. affinis and 166 of its sexual counterpart. The proteins upregulated in D. affinis were mostly associated with protein metabolism (including folding, transport and proteolysis), ribosome biogenesis, gene expression and translation, whereas in the sexual counterpart they were largely related to starch and sucrose metabolism, energy generation and photosynthesis. Ultra-performance liquid chromatography–tandem spectrometry (UHPLC–MS/MS) was used to assess the levels of indole-3-acetic acid (IAA); the cytokinins 6-benzylaminopurine (BA), trans-zeatine (Z), trans-zeatin riboside (ZR), dihydrozeatine (DHZ), dihydrozeatin riboside (DHZR), isopentenyl adenine (iP) and isopentenyl adenosine (iPR); abscisic acid (ABA); the gibberellins GA3 and GA4; salicylic acid (SA); and the brassinosteroids brassinolide (BL) and castasterone (CS). IAA, the cytokinins Z, ZR and iPR, the gibberellin GA4, castasterone and ABA accumulated more in the sexual gametophyte than in the apogamous one. When comparing the three apogamous stages, BA and SA peaked in the filamentous stage, GA3 and BL in the spatulate stage, and DHZR in the cordate gametophytes. Overall, the results reveal marked metabolic differences between apogamous and sexual gametophytes and support the use of fern gametophytes as a valuable experimental system to deepen our understanding of plant reproduction.