Efficient control of gene expression by a tetracycline-dependent transactivator in single Dictyostelium discoideum cells

Blaauw, Mieke; Linskens, Maarten H.K.; Haastert, Peter J.M. van

doi:10.1016/s0378-1119(00)00227-4

Cited by 57 publications

(56 citation statements)

References 33 publications

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“…The resulting constructs were designated T1, T2, T3, T4, R1, R2, and R7, respectively. Each of the constructs was subcloned into the MB38 vector for tetracycline-regulated expression in Dictyostelium (50).…”

Section: Methodsmentioning

confidence: 99%

Two modes of microtubule sliding driven by cytoplasmic dynein

Shima

Kon

Imamula

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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Dynein is a huge multisubunit microtubule (MT)-based motor, whose motor domain resides in the heavy chain. The heavy chain comprises a ring of six AAA (ATPases associated with diverse cellular activities) modules with two slender protruding domains, the tail and stalk. It has been proposed that during the ATP hydrolysis cycle, this tail domain swings against the AAA ring as a lever arm to generate the power stroke. However, there is currently no direct evidence to support the model that the tail swing is tightly linked to dynein motility. To address the question of whether the power stroke of the tail drives MT sliding, we devised an in vitro motility assay using genetically biotinylated cytoplasmic dyneins anchored on a glass surface in the desired orientation with a biotin-streptavidin linkage. Assays on the dyneins with the site-directed biotin tag at eight different locations provided evidence that robust MT sliding is driven by the power stroke of the tail. Furthermore, the assays revealed slow MT sliding independent of dynein orientation on the glass surface, which is mechanically distinct from the sliding driven by the power stroke of the tail.AAA protein ͉ motility ͉ motor protein ͉ power stroke

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Section: Methodsmentioning

confidence: 99%

Two modes of microtubule sliding driven by cytoplasmic dynein

Shima

Kon

Imamula

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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“…For RNA interference (RNAi) experiments, cells were cotransformed with a raptor (Tet-off) expression vector and a vector that encodes the tetracycline-dependent transrepressor (Blaauw et al, 2000). Cells grown in the presence of tetracycline did not express the engineered raptor RNAi; however, upon removal of tetracycline, raptor RNAi was induced and, with time, endogenous raptor mRNA was depleted ( Fig.…”

Section: Phagocytosis Does Not Correlate With Torc1 Activitymentioning

confidence: 99%

TOR complex 2 (TORC2) in Dictyostelium suppresses phagocytic nutrient capture independently of TORC1-mediated nutrient sensing

Rösel

Khurana

Majithia

et al. 2012

Journal of Cell Science

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SummaryThe TOR protein kinase functions in two distinct complexes, TOR complex 1 (TORC1) and 2 (TORC2). TORC1 is required for growth in response to growth factors, nutrients and the cellular energy state; TORC2 regulates AKT signaling, which can modulate cytoskeletal polarization. In its ecological niche, Dictyostelium engulf bacteria and yeast for nutrient capture. Despite the essential role of TORC1 in control of cellular growth, we show that nutrient particle capture (phagocytosis) in Dictyostelium is independent of TORC1-mediated nutrient sensing and growth regulation. However, loss of Dictyostelium TORC2 components Rictor/Pia, SIN1/RIP3 and Lst8 promotes nutrient particle uptake; inactivation of TORC2 leads to increased efficiency and speed of phagocytosis. In contrast to phagocytosis, we show that macropinocytosis, an AKT-dependent process for cellular uptake of fluid phase nutrients, is not regulated by either of the TOR complexes. The integrated and balanced regulation of TORC1 and TORC2 might be crucial in Dictyostelium to coordinate growth and energy needs with other essential TOR-regulated processes.

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“…In such a case, a target gene's activity can be inhibited by using an antisense or a dominant negative construct under the control of a regulatable promoter, e.g., the tetracycline ''off'' system (Blaauw et al 2000). More sophisticated manipulations such as gene ''knock-ins'' are also routinely used to modify genes in the genome.…”

Section: Dictyostelium Genomics and Molecular Geneticsmentioning

confidence: 99%

Dictyostelium Finds New Roles to Model

Williams

2010

Genetics

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Any established or aspiring model organism must justify itself using two criteria: does the model organism offer experimental advantages not offered by competing systems? And will any discoveries made using the model be of wider relevance? This review addresses these issues for the social amoeba Dictyostelium and highlights some of the organisms more recent applications. These cover a remarkably wide gamut, ranging from sociobiological to medical research with much else in between.

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Efficient control of gene expression by a tetracycline-dependent transactivator in single Dictyostelium discoideum cells

Cited by 57 publications

References 33 publications

Two modes of microtubule sliding driven by cytoplasmic dynein

Two modes of microtubule sliding driven by cytoplasmic dynein

TOR complex 2 (TORC2) in Dictyostelium suppresses phagocytic nutrient capture independently of TORC1-mediated nutrient sensing

Dictyostelium Finds New Roles to Model

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