MAPK signaling to the early secretory pathway revealed by kinase/phosphatase functional screening

Farhan, Hesso; Wendeler, Markus W.; Mitrović, Sandra; Fava, Eugenio; Silberberg, Yael; Sharan, Roded; Zerial, Marino; Hauri, Hans‐Peter

doi:10.1083/jcb.200912082

Cited by 174 publications

(226 citation statements)

References 51 publications

(73 reference statements)

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“…For example, signaling by growth factors (e.g. MAPK/ERK) at plasma membranes affects the early secretory pathway (anterograde transport) via the ERES (Farhan et al, 2010). Thus, it is important to investigate the overall balance of membrane trafficking between the relevant organelles, as well as the plasma membrane, to elucidate the changes in Golgi and ER morphology that occur during mitosis more fully.…”

Section: Discussionmentioning

confidence: 99%

Semi-Intact Cell Systems - Application to the Analysis of Membrane Trafficking Between the Endoplasmic Reticulum and the Golgi Apparatus and of Cell Cycle-Dependent Changes in the Morphology of These Organelles

Murata¹,

Kano²

2012

Crosstalk and Integration of Membrane Trafficking Pathways

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Section: Discussionmentioning

confidence: 99%

Semi-Intact Cell Systems - Application to the Analysis of Membrane Trafficking Between the Endoplasmic Reticulum and the Golgi Apparatus and of Cell Cycle-Dependent Changes in the Morphology of These Organelles

Murata¹,

Kano²

2012

Crosstalk and Integration of Membrane Trafficking Pathways

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“…Farhan and colleagues investigated the role of human kinases and phosphatases in the distribution of membrane markers of the early secretory pathway, and showed that growth factor signalling regulates ER export through ERK2. Besides the MAPK signalling pathway, a number of other signalling networks, such as those involving insulin, phosphatidylinositol or toll-like receptor signalling, were also found to have a role in the regulation of the early secretory pathway (Farhan et al, 2010).…”

Section: Stedmentioning

confidence: 99%

“…Such an automated, integrated approach has recently been used to identify, at genome level, those genes that, when knocked down, cause alterations in biosynthetic protein trafficking (Simpson et al, 2012) and organelle integrity (Farhan et al, 2010;Kondylis et al, 2011;Simpson et al, 2012). Farhan and colleagues investigated the role of human kinases and phosphatases in the distribution of membrane markers of the early secretory pathway, and showed that growth factor signalling regulates ER export through ERK2.…”

Section: Stedmentioning

confidence: 99%

“…The involvement of a gene in the early secretory pathway can also be investigated in intact cells, by knocking it down using RNA interference (RNAi) and analysing the effect on trafficking and/or the morphology of the secretory pathway components. A number of such RNAi-based screens have been performed in metazoans (Bard et al, 2006;D'Ambrosio and Vale, 2010;Farhan et al, 2010;Kondylis et al, 2011;Simpson et al, 2012;Wendler et al, 2010). For example, two genome-wide screens identified new genes with a role in protein secretion by first analysing the effects of gene knockdown on protein secretion by using enzymatic bulk readout analysis, followed by systematic microscopy-based analyses to better understand the role of the identified genes in regulating the early secretory pathway or Golgi integrity (Bard et al, 2006;Wendler et al, 2010).…”

Section: Stedmentioning

confidence: 99%

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Imaging ER-to-Golgi transport: towards a systems view

Verı́ssimo

Pepperkok

2013

Journal of Cell Science

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SummaryProteins synthesised at the endoplasmic reticulum (ER) have to undergo a number of consecutive and coordinated steps to reach the Golgi complex. To understand the dynamic complexity of ER-to-Golgi transport at the structural and molecular level, light microscopy approaches are fundamental tools that allow in vivo observations of protein dynamics and interactions of fluorescent proteins in living cells. Imaging protein and organelle dynamics close to the ultra-structural level became possible by combining light microscopy with electron microscopy analyses or super-resolution light microscopy methods. Besides, increasing evidence suggests that the early secretory pathway is tightly connected to other cellular processes, such as signal transduction, and quantitative information at the systems level is fundamental to achieve a comprehensive molecular understanding of these connections. High-throughput microscopy in fixed and living cells in combination with systematic perturbation of gene expression by, e.g. RNA interference, will open new avenues to gain such an understanding of the early secretory pathway at the systems level. In this Commentary, we first outline examples that revealed the dynamic organisation of ER-to-Golgi transport in living cells. Next, we discuss the use of advanced imaging methods in studying ER-to-Golgi transport and, finally, delineate the efforts in understanding ER-to-Golgi transport at the systems level.

