Page 266 - PC2019 Program & Proceedings

PLANT CANADA 2019

P105. Investigating the role of e3 ubiquitin ligases in the Brassicaceae self-incompatible pollen
response

*
Beronilla, P. ; D. Goring
University of Toronto
Following pollination, the dry stigmas in the Brassicaceae rapidly regulate pollen-stigma interactions,
thereby leading to the acceptance of compatible pollen or the rejection of self-incompatible pollen. The
self-incompatible pollen response inhibits pollen hydration and germination, ultimately deterring
fertilization. The objective of this research is to investigate the role of the members of the Plant-U-Box
(PUB) family of E3 Ubiquitin ligases in the self-incompatibility (SI) pathway in Brassicaceae. Previous
studies showed that the ARM-Repeat Containing 1 (ARC1) gene, which is conserved in self-incompatible
species, is required for SI as ARC1-knockdown constructs in transgenic Brassica napusand Arabidopsis
lyrata confer a partial breakdown of SI. What remains unknown is how the SI response would be affected
when ARC1 is completely knocked out and whether the closely-related PUB17 gene may play a
redundant function. The two research goals to approach this question both utilize the CRISPR/Cas9
system to generate knockout deletion mutations. First, ARC1 deletion mutations will be generated in
compatible Capsella rubellaand then crossed into self-incompatible Capsella grandiflora. Second, a self-
incompatible Arabidopsis thaliana C24 line carrying the A. lyrataSCRband SRKbtransgeneswill be used
to investigate PUB17 as this line has a robust SI phenotype despite lacking a functional ARC1 protein.
Currently, CRISPR/Cas9 constructs are being used to generate ARC1 and PUB17 knockouts and SI in the
mutants will be observed through phenotypic assays. Overall, this research will further elucidate the role
of E3 ubiquitin ligases in the rejection of self-incompatible pollen in the SI pathway.

Paula Beronilla (paula.beronilla@mail.utoronto.ca)

P106. Characterization of Camelina sativa germination: The effect of gibberellins on vacuolation
*
Gomes, M. ; E. Nambara
University of Toronto

Camelina sativa is an important oilseed crop, which has recently gained attention in biofuel production as
more information emerges about its genomic characteristics and crop performance. The aim of this
research is to compare germination of Camelina with that of Arabidopsis, as molecular mechanisms of
germination are well studied in this species. Arabidopsis protein storage vacuoles (PSVs) increase in size
and decrease in number as a consequence of germination progression. Through diphenyl boric acid
aminoethyl ester (DPBA) staining of seed embryo and endosperm, Camelina showed similar results to
Arabidopsis, however with distinct time points of vacuolation. The rate of this process increases by the
addition of gibberellins (GA). Vacuolation was increased in seeds treated with exogenous GA and
compared to seeds where endogenous GA was inhibited by paclobutrazol, a GA inhibitor. This result
indicates that, similar to Arabidopsis, vacuolation is an indication of germination in Camelina and can be
regulated by GA. It was also observed that micropylar regions of the endosperm had higher ratios of
vacuolated cells, suggesting that these regions may have different roles in nutrient transfer and may be
more sensitive to hormonal regulation during germination.

Malaika Gomes (malaika.gomes@mail.utoronto.ca)

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