Genetic and molecular characterization of apospory in Paspalum sp.

ORTIZ, JUAN PABLO AMELIO; PESSINO, SILVINA C.; QUARIN, CAMILO L.; PUPILLI, FULVIO; STEIN, JULIANA; MARTÍNEZ, ERIC J.; ESPINOZA, FRANCISCO; FELITTI, S. A.; RODRIGUEZ, MARÍA P.; LASPINA, N.; SIENA, LORENA A.; OCHOGAVIA, A.C.; PODIO, M.; SARTOR, M.; HOJSGAARD, DIEGO H.; URBANI, MARIO H.

Brasilia, Brasil

Congreso; XX International Congress on Sexual Plant Reproduction; 2008

International Association of Sexual Plant Reproduction Research – IASPRR

The genus Paspalum comprises numerous grass species that are important forage resources for the tropical and subtropical regions of the Americas. Ploidy levels within the genus range from diploid to 16-ploid, with modes of reproduction from allogamy to apomixis. An important proportion of Paspalum species form agamic complexes, where diploid sexual self-incompatible cytotypes, have pseudogamous, self-compatible apomictic polyploid counterparts. Apospory is the more frequent type of gametophytic apomixis in the genus: megagametophytes develop directly from sporophytic nucelar cells (2n) and show diverse structural organization. The aposporous embryo sac configuration usually comprise: the egg cell and a large central cell bearing two polar nuclei, and may include one or two synergids but antipodal cell are usually absent. The parthenogenetic development of the embryo from the egg cell, and the formation of the endosperm after the fertilization of the polar nuclei (pseudogamy) completes seed formation. Apospory and meiotic embryo sacs can occur simultaneously in the same plant and even in the same ovule. Gametophytic apomixis tends to occur in polyploids, and then most often at the tetraploid or greater levels. Over the last 10 years our group has worked on the characterization of apomixis in the genus Paspalum mainly focussing on: 1) the mode of inheritance of apospory in tetraploid P. notatum and the identification of molecular markers linked to the trait, 2) the generation of a genetic map of the species and the characterization of the linkage group carrying the apospory locus, 3) the detection of genes and pathways involved in the aposporous and meiotic embryo sac development in P. notatum and 4) the study of the functionality of apomixis components at the diploid level in P. rufum. Inheritance of apospory in P. notatum was investigated by using the tetraploid (2n=4x=40) genotypes Q4188 (a fully sexual plant used as pistilate parent) and Q4117 (a natural obligate apomictic plant employed as pollen donor). Genetic analysis of F1 progenies and several segregating populations derived from it indicated that the trait is controlled by a single dominant locus with a distorted segregation ratio, possibly due to a pleiotropic partial lethal effect, or a partial linkage to a lethal factor. Pollen viability determinations revealed a significantly higher level of non-viable pollen in the aposporous genotype Q4117 than in the sexual Q4188 and analysis of male meiosis of both genotypes showed that Q4117 could present a genomic rearrangement. A full genetic linkage map of the species was developed based on an F1 family derived from the same parental plants using single dose AFLP markers. In general, polysomic inheritance was observed for most markers, but the linked group carrying apospory showed preferential chromosome pairing and a strong restriction in recombination. Several AFLP markers 100% linked to apospory were detected. Results obtained so far denote apospory in P. notatum is controlled by a complex locus spanning a large non-recombinant chromosomal segment, which includes non-coding sequences, retroelements and cytosine methylation. The apospory region in P. notatum is related to rice chromosomes 2 and 12. Transcriptome surveys aimed at the identification of genes differentially expressed in aposporous and sexual genotypes of P. notatum allowed isolation of 65 candidates, out of which two thirds (45) could be successfully annotated. Several of them were identical to those related to apospory in Poa pratensis. Sequences belong to signal transduction (including several members of an ERK cascade), transcription, proteolysis, cell cycle control, cell wall composition and repetitive elements functional classes. A subgroup of candidates silenced in aposporous plants mapped close to apo-region in P. notatum. Curiously, RNA in situ tissue hybridization showed that many of the genes are differentially expressed not only in the ovule but also in other tissues like the tapetum of anthers and pollen mother cells. On the other hand, the previous observation of ovules containing an aposporous embryo sac besides the meiotic one in the diploid genotype of P. rufum Q3754, suggested that at least some species have the potential for apomictic reproduction at the diploid level. Progeny tests carried out on 2 experimental families derived from a controlled cross (H1) and the induced self-pollination of Q3754 (S1) showed that all progenies of H1 derived from sexual reproduction (n + n), but 5 out of 95 plants from S1 were of clonal origin (2n + 0). Further experiments, carried out on a third family (M1) obtained after crossing Q3754 with the tetraploid plant Q3785 showed diploids (75 %), triploids (20 %) and tetraploid (5 %) progenies. Triploids and the tetraploids may have originated from functional aposporous embryo sacs. Likewise, the reconstruction of the developmental route of individual seeds demonstrated that 27.5 % of them derived from fertilized aposporous embryo sacs (2n + n). These results indicate that components of gametophytic apomixis are effectively expressed at the diploid level in P. rufum and could be the foundation of a recurrent auto-polyploidization process in the species.