Technology of the Month
In-Cell Discovery of RNA–Protein Interactions

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ADVANTAGES:

Does not rely on direct RNA pulldowns, thus circumventing a step that is often plagued by low efficiency.

Avoids crosslinking and the associated inefficiency and biases.

Not limited by low expression levels of the test RNA or proteins.

Can identify proteomes of specific RNA regions and provide RNA region-specific resolution of protein binding.

Can identify transient RNA–protein interactions.

One cell line is used for all applications.

The protein library is not limited in size and can be expanded or

CHALLENGES:

incPRINT is a primary binding assay that does not consider the developmental context or timing of RNA–protein interactions.

A subset of proteins may interact with MS2 stem-loops but not with the test RNA.

Protein tags may alter protein expression or binding.

Some interactions may be indirect: additional validation assays are necessary to establish direct RNA binding.

Some RNA–protein interactions may require additional factors that are not expressed in the cell line used for the assay.

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