BY‐2 Cells: Culture and Transformation for Live Cell Imaging

Brandizzí, Federica; Irons, Sarah L.; Kearns, Anne; Hawes, Chris

doi:10.1002/0471143030.cb0107s19

Cited by 48 publications

(35 citation statements)

References 32 publications

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“…Shown are representative epifluorescence micrographs of tobacco (Nicotiana tabacum) Bright Yellow-2 (BY-2) suspensioncultured cells, which serve as a well-characterized system for studying protein localization in plant cells. 18 BY-2 cells were transiently transformed via biolistic bombardment with plasmid Dna encoding full-length guayule SrPP 12 (GenBank number aF541942) C-terminally fused to the n terminus of the green fluorescent protein (SrPP-GFP). Following bombardment, cells were incubated in linoleic acid, which induces a dramatic increase in the number and size of lipid droplets in these cells (see ref.…”

Section: Figure 2 Expression Pattern Of Ldaps In Oil Palm (E Guineementioning

confidence: 99%

Lipid droplet-associated proteins (LDAPs) are involved in the compartmentalization of lipophilic compounds in plant cells

Gidda

Watt²,

Collins-Silva

et al. 2013

Plant Signaling & Behavior

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Lipid droplets play an essential role in the life cycle of plants by housing lipid storage compounds, usually triacylglycerols, in seeds that are mobilized to support post-germinative growth, prior to photosynthetic establishment. Given this critical role in plant growth and development, the majority of research on plant lipid droplets has focused on their function in seed tissues. For instance, purification of lipid droplets from plant oilseeds resulted in the identification and characterization of the oleosins, which are an abundant class of lipid droplet-surface-associated proteins important for stabilizing lipid droplets during seed desiccation 1-3 and possibly serving as sites for recruitment of lipases that facilitate the breakdown of stored triacylglycerols during seedling establishment. 4 It is now appreciated, however, that lipid droplets have numerous functions beyond lipid storage in seeds and that they are present in nearly all plant cell types, many of which do not accumulate appreciable amounts of lipid, such as the cells in leaves, stems, and roots.5 There is also emerging evidence that lipid droplets are highly dynamic organelles involved in a variety of cellular processes and physiological responses, some of which appear to be conserved among eukaryotes. [6][7][8] Nevertheless, the precise functions of lipid droplets in non-seed cell types in plants are currently poorly understood.In an effort to increase our understanding of lipid droplet biogenesis and functions in plants, we recently characterized the proteome of lipid droplets isolated from the mesocarp of avocado (Persea americana).9 This tissue was selected for analysis since it is a rich source of non-seed lipid droplets that lack the abundant oleosins found in oilseed tissues. Briefly, proteins enriched in the isolated avocado mesocarp lipid droplet fraction were identified using a combination of multi-dimensional protein identification technology and peptide mass fingerprinting, using an avocado RNAseqderived "proteome" for query. Two of the most abundant proteins associated with these lipid droplets were highly similar (86%) in sequence to each other and, thus, were annotated as lipid dropletassociated protein 1 (LDAP1) and LDAP2 ( Fig. 1A; Pam_LDAP1 and Pam_LDAP2). We also showed previously that LDAP1 and LDAP2 gene expression during development of avocado mesocarp increased in correlation with oil accumulation.9 Interestingly, transcriptome analysis of various tissues of oil palm (Elaeis guineensis) revealed the presence of three LDAP-like genes, of which one of While lipid droplets have traditionally been considered as inert sites for the storage of triacylglycerols and sterol esters, they are now recognized as dynamic and functionally diverse organelles involved in energy homeostasis, lipid signaling, and stress responses. unlike most other organelles, lipid droplets are delineated by a half-unit membrane whose protein constituents are poorly understood, except in the specialized case of oleosins, which are associated with seed li...

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Section: Figure 2 Expression Pattern Of Ldaps In Oil Palm (E Guineementioning

confidence: 99%

Lipid droplet-associated proteins (LDAPs) are involved in the compartmentalization of lipophilic compounds in plant cells

Gidda

Watt²,

Collins-Silva

et al. 2013

Plant Signaling & Behavior

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“…21,22 Overall, the data show that plant cells, like mammalian cells, but unlike Saccharomyces, do not accumulate tRNAs derived from intronless and intron-containing precursors in their nucleus during nutrient deprivation, indicating that nutrient-related regulation of nuclear tRNA export is not entirely conserved among evolutionarily diverse organisms.…”

Section: Discussionmentioning

confidence: 89%

Plants, like mammals, but unlikeSaccharomyces,do not regulate nuclear-cytoplasmic tRNA trafficking in response to nutrient stress

