CB 399: RNAi Screening: From Design to Data Analysis Spring 2011

Intellectual Unit:

RNAi Screening: From Design to Data Analysis
Course Director: Norbert Perrimon
Course Lecturers: Steve Elledge, Stephanie Mohr, Norbert Perrimon and Caroline Shamu

Curriculum Fellow: Leah Brault, lbrault@genetics.med.harvard.edu

Tissue culture cells have provided a powerful system for studying many fundamental problems in signal transduction, cell differentiation and physiology. However, functional studies in cultured cells were hampered in the past by the lack of a powerful method for perturbing gene activities. A turning point came with the discovery of RNA interference and its rapid rise from small scale to genome-scale screening. Today, the most commonly used approaches are based on long dsRNA for Drosophila cells, and either synthetic siRNAs or vector-expressed short hairpin RNAs (shRNAs) for mammalian cells. Driven by genome-sequence data, RNAi is now widely used in high-throughput (HT) screens in both basic and applied biology. It is a powerful method for addressing many cell biological questions, and its amenability for use in modifier screens in addition to direct loss of function screening has made it particularly useful for the analysis of signal transductions pathways. RNAi has also become a method of choice for key steps in the development of therapeutic agents, from target discovery and validation to the analysis of the mechanisms of action of small molecules. This nanocourse will focus on both the technology and applications of RNAi screens.

 

Schedule:

First Session: Tuesday, February 8, 2011, 1 - 4:30 PM
Location: TMEC Building, Room 209

Second Session: Monday, February 14, 2011, 2- 4:30 PM
Location: TMEC Building, Room 333

 

Recommended Reading

Boutros M, and Ahringer J. 2008.  The art and design of genetic screens: RNA interference. Nat Rev Genet  9(7):554-66.

Echeverri C and Perrimon N. 2006.  High-throughput RNAi Screening in cells: A User's Guide. Nat Rev Genet  7(5):373-84.

Friedman, A & Perrimon, N., 2006.  A functional genomic RNAi screen for novel regulators of RTK/ERK signaling. Nature  444(7116):230-4.

Schlabach MR, et al.  2008.  Cancer proliferation gene discovery through functional genomics. Science 319(5863):620-4.

Brass AL, Huang I-C, Benita Y, John SP, Krishnan MN, Feeley EM, Ryan BJ, Weyer JL, van der Weyden L, Fikrig E, Adams DJ, Xavier RJ, Farzan M, Elledge SJ.  2009.  The IFITM Proteins Mediate Cellular Resistance to Influenza A H1N1 Virus, West Nile Virus, and Dengue Virus. Cell 139:1243-1254.

Birmingham A, Selfors LM, Forster T, Wrobel D, Kennedy CJ, Shanks E, Santoyo-Lopez J, Dunicann DJ, Long A, Kelleher D, Smith Q, Beijersbergen RL, Ghazal P, Shamu CE. 2009. Statistical methods for analysis of high-throughput RNA interference screens. Nat Methods. 2009 Aug;6(8):569-75.

Mohr S, Bakal C, Perrimon N.  2010.  Genomic screening with RNAi: results and challenges.  Annu Rev Biochem  79:37-64.

Neumann B, Walter T, Hériché JK, Bulkescher J, Erfle H, Conrad C, Rogers P, Poser I, Held M, Liebel U, Cetin C, Sieckmann F, Pau G, Kabbe R, Wünsche A, Satagopam V, Schmitz MH, Chapuis C, Gerlich DW, Schneider R, Eils R, Huber W, Peters JM, Hyman AA, Durbin R, Pepperkok R, Ellenberg J.  2010.  Phenotypic profiling of the human genome by time-lapse microscopy reveals cell division genes.  Nature  464(7289):721-7.

Collinet C, Stöter M, Bradshaw CR, Samusik N, Rink JC, Kenski D, Habermann B, Buchholz F, Henschel R, Mueller MS, Nagel WE, Fava E, Kalaidzidis Y, Zerial M.  2010.  Systems survey of endocytosis by multiparametric image analysis.  Nature  464(7286):243-9.

 

DROP DEADLINE: Tuesday, February 1, 2011

 

AUDITORS (Post-Docs, Faculty, or Staff) DO NOT NEED TO SIGN UP TO ATTEND THE 1st SESSION.  PLEASE DO NOT ENROLL.

 

Instructions for Registered Students

 The first meeting of the "RNAi Screening: From Design to Data Analysis" nanocourse will take place on Tuesday, February 8th from 1:00 - 4:30 p.m. Reference material and the course assignment are currently posted below. It is up to the individual student to become familiar and comfortable with the terms and definitions needed to understand the primary literature and the lectures. The extent of each student's preparation will depend on that student's prior knowledge of the subject matter. Following the first meeting of this nanocourse, the lecture material will be placed online.

 

The second meeting of this course will be on Monday, February 14th from 2:00 – 4:30 p.m. Please arrive on time for this session!  In preparation for this session, selected publications from the recommended readings list above will be required for you to read.  These are posted as .pdfs below.  The required readings are:

Friedman, A & Perrimon, N., 2006.  A functional genomic RNAi screen for novel regulators of RTK/ERK signaling. Nature  444(7116):230-4.

Schlabach MR, et al.  2008.  Cancer proliferation gene discovery through functional genomics. Science 319(5863):620-4.

Brass AL, Huang I-C, Benita Y, John SP, Krishnan MN, Feeley EM, Ryan BJ, Weyer JL, van der Weyden L, Fikrig E, Adams DJ, Xavier RJ, Farzan M, Elledge SJ.  2009.  The IFITM Proteins Mediate Cellular Resistance to Influenza A H1N1 Virus, West Nile Virus, and Dengue Virus. Cell 139:1243-1254.

 

All students are required to submit a 1-page description of an RNAi screen which addresses a specific biological question. (“Specific” is to be taken literally…studying “cell death”, “cell viability” or “effects on antennae” are too broad.  Be exact in your question!)  In your answer you should describe the primary screening assay, at least one secondary assay, appropriate controls, data analysis and validation strategies. Note that you are permitted to describe an assay that is related to your thesis work but you should not recycle a previous assignment or specific aims from your thesis proposal. These assignments will be the basis for discussion on the 14th, so work hard on them. You should also be ready to ask questions and give constructive feedback to fellow students on Monday.

This assignment must be submitted to the curriculum fellow (lbrault@genetics.med.harvard.edu) by Friday, February 11th at 12:00 p.m. (noon).

Immediately following the discussion session, you will be given the opportunity to tour the ICCB-Longwood screening facility and the Drosophila RNAi Screening Center. Participation in the tour is not required to receive nanocourse credit.