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Brr2 plays a role in spliceosomal activation in addition to U4/U6 unwinding.

Zhang L, Li X, Hill RC, Qiu Y, Zhang W, Hansen KC, Zhao R - Nucleic Acids Res. (2015)

Bottom Line: This N-terminal truncation does not significantly affect Brr2's helicase activity.However, the truncation significantly impairs spliceosomal activation, leading to a dramatic reduction of U5, U6 snRNAs and accumulation of U1 snRNA in the B(act) complex.The N-terminal domain of Brr2 does not seem to be directly involved in regulating U1/5'ss unwinding.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Genetics, University of Colorado School of Medicine, Aurora, CO 80045, USA.

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Brr2-Δ120 associates with less U5 snRNP, tri-snRNP, NTC, and U2 snRNP proteins in two independent pull down and mass spectrometry experiments (see Table 1 for raw data). Ratio of averaged integrated peak area (Brr2-Δ120/Brr2 WT) for each group of proteins is shown with this ratio for Brr2 normalized to 1.
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Figure 8: Brr2-Δ120 associates with less U5 snRNP, tri-snRNP, NTC, and U2 snRNP proteins in two independent pull down and mass spectrometry experiments (see Table 1 for raw data). Ratio of averaged integrated peak area (Brr2-Δ120/Brr2 WT) for each group of proteins is shown with this ratio for Brr2 normalized to 1.

Mentions: We next evaluated spliceosomal proteins associated with Brr2 WT and Brr2-Δ120 proteins using mass spectrometry after pulling down TAP-tagged Brr2 on IgG resin using the same amount of cells and resin (Table 1). The mass spectrometry data carried out in two independent experiments demonstrated again that TAP-Brr2 primarily pulls down U5 and tri-snRNPs under this experimental condition, but there are some NTC/Prp19 complex and U2 snRNP proteins present as well. Brr2 and Brr2-Δ120 protein levels are similar (Brr2-Δ120 protein level is slightly higher than WT Brr2 in experiment 2). Brr2-Δ120 associates with reduced levels of U5 and tri-snRNPs proteins (Figure 8), consistent with the reduced U4/U5/U6 snRNA associations (Figure 7b) and lower amount of tri-snRNP formed (Figure 4). Interestingly, multiple protein components of the U2 snRNP are more significantly reduced, judging by both the spectral counts and integrated peak areas (which typically provide a more accurate quantification of proteins) (Table 1).


Brr2 plays a role in spliceosomal activation in addition to U4/U6 unwinding.

Zhang L, Li X, Hill RC, Qiu Y, Zhang W, Hansen KC, Zhao R - Nucleic Acids Res. (2015)

Brr2-Δ120 associates with less U5 snRNP, tri-snRNP, NTC, and U2 snRNP proteins in two independent pull down and mass spectrometry experiments (see Table 1 for raw data). Ratio of averaged integrated peak area (Brr2-Δ120/Brr2 WT) for each group of proteins is shown with this ratio for Brr2 normalized to 1.
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Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4381053&req=5

Figure 8: Brr2-Δ120 associates with less U5 snRNP, tri-snRNP, NTC, and U2 snRNP proteins in two independent pull down and mass spectrometry experiments (see Table 1 for raw data). Ratio of averaged integrated peak area (Brr2-Δ120/Brr2 WT) for each group of proteins is shown with this ratio for Brr2 normalized to 1.
Mentions: We next evaluated spliceosomal proteins associated with Brr2 WT and Brr2-Δ120 proteins using mass spectrometry after pulling down TAP-tagged Brr2 on IgG resin using the same amount of cells and resin (Table 1). The mass spectrometry data carried out in two independent experiments demonstrated again that TAP-Brr2 primarily pulls down U5 and tri-snRNPs under this experimental condition, but there are some NTC/Prp19 complex and U2 snRNP proteins present as well. Brr2 and Brr2-Δ120 protein levels are similar (Brr2-Δ120 protein level is slightly higher than WT Brr2 in experiment 2). Brr2-Δ120 associates with reduced levels of U5 and tri-snRNPs proteins (Figure 8), consistent with the reduced U4/U5/U6 snRNA associations (Figure 7b) and lower amount of tri-snRNP formed (Figure 4). Interestingly, multiple protein components of the U2 snRNP are more significantly reduced, judging by both the spectral counts and integrated peak areas (which typically provide a more accurate quantification of proteins) (Table 1).

Bottom Line: This N-terminal truncation does not significantly affect Brr2's helicase activity.However, the truncation significantly impairs spliceosomal activation, leading to a dramatic reduction of U5, U6 snRNAs and accumulation of U1 snRNA in the B(act) complex.The N-terminal domain of Brr2 does not seem to be directly involved in regulating U1/5'ss unwinding.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Genetics, University of Colorado School of Medicine, Aurora, CO 80045, USA.

Show MeSH