Page 263 - PC2019 Program & Proceedings

PLANT CANADA 2019

TOPIC 8: Cell Biology (Posters P99-P109)

P99. Characterization of the pokeweed antiviral protein (PAP) interactome by proximity-
dependent biotin identification
*
Chivers, J. ; K. Hudak
York University

Ribosome-inactivating proteins (RIPs) are produced primarily by plants, and are named for their
enzymatic ability to remove a purine base from the sarcin-ricin loop of the large ribosomal RNA.
However, little is known about the biological function of RIPs in the plant. Phytolacca americana, the
American pokeweed, produces a RIP called pokeweed antiviral protein (PAP). We suspect that this
protein is involved in defense against pathogens and adaptation to environmental stress. The objective of
this work is to investigate the biological role of PAP by mapping out the PAP-protein interactome.
Proximity dependent biotin ligase labelling (BioID) will be used to identify PAP interactors in both the
apoplast, where PAP has been shown to primarily localize, and the cytoplasm, where the ribosomal target
resides. PAP-biotin ligase fusions will be expressed transiently in Nicotiana tabacum, followed by affinity
purification of biotinylated proteins and their identification using mass spectrometry. These protein
interactors are important to understand how PAP functions in plant systemic responses while sequestered
in the apoplast, as well as identify ribosomal proteins that mediate rRNA depurination by PAP in the
cytoplasm. This work will represent the first protein interactome mapping for a RIP; identification of PAP
interactors in living plant cells will help elucidate PAP’s function and contribute to characterization of the
biological role of RIPs in general.

Jennifer Chivers (jchivers@yorku.ca)

P100. Understanding differential development and behaviour of plastids using the Arabidopsis
immutans mutant
*
Burnside, M. ; K. Barton; N. Mathur; J. Mathur
University of Guelph

Plastids are characteristic organelles of plants. All plastids arise from pro-plastids and subsequent
differentiation allows their categorization into three broad types; leucoplasts when they remain colourless,
chloroplasts when they develop chlorophyll and chromoplasts when they predominantly have pigments
other than chlorophyll. While the plastid types can interconvert, they are usually located in different
regions and organs of a plant. Thus, chloroplasts are mainly found in leaves while leucoplasts occur
primarily in roots. Their different locations make it difficult to study the cellular factors involved in
plastid interconversions and the mechanisms whereby one particular type may predominate in a cell. An
Arabidopsis thaliana mutant, immutans (im) develops leaves with randomly distributed green and white
sectors corresponding to the presence of chloroplasts and leucoplasts, respectively. The introduction of a
stroma targeted GFP revealed a range of plastid sizes and shapes in the mutant and as observed by
transmission electron microscopy the different shapes correlated with differences in internal membrane
architecture. Additional fluorescent protein probes targeted to the endoplasmic reticulum, peroxisomes,
and mitochondria have been introduced in im to obtain more insights on the interactions of chloroplasts
and leucoplasts with other organelles. The creation of transgenic im lines has resulted in a new set of
fluorescent protein-based tools for studying the development and interconversion of plastids and
understanding their differential responses to environmental stimuli.

Mariel Burnside (mburnsid@uoguelph.ca)

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