Fig. 3: Spt5 stimulates Xrn2-dependent RNA degradation. | Nature Communications

Fig. 3: Spt5 stimulates Xrn2-dependent RNA degradation.

From: DSIF factor Spt5 coordinates transcription, maturation and exoribonucleolysis of RNA polymerase II transcripts

Fig. 3

a Spt5 can stimulate Xrn2 exoribonucleolytic activity towards 3’FAM-labelled 5’-monophosphate RNA (26 nt) in the context of the Pol II complex. Complexes were immobilised on beads using biotinylated non-template DNA (as depicted in Supplementary Fig. 4b). The extent of full-length RNA degradation is quantified for samples containing Xrn2 and indicated above RNA bands (n = 3). Source data are provided as a Source Data file. b Kow5-sCTR (Spt5 region 720 to 874 aa, N-terminal His-tag) stimulates degradation by Xrn2. Fluorescence polarisation anisotropy assay (as depicted in Supplementary Fig. 4d) comparing RNA degradation kinetics of Xrn2 alone or Kow5-sCTR (compare to Supplementary Fig. 4e). Construct used with N-terminal His-tag. Source data are provided as a Source Data file. c The affinity of Kow5-sCTR to RNA was evaluated using a polarisation anisotropy assay. The 3’FAM-labelled 5’-monophosphate-RNA substrate was titrated with increasing amounts of purified protein. Construct used with N-terminal His-tag. Source data are provided as a Source Data file.

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