Spliceosome disassembly catalyzed by Prp43 and its associated components Ntr1 and Ntr2.
Two novel yeast splicing factors required for spliceosome disassembly have been identified. Ntr1 and Ntr2 (NineTeen complex-Related proteins) were identified for their weak association with components of the Prp19-associated complex. Unlike other Prp19-associated components, these two proteins were primarily associated with the intron-containing spliceosome during the splicing reaction. Extracts depleted of Ntr1 or Ntr2 exhibited full splicing activity, but accumulated large amounts of lariat-intron in the spliceosome after splicing, indicating that the normal function of the Prp19-associated complex in spliceosome activation was not affected, but spliceosome disassembly was hindered. Immunoprecipitation analysis revealed that Ntr1 and Ntr2 formed a stable complex with DExD/H-box RNA helicase Prp43 in the splicing extract. Ntr1 interacted with Prp43 through the N-terminal G-patch domain, with Ntr2 through a middle region, and with itself through the carboxyl half of the protein. The affinity-purified Ntr1-Ntr2-Prp43 complex could catalyze disassembly of the spliceosome in an ATP-dependent manner, separating U2, U5, U6, NTC (NineTeen Complex), and lariat-intron. This is the first demonstration of physical disassembly of the spliceosome, catalyzed by a complex containing a DExD/H-box RNA helicase and two accessory factors, which might function in targeting the helicase to the correct substrate.
Tsai
Rong-Tzong
RT
Institute of Molecular Biology, Academia Sinica, Nankang, Taipei, Taiwan, Republic of China.
Fu
Ru-Huei
RH
Yeh
Fu-Lung
FL
Tseng
Chi-Kang
CK
Lin
Yu-Chieh
YC
Huang
Yu-Hsin
YH
Cheng
Soo-Chen
SC
D000255:Adenosine Triphosphate / Q000737:chemistry / Q000378:metabolism; D002474:Cell-Free System / Q000737:chemistry / Q000378:metabolism; D053487:DEAD-box RNA Helicases; D020365:RNA Helicases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D012326:RNA Splicing / Q000502:physiology*; D000072260:RNA Splicing Factors; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D017381:Spliceosomes / Q000737:chemistry / Q000378:metabolism*
C507653:Ntr2 protein, S cerevisiae; C079877:PRP19 protein, S cerevisiae; D000072260:RNA Splicing Factors; D029701:Saccharomyces cerevisiae Proteins; C507652:Spp382 protein, S cerevisiae; D000255:Adenosine Triphosphate; C109034:PRP43 protein, S cerevisiae; D053487:DEAD-box RNA Helicases; D020365:RNA Helicases