Extended Data Fig. 1: MS2-binding analysis using NLS-cpMCP-S52. | Nature Methods

Extended Data Fig. 1: MS2-binding analysis using NLS-cpMCP-S52.

From: RNA-stabilized coat proteins for sensitive and simultaneous imaging of distinct single mRNAs in live cells

Extended Data Fig. 1

(a) Schematic depicting the RNA binding-mediated nuclear export of NLS-cpMCP-S52, which is imported into and retained within the nucleus in its unbound form. Upon overexpression and nuclear export of MS2-tagged mRNA, protein-RNA binding leads to translocation of NLS-cpMCP-S52 to the cytoplasm. (b) Widefield fluorescence microscopy images of HEK293FT cells co-transfected to express mVenus-NLS-cpMCP-S52 and either H2B-mCherry-pA or H2B-mCherry-16xMS2-pA transcripts show that expression of MS2-tagged mRNA leads to nuclear exclusion of NLS-cpMCP-S52, indicating that it is capable of binding to MS2 loops. Insets in the bottom right display H2B-mCherry expression (c) HEK293FT cells were transfected to express mVenus-cpMCP-S52 (with no NLS) or the degron-appended mVenus-dMCP-S52 (both lacking an NLS). These cells were cotransfected to express H2B-mCherry-pA or H2B-mCherry-16xMS2-pA transcripts. The images show that the fluorescence of mVenus-dMCP-S52 is heavily reduced in the absence of MS2 RNA, and that the fluorescence is partially recovered in the presence of MS2 RNA.

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