Sexual and Apogamous Species of Woodferns Show Different Protein and Phytohormone Profiles

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 vs. 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, different regulation was found in 206 proteins of D. affinis and 166 of its sexual counterpart. The proteins upregulated in D. affinis are mostly associated to protein metabolism (including folding, transport, and proteolysis), ribosome biogenesis, gene expression and translation, while in the sexual counterpart, they account largely for starch and sucrose metabolism, generation of energy and photosynthesis. Likewise, ultra-performance liquid chromatography-tandem spectrometry (UHPLC-MS/MS) was used to assess the levels of indol-3-acetic acid (IAA); the cytokinins: 6-benzylaminopurine (BA), trans-Zeatine (Z), trans-Zeatin riboside (ZR), dyhidrozeatine (DHZ), dyhidrozeatin riboside (DHZR), isopentenyl adenine (iP), 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, iPR, the gibberellin GA4, the brassinosteoids 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 filamentous, GA3 and BL in spatulate and DHRZ in cordate gametophytes. The results point to the existence of large metabolic differences between apogamous and sexual gametophytes, and invite to consider the fern gametophyte as a good experimental system to deepen our understanding of plant reproduction.

The Shared Proteome of the Apomictic Fern Dryopteris affinis ssp. affinis and Its Sexual Relative Dryopteris oreades

“…Out of 879 quantifiable proteins [27], we detected a large number (417) that are shared by the gametophytes of apomictic DA and its sexual parent DO. These results, together with a previous study using the same two species [27], enlarge the proteomic profile of both ferns.…”

“…Out of 879 quantifiable proteins [27], we detected a large number (417) that are shared by the gametophytes of apomictic DA and its sexual parent DO. These results, together with a previous study using the same two species [27], enlarge the proteomic profile of both ferns. However, unlike for DA [25], transcriptome data for DO are lacking, such that most certainly fewer proteins could be identified in DO than in DA due to sequence divergence.…”

“…Thanks to the advent of high-throughput technologies, it has been possible to perform transcriptomics by next-generation sequencing (NGS) and shotgun proteomics by tandem mass spectrometry over recent years. However, although there are now almost 100 studies on fern transcriptomes, the proteome has been studied in only a few species: Azolla microphylla [15]; Ceratopteris richardii [16], Cyathea delgadii [17], Diplazium maximum [18], Lygodium japonicum [19], Mohria caffrorum [20], Pteris vittata [21], Pteridium aquilinum [22,23], Struthiopteris spicant [24], and also in the apogamous DA and its sexual relative DO [25][26][27]. In this regard, we previously compared the protein profiles of gametophytes of DA and DO from a total of 879 quantifiable proteins.…”

Transcriptomic analyses in the gametophyte of Dryopteris affinis: apomixis and more

“…Vegetative and reproductive development of ferns is underexplored compared with other plant groups. In this regard, the most studied aspects in ferns are: photomorphogenesis (Wada, 2007), spore germination (Salmi et al, 2005; Suo et al, 2015), cell polarity (Salmi and Bushart, 2010), cell wall composition (Eeckhout et al, 2014), reproduction (Kaźmierczak, 2010; Lopez and Renzaglia, 2014; Valledor et al, 2014; de Vries et al, 2016; Domzalska et al, 2017; Grossmann et al, 2017; Rivera et al, 2018; Wyder et al, 2020; Fernández et al, 2021; Ojosnegros et al, 2022; 2023), or adaptation to the environment (Gill et al, 2009; Thagela et al, 2016; Sareen et al, 2019). In particular, these studies show that ferns are able to survive atmospheric conditions with a higher amount of carbon dioxide than we have now, and to resist in situations of salinity, drought, or soil contamination with heavy metals (Rathinasabapathi, 2006; Wang et al, 2010).…”

Evolutionary Analysis and Functional Identification of Ancient Brassinosteroid Receptors in Ceratopteris richardii

“…Our sequence alignments reveal that the BR receptor-like proteins in ferns are more closely related to seed-plant BRL2, which cannot bind BRs. Yet, active BRs and physiological responses to BRs have been detected in ferns [ 26 , 27 , 33 ], so how BRs function in ferns remains unknown. We found that although the presumed BR receptors in Ceratopteris richardii (C-fern) cannot directly complement the dwarf phenotype of Arabidopsis bri1 mutants, their heterologous expression in Arabidopsis can activate BR responsive genes, indicating that they may have weak BR functions.…”