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BioAssay: AID 686937

Genes involved in protein secretion and Golgi organization

Yeast genetics and in vitro biochemical analysis have identified numerous genes involved in protein secretion. As compared with yeast, however, the metazoan secretory pathway is more complex and many mechanisms that regulate organization of the Golgi apparatus remain poorly characterized. We performed a genome-wide RNA-mediated interference screen in a Drosophila cell line to identify genes more ..
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 Tested RNAi Reagents
 Tested RNAi Reagents
All(22533)
 
 
Active(1068)
 
 
Inactive(21465)
 
 
 Screened Genes
 Screened Genes
AID: 686937
Data Source: Drosophila RNAi Screening Center (DRSC) (DRSC-P34)
BioAssay Type: Primary, Primary Screening, Single Concentration Activity Observed
Depositor Category: Assay Vendor
Deposit Date: 2013-04-13

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Description:
Title: Functional genomics reveals genes involved in protein secretion and Golgi organization

Abstract:
Yeast genetics and in vitro biochemical analysis have identified numerous genes involved in protein secretion. As compared with yeast, however, the metazoan secretory pathway is more complex and many mechanisms that regulate organization of the Golgi apparatus remain poorly characterized. We performed a genome-wide RNA-mediated interference screen in a Drosophila cell line to identify genes required for constitutive protein secretion. We then classified the genes on the basis of the effect of their depletion on organization of the Golgi membranes. Here we show that depletion of class A genes redistributes Golgi membranes into the endoplasmic reticulum, depletion of class B genes leads to Golgi fragmentation, depletion of class C genes leads to aggregation of Golgi membranes, and depletion of class D genes causes no obvious change. Of the 20 new gene products characterized so far, several localize to the Golgi membranes and the endoplasmic reticulum.
Protocol
The primary assay used a genome-wide library of ~22,000 double-stranded RNAs (dsRNAs) that have been previously used in several screens. Cells secreting ss-HRP were plated in 384-well plates containing dsRNA. After 5 days, ss-HRP production was induced, and 12 hours later peroxidase activity released into the supernatant was measured by chemiluminescence. Each plate included wells with dsRNA encoding Syntaxin 5 and GFP as positive and negative controls. The screen was carried out in duplicate. Because most dsRNAs did not inhibit HRP secretion, the average for a given plate was very close to that of non-treated wells. Therefore, the z-score of peroxidase activity in each well was used to compare the effects of each dsRNA on secretion across the whole set of plates.
Comment
The activity scores are given using the standard DRSC scoring system for hits:
1 = weak hit
2 = medium hit
3 = strong hit
In many assays, only 2 is used.
Result Definitions
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TIDNameDescriptionHistogramTypeUnit
OutcomeThe BioAssay activity outcomeOutcome
ScoreThe BioAssay activity ranking scoreInteger
1extHRP replicate 1Float
2extHRP replicate 2Float
3extHRP replicate 3Float
4extHRP replicate 4Float
5extHRP replicate 5Float
6extHRP replicate 6Float
7intHRP replicate 1Float
8intHRP replicate 2Float
9intHRP replicate 3Float
10intHRP replicate 4Float
11z-score replicate 1Float
12z-score replicate 2Float
13z-score replicate 3Float
14z-score replicate 4Float
15z-score replicate 5Float
16z-score replicate 6Float
17Phenotype replicate 1String
18Phenotype replicate 2String
19Phenotype replicate 3String
20Phenotype replicate 4String
21Predicted Target GeneString

RNAi Target.
Additional Information
Substance Type: Nucleotide

Data Table (Concise)
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