Asexual Female Gametogenesis Involves Contact with a Sexually-Fated Megaspore in Apomictic Hieracium

Abstract

Apomixis results in asexual seed formation where progeny are identical to the maternal plant. In ovules of apomictic species of the subgenus, meiosis of the megaspore mother cell generates four megaspores. Aposporous initial (AI) cells form during meiosis in most ovules. The sexual pathway terminates during functional megaspore (FM) differentiation, when an enlarged AI undergoes mitosis to form an aposporous female gametophyte. Then, the mitotically programmed FM dies along with the three other megaspores by unknown mechanisms. Transcriptomes of laser-dissected AIs, ovule cells, and ovaries from apomicts and AI-deficient mutants were analyzed to understand the pathways involved. The steps leading to AI mitosis and sexual pathway termination were determined using antibodies against arabinogalactan protein epitopes found to mark both sexual and aposporous female gametophyte lineages at inception. At most, four AIs differentiated near developing megaspores. The first expanding AI cell to contact the FM formed a functional AI that underwent mitosis soon after megaspore degeneration. Transcriptome analyses indicated that the enlarged, laser-captured AIs were arrested in the S/G2 phase of the cell cycle and were metabolically active. Further comparisons with AI-deficient mutants showed that AIs were enriched in transcripts encoding homologs of genes involved in, and potentially antagonistic to, known FM specification pathways. We propose that AI and FM cell contact provides cues required for AI mitosis and megaspore degeneration. Specific candidates to further interrogate AI-FM interactions were identified here and include arabinogalactan protein family genes.

Controlling Apomixis: Shared Features and Distinct Characteristics of Gene Regulation

“…LAM in combination with RNA-Seq recently has also shed light onto the cell type specification of the aposporous initial cell (AIC) as compared to early developing embryo sacs and somatic ovule tissues in Hieracium praealtum. These studies suggest advanced acquisition of gametophytic fate by the AIC [108,109] (Table 2).…”

“…It encodes a GH3 family protein involved in modulation of auxin response. GH3 was further identified as higher expressed in AICs than in embryo sacs of Hieracium praealtum, further suggesting its importance for megasporogenesis in apomicts [109]. Also the homologue of the nonethylene receptor HISTIDINE KINASE1 (HK1) is higher expressed in apomictic as compared to sexual nucelli in Boechera.…”

“…Especially during megasporogenesis and preparation of the MMC/AIC for (apo)meiosis, processes related to translation and ribosome biogenesis are enriched both in sexual A. thaliana and apomictic Boechera [103,107]. Further evidence for the importance of ribosome biogenesis and function, and nucleosome assembly for megasporogenesis in apomicts comes from transcriptome analysis of Hieracium subgenus Pilosellum, as related functions are enriched in the AIC as compared to developing embryo sacs [109]. While it has long been noticed that a cycle of ribosome degradation and reassembly is associated with plant meiosis [117], the relevance of this processes for sexual and apomictic reproduction has not been described in detail to date.…”

Gene expression in isolated cowpea (Vigna unguiculata L. Walp) cells from meiosis to seed initiation

“…Cell cycle genes are relatively well conserved across plant species (Wang et al, 2004, Scofield et al, 2014). Analyses of the expression of cell cycle genes known to block and/or promote passage through various phases of the cell cycle in isolated cell types has been used to predict the stage of cell cycle arrest (Juranic et al, 2018, Sornay et al, 2015, Sukawa and Okamoto, 2018, Velappan et al, 2017, Komaki and Sugimoto, 2012). A core set of cell cycle genes from Arabidopsis was used to identify 141 non-redundant cowpea genes encoding proteins with various roles in cell cycle regulation, including 46 cyclins (CYC), 12 cyclin-dependent kinases (CDKs) and 5 CDK inhibitors (interactor/inhibitor of cyclin-dependent kinases/Kip-related proteins; KRP/ICKs) in addition to other key regulators (Supplemental Table 15 and 16).…”

“…In each case the best match based on bit-score was chosen for further analysis. To identify cell-cycle related cowpea genes, the amino acid sequences of a set of cell-cycle genes from Arabidopsis (Juranic et al, 2018) were used in BlastP searches against the predicted cowpea protein sequences. BlastP results with an e value < 1e-20 were chosen, and the annotation of cowpea sequences matching multiple Arabidopsis genes was chosen based on the highest bit score.…”