Johnstone

Mullen²,

Mangroo³

2011

Plant Signaling & Behavior

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“…Given the well-documented role of Tsg101 (the mammalian homolog of Vps23) in HIV budding from the plasma membrane in mammalian cells (21,22) and that yeast Vps23p is redirected to peroxisomes in yeast cells coexpressing TBSV p33 (23), we tested whether, in an analogous manner, CIRV causes the relocalization of Vps23 to mitochondria in plant cells. Toward this end, N. tabacum BY-2 suspension-cultured cells-which serve as a well-characterized system for studying protein targeting in plant cells (57), including viral proteins (58, 59)-were transiently (co)transformed (via biolistic bombardment) with plasmids encoding fulllength CIRV and/or an N-terminal Myc epitope-tagged version of Arabidopsis Vps23 (Myc-Vps23) and then processed for immunoepifluorescence microscopy.…”

Section: Resultsmentioning

confidence: 99%

A Unique N-Terminal Sequence in the Carnation Italian ringspot virus p36 Replicase-Associated Protein Interacts with the Host Cell ESCRT-I Component Vps23

Richardson

Clendening

Sheen

et al. 2014

J Virol

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Like most positive-strand RNA viruses, infection by plant tombusviruses results in extensive rearrangement of specific host cell organelle membranes that serve as the sites of viral replication. The tombusvirus Tomato bushy stunt virus (TBSV) replicates within spherules derived from the peroxisomal boundary membrane, a process that involves the coordinated action of various viral and cellular factors, including constituents of the endosomal sorting complex required for transport (ESCRT). ESCRT is comprised of a series of protein subcomplexes (i.e., ESCRT-0 -I, -II, and -III) that normally participate in late endosome biogenesis and some of which are also hijacked by certain enveloped retroviruses (e.g., HIV) for viral budding from the plasma membrane. Here we show that the replication of Carnation Italian ringspot virus (CIRV), a tombusvirus that replicates at mitochondrial membranes also relies on ESCRT. In plant cells, CIRV recruits the ESCRT-I protein, Vps23, to mitochondria through an interaction that involves a unique region in the N terminus of the p36 replicase-associated protein that is not conserved in TBSV or other peroxisome-targeted tombusviruses. The interaction between p36 and Vps23 also involves the Vps23 C-terminal steadiness box domain and not its N-terminal ubiquitin E2 variant domain, which in the case of TBSV (and enveloped retroviruses) mediates the interaction with ESCRT. Overall, these results provide evidence that CIRV uses a unique N-terminal sequence for the recruitment of Vps23 that is distinct from those used by TBSV and certain mammalian viruses for ESCRT recruitment. Characterization of this novel interaction with Vps23 contributes to our understanding of how CIRV may have evolved to exploit key differences in the plant ESCRT machinery. IMPORTANCEPositive-strand RNA viruses replicate their genomes in association with specific host cell membranes. To accomplish this, cellular components responsible for membrane biogenesis and modeling are appropriated by viral proteins and redirected to assemble membrane-bound viral replicase complexes. The diverse pathways leading to the formation of these replication structures are poorly understood. We have determined that the cellular ESCRT system that is normally responsible for mediating late endosome biogenesis is also involved in the replication of the tombusvirus Carnation Italian ringspot virus (CIRV) at mitochondria. Notably, CIRV recruits ESCRT to the mitochondrial outer membrane via an interaction between a unique motif in the viral protein p36 and the ESCRT component Vps23. Our findings provide new insights into tombusvirus replication and the virusinduced remodeling of plant intracellular membranes, as well as normal ESCRT assembly in plants.T ombusviruses are positive-strand RNA [(ϩ)RNA] viruses that infect a wide range of plant species and replicate at host cell membranes derived specifically from either peroxisomes (e.g., Tomato bushy stunt virus [TBSV]) or mitochondria (e.g., Carnation Italian ringspot virus [CIRV]) (1). Up...

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BY‐2 Cells: Culture and Transformation for Live Cell Imaging

Cited by 48 publications

References 32 publications

Lipid droplet-associated proteins (LDAPs) are involved in the compartmentalization of lipophilic compounds in plant cells

Lipid droplet-associated proteins (LDAPs) are involved in the compartmentalization of lipophilic compounds in plant cells

Plants, like mammals, but unlikeSaccharomyces,do not regulate nuclear-cytoplasmic tRNA trafficking in response to nutrient stress

A Unique N-Terminal Sequence in the Carnation Italian ringspot virus p36 Replicase-Associated Protein Interacts with the Host Cell ESCRT-I Component Vps23

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