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. 2022 May 23;11:e74410. doi: 10.7554/eLife.74410

Figure 3. Nonsense-mediated mRNA decay (NMD), Edc3, Dhh1, and Pat1 substrates are still degraded by decapping-dependent 5′ to 3′ decay in the absence of active decapping enzyme recruitment.

(A) Schematics of dcp2 alleles that eliminate Edc3, Upf1, or Pat1 binding to Dcp2. (B) Deletion of XRN1, but not deletions of SKI2 or SKI7, causes significant stabilization of NMD substrates in HA-dcp2-U1D1-U1D2 cells, and Edc3 and Dhh1 substrates in HA-dcp2-E3D or E3D1 cells. (C) Deletion of XRN1, but not deletions of SKI2 or SKI7, causes significant stabilization of Pat1 substrates in HA-dcp2-LD1-9 cells. Northern analyses in B and C as in Figures 1 and 2. Bar graphs in lower panels of B and C depict relative levels of decapping substrates in different strains determined from average ± SEM of three independent experiments. One representative northern blot for each transcript is shown in the upper panels. In the upper panel of B, lower case letters denote SCR1 blots duplicated for clarity of presentation. See also Figure 3—figure supplement 1.

Figure 3—source data 1. Northern analyses of different nonsense-mediated mRNA decay (NMD), Edc3, and Dhh1 substrates in cells harboring the HA-dcp2-U1D1-U1D2, E3D, or E3D1 alleles and deletions of XRN1, SKI2, or SKI7 (Figure 3B).
Figure 3—source data 2. Northern analyses of different Pat1 substrates in cells harboring the HA-dcp2-LD1-9 allele and deletions of XRN1, SKI2, or SKI7 (Figure 3C).

Figure 3.

Figure 3—figure supplement 1. Nonsense-mediated mRNA decay (NMD), Edc3, and Dhh1 substrates are still degraded by decapping-dependent 5′ to 3′ decay in the absence of active recruitment of the decapping enzyme.

Figure 3—figure supplement 1.

Bar graphs of average ± SEM for a subset of the northern analyses depicted in Figure 3B. The relative levels of each decapping substrate in different strains were determined from three independent experiments, with one representative blot for each transcript shown in Figure 3B. In each case, the relative RNA levels in different mutants were determined by comparison to the levels of the same transcripts in wild-type cells.