RNA Interference

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RNA Interference
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Introduction

• This phenomenon was fully understood first in C.
  elegans.
• si RNA and mi RNA
• ds RNAs are acted up by Dicer and RISC
• Primed and unprimed ds RNAs
• Result in Silencing

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Small Interfering RNA-Mediated Gene Silencing in
T Lymphocytes

• Si RNA-mediated RNAi mechanism using a
  thymoma-derived cell line model to demonstrate
  for the first time the occurrence of RNAi in
  primary T-lymphocytes.

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Materials and Methods

• Cell Culture E10
• Transfection
• Flowcytometry
• Northern Hybridization

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Results
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Activation of the interferon system by
short-interfering RNAs

• The authors found that transfection of siRNA
  results in interferon (INF)-mediated activation
  of the jak-Stat pathway and global upregulation
  of IFN-stimulated genes.
• This effect is mediated by the dsRNA-dependent
  protein kinase, PKR, which is activated by
  21-base-pair (bp) siRNAs and required for
  upregulation of IFN-beta in response to siRNA.

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Materials and Methods

• Cell Cultures-S2 Drosophila Schneider cells
• Vector construction
• Protein analysis
• Semi-quantative RT-PCR analysis

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Discussions

• RNAi- Decrease T-Cell surface protein expression.
• RNAi- Responsible for Silencing as well as
  activation.
• Decrease expression of CD4 and CD8 proteins.
• Specificity in silencing.
• Expression Varied with Time, Concentration
  (siRNA), and Effective sequences.
• RNAi- Results in increase in expression

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Therapeutic Uses

• TRANSPLANT REJECTION There is a decrease in the
  transplant rejection with the decrease in the
  expression of CD4 and CD8 proteins on the Surface
  of the T-Lymphocytes.
• RESISTANCE OT VIRAL INFECTION Indirect
  upregulation of interferons gives us a better
  chance of fighting off viral infection. As
  interferons and cytokines are the primary line of
  defense against viral infection.

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Authors

• Michael T. McManus, Brian B. Haines, Christopher
  P. Dillon, Charles E. Whitehurst, Luk van Parijs,
  Jianzhu Chen, and Phillip A. Sharp2
• Carol A. Sledz1,2,Michelle Holko1,3,Michael J. de
  Veer1, Robert H. Silverman1,2 and Bryan R.
  G.Williams14
• THANK YOU