Agrobacterium tumefaciens-Mediated Transient Transformation of Arabidopsis thaliana Leaves

Mangano, Silvina; González, Cintia Daniela; Petruccelli, Silvana

doi:10.1007/978-1-62703-580-4_8

Cited by 28 publications

(19 citation statements)

References 16 publications

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“…Bacterial cultures were adjusted to an optical density at 600 nm of 0.5 using infiltration buffer, and N. benthamiana leaves were pressure infiltrated through the abaxial epidermis. A. tumefaciensmediated transient transformation of Arabidopsis was performed in leaves of 4-week-old plants grown under short-day conditions according to the protocol described by Mangano et al (2014). For stable transformation of Arabidopsis, the floral dip technique was used (Clough and Bent, 1998), and eight to 24 independent transgenic lines were identified in the T1 generation by Basta selection for each construct analyzed.…”

Section: Plant Transformationmentioning

confidence: 99%

The IQD Family of Calmodulin-Binding Proteins Links Calcium Signaling to Microtubules, Membrane Subdomains, and the Nucleus

et al. 2017

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ORCID IDs: 0000-0002-3493-4800 (K.B.); 0000-0002-7146-043X (B.M.).Calcium (Ca 2+ ) signaling and dynamic reorganization of the cytoskeleton are essential processes for the coordination and control of plant cell shape and cell growth. Calmodulin (CaM) and closely related calmodulin-like (CML) polypeptides are principal sensors of Ca 2+ signals. CaM/CMLs decode and relay information encrypted by the second messenger via differential interactions with a wide spectrum of targets to modulate their diverse biochemical activities. The plant-specific IQ67 DOMAIN (IQD) family emerged as possibly the largest class of CaM-interacting proteins with undefined molecular functions and biological roles. Here, we show that the 33 members of the IQD family in Arabidopsis (Arabidopsis thaliana) differentially localize, using green fluorescent protein (GFP)-tagged proteins, to multiple and distinct subcellular sites, including microtubule (MT) arrays, plasma membrane subdomains, and nuclear compartments. Intriguingly, the various IQD-specific localization patterns coincide with the subcellular patterns of IQD-dependent recruitment of CaM, suggesting that the diverse IQD members sequester Ca 2+ -CaM signaling modules to specific subcellular sites for precise regulation of Ca 2+ -dependent processes. Because MT localization is a hallmark of most IQD family members, we quantitatively analyzed GFP-labeled MT arrays in Nicotiana benthamiana cells transiently expressing GFP-IQD fusions and observed IQD-specific MT patterns, which point to a role of IQDs in MT organization and dynamics. Indeed, stable overexpression of select IQD proteins in Arabidopsis altered cellular MT orientation, cell shape, and organ morphology. Because IQDs share biochemical properties with scaffold proteins, we propose that IQD families provide an assortment of platform proteins for integrating CaM-dependent Ca 2+ signaling at multiple cellular sites to regulate cell function, shape, and growth.

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Section: Plant Transformationmentioning

confidence: 99%

The IQD Family of Calmodulin-Binding Proteins Links Calcium Signaling to Microtubules, Membrane Subdomains, and the Nucleus

et al. 2017

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“…As is true for stable transformation, it is now routine to add AS during transformation. Pre-treating A. tumefaciens with AS before agroinfiltration of in A. thaliana leaves can also increase the transient transformation efficiency (Mangano et al, 2014). Detergents/surfactants are also used to improve cuticular penetration of fluid on leaf surfaces.…”

Section: Transient Transformation In Adult Leaves Via Agrofiltrationmentioning

confidence: 99%

Agrobacterium-Mediated Plant Transformation: Biology and Applications

Hwang

Lai

2017

The Arabidopsis Book

216

142

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“…The leaves of 4- to 6-week old N. benthamiana were co-infiltrated with Agrobacterium tumefaciens GV3101 strains harboring either a YFP-fused protein or an mCer3-fused protein, in addition to the Agrobacterium strain C58C1 harboring the viral silencing suppressor helper complex pCH32 (109). Before infiltration into Nicotiana leaves, Agrobacterium cultures grown overnight were treated with 200uM acetosyringone in a modified induction buffer for 4 hours (110). This induced culture was re-suspended in 10mM MES 10mM MgSO4 and directly infiltrated into young leaves.…”

Section: Methodsmentioning

confidence: 99%

NUCLEAR FACTOR Y, subunit C (NF-YC) transcription factors are positive regulators of photomorphogenesis inArabidopsis thaliana

Myers

Kumimoto

Siriwardana

et al. 2016

Preprint

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Recent reports suggested that NF-Y transcription factors are positive regulators of skotomorphogenesis in Arabidopsis thaliana. Three NF-YC genes (NF-YC3, NF-YC4, and NF-YC9) are known to have overlapping functions in photoperiod dependent flowering and previous studies demonstrated that they interact with basic leucine zipper (bZIP) transcription factors. This included ELONGATED HYPOCOTYL 5 (HY5), which has well-demonstrated roles in photomorphogenesis. Similar to hy5 mutants, we report that nf-yc3 nf-yc4 nf-yc9 triple mutants failed to inhibit hypocotyl elongation in all tested light wavelengths. Surprisingly, nf-yc3 nf-yc4 nf-yc9 hy5 mutants had synergistic defects in light perception, suggesting that NF-Ys represent a parallel light signaling pathway. As with other photomorphogenic transcription factors, nf-yc3 nf-yc4 nf-yc9 triple mutants also partially suppress the short hypocotyl and dwarf rosette phenotypes of CONSTITUTIVE PHOTOMORPHOGENIC 1 (cop1) mutants. Thus, our data strongly suggest that NF-Y transcription factors have important roles as positive regulators of photomorphogenesis, and in conjunction with other recent reports implies that the NF-Y are multifaceted regulators of early seedling development.Author SummaryLight perception is critically important for the fitness of plants in both natural and agricultural settings. Plants not only use light for photosynthesis, but also as a cue for proper development. As a seedling emerges from soil it must determine the light environment and adopt an appropriate growth habit. When blue and red wavelengths are the dominant sources of light, plants will undergo photomorphogenesis. Photomorphogenesis describes a number of developmental responses initiated by light in a seedling, and includes shortened stems and establishing the ability to photosynthesize. The genes regulating photomorphogenesis have been studied extensively, but a complete picture remains elusive. Here we describe the finding that NUCLEAR FACTOR-Y (NF-Y) genes are positive regulators of photomorphogenesis - i.e., in plants where NF-Y genes are mutated, they display some characteristics of dark grown plants, even though they are in the light. Our data suggests that the roles of NF-Y genes in light perception do not fit in easily with those of other described pathways. Thus, studying these genes promises to help develop a more complete picture of how light drives plant development.

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Agrobacterium tumefaciens-Mediated Transient Transformation of Arabidopsis thaliana Leaves

Cited by 28 publications

References 16 publications

The IQD Family of Calmodulin-Binding Proteins Links Calcium Signaling to Microtubules, Membrane Subdomains, and the Nucleus

The IQD Family of Calmodulin-Binding Proteins Links Calcium Signaling to Microtubules, Membrane Subdomains, and the Nucleus

Agrobacterium-Mediated Plant Transformation: Biology and Applications

NUCLEAR FACTOR Y, subunit C (NF-YC) transcription factors are positive regulators of photomorphogenesis inArabidopsis thaliana

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