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“…Treatment with chemical compounds or specific gene silencing by RNA interference are commonly used at the scale of individual experiments to highthroughput studies. Visual inspection by expert biologists has been performed for several decades, ranging from early studies by microscopists like Ramon y Cajal to contemporary large scale, high-throughput screens (1)(2)(3)(4). Although human observation may be very accurate, the three major drawbacks are that (i) it lacks quantitative measures, (ii) it may be biased, and (iii) it is time consuming.…”

mentioning

confidence: 99%

Closed-form density-based framework for automatic detection of cellular morphology changes

Duong

Goud

Schauer

2012

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

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A primary method for studying cellular function is to examine cell morphology after a given manipulation. Fluorescent markers attached to proteins/intracellular structures of interest in conjunction with 3D fluorescent microscopy are frequently exploited for functional analysis. Despite the central role of morphology comparisons in cell biological approaches, few statistical tools are available that allow biological scientists without a high level of statistical training to quantify the similarity or difference of fluorescent images containing multifactorial information. We transform intracellular structures into kernels and develop a multivariate two-sample test that is nonparametric and asymptotically normal to directly and quantitatively compare cellular morphologies. The asymptotic normality bypasses the computationally intensive calculations used by the usual resampling techniques to compute the P-value. Because all parameters required for the statistical test are estimated directly from the data, it does not require any subjective decisions. Thus, we provide a black-box method for unbiased, automated comparison of cell morphology. We validate the performance of our test statistic for finite synthetic samples and experimental data. Employing our test for the comparison of the morphology of intracellular multivesicular bodies, we detect changes in their distribution after disruption of the cellular microtubule cytoskeleton with high statistical significance in fixed samples and live cell analysis. These results demonstrate that density-based comparison of multivariate image information is a powerful tool for automated detection of cell morphology changes. Moreover, the underlying mathematics of our test statistic is a general technique, which can be applied in situations where two data samples are compared.hypothesis test | integrated density functional | optimal bandwidth selection | quantitative cell comparison F luorescent markers attached to proteins of interest in conjunction with modern fluorescent microscopy technologies are a useful proxy for studying subcellular compartments and their behavior after a given manipulation. Treatment with chemical compounds or specific gene silencing by RNA interference are commonly used at the scale of individual experiments to highthroughput studies. Visual inspection by expert biologists has been performed for several decades, ranging from early studies by microscopists like Ramon y Cajal to contemporary large scale, high-throughput screens (1-4). Although human observation may be very accurate, the three major drawbacks are that (i) it lacks quantitative measures, (ii) it may be biased, and (iii) it is time consuming.The structural features of cells and the topological relationships between the numerous intracellular compartments give rise to multivariate data whose unbiased, automatic comparison is a major challenge. Importantly, alterations in cellular morphology also occur in many diseases, including cancer, requiring quantitative tools for their detection. Give...

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MAPK signaling to the early secretory pathway revealed by kinase/phosphatase functional screening

Abstract: An RNAi screen determines that the early secretory pathway is subject to phosphoregulation via a variety of signaling pathways, including a link between growth factor signaling and ER export.

Cited by 174 publications

References 51 publications

Semi-Intact Cell Systems - Application to the Analysis of Membrane Trafficking Between the Endoplasmic Reticulum and the Golgi Apparatus and of Cell Cycle-Dependent Changes in the Morphology of These Organelles

Semi-Intact Cell Systems - Application to the Analysis of Membrane Trafficking Between the Endoplasmic Reticulum and the Golgi Apparatus and of Cell Cycle-Dependent Changes in the Morphology of These Organelles

Imaging ER-to-Golgi transport: towards a systems view

Closed-form density-based framework for automatic detection of cellular morphology changes

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