sirna screen
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2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Micha Müller ◽  
Merve Avar ◽  
Daniel Heinzer ◽  
Marc Emmenegger ◽  
Adriano Aguzzi ◽  
...  
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2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Micha Müller ◽  
Lucas Pelkmans ◽  
Scott Berry

AbstractCoordination of RNA abundance and production rate with cell size has been observed in diverse organisms and cell populations. However, how cells achieve such ‘scaling’ of transcription with size is unknown. Here we describe a genome-wide siRNA screen to identify regulators of global RNA production rates in HeLa cells. We quantify the single-cell RNA production rate using metabolic pulse-labelling of RNA and subsequent high-content imaging. Our quantitative, single-cell measurements of DNA, nascent RNA, proliferating cell nuclear antigen (PCNA), and total protein, as well as cell morphology and population-context, capture a detailed cellular phenotype. This allows us to account for changes in cell size and cell-cycle distribution (G1/S/G2) in perturbation conditions, which indirectly affect global RNA production. We also take advantage of the subcellular information to distinguish between nascent RNA localised in the nucleolus and nucleoplasm, to approximate ribosomal and non-ribosomal RNA contributions to perturbation phenotypes. Perturbations uncovered through this screen provide a resource for exploring the mechanisms of regulation of global RNA metabolism and its coordination with cellular states.


2020 ◽  
Vol 11 (11) ◽  
Author(s):  
Ayse Ertay ◽  
Huiquan Liu ◽  
Dian Liu ◽  
Ping Peng ◽  
Charlotte Hill ◽  
...  

AbstractTriple-negative breast cancer (TNBC) is the most aggressive type of breast cancer that lacks the oestrogen receptor, progesterone receptor and human epidermal growth factor receptor 2, making it difficult to target therapeutically. Targeting synthetic lethality is an alternative approach for cancer treatment. TNBC shows frequent loss of phosphatase and tensin homologue (PTEN) expression, which is associated with poor prognosis and treatment response. To identify PTEN synthetic lethal interactions, TCGA analysis coupled with a whole-genome siRNA screen in isogenic PTEN-negative and -positive cells were performed. Among the candidate genes essential for the survival of PTEN-inactive TNBC cells, WDHD1 (WD repeat and high-mobility group box DNA-binding protein 1) expression was increased in the low vs. high PTEN TNBC samples. It was also the top hit in the siRNA screen and its knockdown significantly inhibited cell viability in PTEN-negative cells, which was further validated in 2D and 3D cultures. Mechanistically, WDHD1 is important to mediate a high demand of protein translation in PTEN-inactive TNBC. Finally, the importance of WDHD1 in TNBC was confirmed in patient samples obtained from the TCGA and tissue microarrays with clinic-pathological information. Taken together, as an essential gene for the survival of PTEN-inactive TNBC cells, WDHD1 could be a potential biomarker or a therapeutic target for TNBC.


PLoS ONE ◽  
2020 ◽  
Vol 15 (10) ◽  
pp. e0240746
Author(s):  
Lauren E. King ◽  
Hui-Hua Zhang ◽  
Cathryn M. Gould ◽  
Daniel W. Thomas ◽  
Lachlan W. Whitehead ◽  
...  

2020 ◽  
Vol 295 (23) ◽  
pp. 7941-7957 ◽  
Author(s):  
Shiho Torii ◽  
Yasuko Orba ◽  
Michihito Sasaki ◽  
Koshiro Tabata ◽  
Yuji Wada ◽  
...  

Chikungunya fever is a re-emerging zoonotic disease caused by chikungunya virus (CHIKV), a member of the Alphavirus genus in the Togaviridae family. Only a few studies have reported on the host factors required for intracellular CHIKV trafficking. Here, we conducted an imaging-based siRNA screen to identify human host factors for intracellular trafficking that are involved in CHIKV infection, examined their interactions with CHIKV proteins, and investigated the contributions of these proteins to CHIKV infection. The results of the siRNA screen revealed that host endosomal sorting complexes required for transport (ESCRT) proteins are recruited during CHIKV infection. Co-immunoprecipitation analyses revealed that both structural and nonstructural CHIKV proteins interact with hepatocyte growth factor–regulated tyrosine kinase substrate (HGS), a component of the ESCRT-0 complex. We also observed that HGS co-localizes with the E2 protein of CHIKV and with dsRNA, a marker of the replicated CHIKV genome. Results from gene knockdown analyses indicated that, along with other ESCRT factors, HGS facilitates both genome replication and post-translational steps during CHIKV infection. Moreover, we show that ESCRT factors are also required for infections with other alphaviruses. We conclude that during CHIKV infection, several ESCRT factors are recruited via HGS and are involved in viral genome replication and post-translational processing of viral proteins.


2020 ◽  
Vol 34 (S1) ◽  
pp. 1-1
Author(s):  
Anand Patwardhan ◽  
Norton Cheng ◽  
JoAnn Trejo

2020 ◽  
Vol 7 (1) ◽  
Author(s):  
Beth K. Neilsen ◽  
David L. Kelly ◽  
Binita Chakraborty ◽  
Hyun Seok Kim ◽  
Michael A. White ◽  
...  

2020 ◽  
Vol 16 (1) ◽  
pp. e1008268 ◽  
Author(s):  
Nenavath Gopal Naik ◽  
Thomas Hong Nguyen ◽  
Lauren Roberts ◽  
Luke Todd Fischer ◽  
Katherine Glickman ◽  
...  

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Gerald Klanert ◽  
Daniel J. Fernandez ◽  
Marcus Weinguny ◽  
Peter Eisenhut ◽  
Eugen Bühler ◽  
...  

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